key: cord-0999349-dlp2hz6k
authors: Mair, Maximilian J.; Berger, Julia M.; Mitterer, Manfred; Gansterer, Margaretha; Bathke, Arne C.; Trutschnig, Wolfgang; Berghoff, Anna S.; Perkmann, Thomas; Haslacher, Helmuth; Lamm, Wolfgang W.; Raderer, Markus; Tobudic, Selma; Fuereder, Thorsten; Buratti, Thomas; Fong, Dominic; Preusser, Matthias
title: Third dose of SARS-CoV-2 vaccination in hemato-oncological patients and health care workers: immune responses and adverse events – a retrospective cohort study
date: 2022-02-07
journal: Eur J Cancer
DOI: 10.1016/j.ejca.2022.01.019
sha: 4d5b43deb0ad6b92c27e7c67349900e864fed739
doc_id: 999349
cord_uid: dlp2hz6k

BACKGROUND: Due to potentially immune-escaping virus variants and waning immunity, a third SARS-CoV-2 vaccination dose is increasingly recommended. However, data in patients with cancer are limited. PATIENTS AND METHODS: We measured anti-SARS-CoV-2 spike protein antibody levels after the third vaccination dose in 439 patients with cancer and 41 health care workers (HCW) at an academic center in Austria and a rural community hospital in Italy. Adverse events were retrieved from questionnaires. RESULTS: Overall, 439 patients and 41 HCW were included. SARS-CoV-2 infections were observed in 62/439 (14.1%) patients before vaccination and in 5/439 (1.1%) patients after ≥1 dose. Longitudinal analysis revealed a decrease of antibody levels between 3 and 6 months after second vaccination in patients with solid tumors (p<0.001) and hematological malignancies without anti-B cell therapies (p<0.001). After the third dose, anti-S levels increased as compared to the first/second dose. Patients receiving B cell-targeted agents had lower antibody levels than patients with hematological malignancies undergoing other treatments (p<0.001) or patients with solid tumors (p<0.001). Moreover, anti-S levels correlated with CD19+ (B cell) and CD56+ (NK cell) counts in peripheral blood. The most frequent adverse events after the third dose were local pain (75/160, 46.9%), fatigue (25/160, 15.6%), and fever/chills (16/160, 10.0%). Patients with cancer had lower anti-S levels than HCW (p=0.015). CONCLUSIONS: This study in patients with cancer shows improved antibody levels after the third vaccination dose at an acceptable side effect profile. Lower antibody levels than in controls underline the need for further follow-up studies and dedicated trials.

Patients with cancer are particularly affected by the COVID-19 pandemic. They are not only prone to adverse outcomes and severe COVID-19 when infected with SARS-CoV-2, but also at risk of disruptions of anticancer treatment or monitoring visits because of SARS-CoV-2 infections [1] [2] [3] [4] . Therefore, large oncological societies recommend SARS-CoV-2 vaccination for all patients with cancer [5] .

Overall, SARS-CoV-2 vaccines are highly effective and well tolerated [6] [7] [8] . Although SARS-CoV-2 vaccination shows immunogenicity in patients with cancer, seroconversion rates and antibody levels are lower compared to healthy cohorts, especially in patients undergoing antineoplastic treatment [9] [10] [11] [12] [13] [14] . Similar results were observed in patients with immunosuppression due to organ transplantation [15] .

With the emergence of variants of concern with increased immune evasion abilities, expanding the overall vaccination coverage and administration of a third booster vaccination is needed to avoid prolonging the pandemic [16] . A third vaccination dose shows the ability to lower the rates of SARS-CoV-2 infection and severe COVID-19 in comparison to non-boostered elderly populations [17] . Studies in immunosuppressed transplant recipients report improved antibody responses after the third dose [18, 19] .

Similarly, first data after the booster dose in cancer patients show higher antibody titers, especially in patients with minimal to moderate responses after the second dose [20] [21] [22] .

However, the existing data are limited by small sample sizes, and large-scale real-life data assessing the immune response after the third dose in patients with hematooncological diseases are scarce. Therefore, we aimed to assess clinical factors impacting humoral immune responses after the third SARS-CoV-2 vaccination dose in a large real-life cohort of patients with hemato-oncological malignancies.

In this retrospective cohort study, we included samples from two cohorts of patients with hemato-oncological malignancies undergoing antineoplastic treatment and/or regular follow-up visits as well as from one control group of health care workers (HCW) as described previously [9] . This study was reviewed and approved by the ethics prior to outpatient visits was performed as previously published and according to institutional practice [23] . Blood samples were stored by the "MedUni Wien Biobank" facility in accordance with Standard Operating Procedures in an ISO 9001:2015certified environment [24] . Patients were included if blood samples after the third SARS-CoV-2 vaccination dose were available. Sampling was performed at 15.5 days after vaccination in median (range: 1-70). Anti-SARS-CoV-2 nucleocapsid (anti-NC) and anti-SARS-CoV-2 spike protein (anti-S) antibodies were measured using Roche Elecsys assays at the Department of Laboratory Medicine as outlined previously and according to institutional practice [9] . Tappeiner" Hospital in Meran/Merano, Italy. All patients in this cohort received the BNT162b2 vaccine as first, second, and third dose. According to national guidelines, the second dose was considered as booster dose in patients with previous SARS-CoV-2 infection. Anti-NC/-S antibodies were measured 21 days after the first dose, while a further analysis was performed 21 days after the second dose in patients who did not show seroconversion (anti-S <50 AU/ml) after the first dose. Moreover, to assess the timely variations of antibody levels, repeated blood sampling at pre-defined timepoints (in median 3, 4.5 and 6 months after the second vaccination) was performed in a longitudinal cohort. Furthermore, lymphocyte subset counts were assessed 6 months after the second vaccination in this cohort. Antibody measurements after the third dose were performed at a median of 18 days (range: 3-66) after vaccination. Anti-NC and anti-S levels were determined using the Abbott SARS-CoV-2 IgG assays on the Abbott Alinity system as published previously [9] . Patient-reported adverse events after vaccination were collected using a standardized questionnaire.

Health care workers (HCW) cohort A cohort of HCW was included as control group. This cohort included physicians, nurses as well as scientific and administrative staff employed at the Division of Oncology, Department of Medicine I (Medical University of Vienna). Blood biobanking was performed as described previously [9, 25] , and antibody levels were measured as outlined for the Vienna patient cohort.

To compare distributions between metric variables, the Mann-Whitney-U, Kruskal-Wallis, and Wilcoxon signed-rank tests were used as appropriate, supplemented by nonparametric methods for longitudinal data [26] . A two-sided p<.05 was considered as significance threshold. Due to the exploratory and hypothesis-generating design of the present study, no adjustment for multiple testing was applied [27] . Statistical analysis was performed using GraphPad Prism 9. 

In total, 26 patients were included in Vienna, while 413 patients were enrolled in Meran.

In the Vienna cohort, the median age was 63 years (range: 28-85) and all patients were treated for solid tumors. The most frequently applied treatments were chemotherapy and immune checkpoint inhibitors (ICI) in 8/26 (30.8%) patients each.

In the Meran cohort, the median age was 70 years (range: Table 1 .

In Vienna, 19 infection between the first and second dose had a mild clinical course and could be managed in an outpatient setting. In total, three patients were infected after the third dose (3 days, 3 weeks, and 4 weeks after vaccine administration, respectively). One patient suffered from acute myeloid leukemia and received 5-azacytidine treatment, J o u r n a l P r e -p r o o f another individual was under active ibrutinib treatment for CLL. Both were admitted to the hospital but did not need admission to the intensive care unit (ICU). In the third patient, rituximab maintenance therapy due to follicular lymphoma had been stopped 6 months prior to infection. Still, the patient had no detectable SARS-CoV-2 antibodies after vaccination and experienced a severe course with ICU admission.

In the Vienna patient cohort, the median anti-S level after the second vaccination was 236 U/ml (range: 0.0-2500 U/ml; below the detection range (0.4 U/ml): 2/24 patients, 8 .3%). After the third vaccination, anti-S concentrations of these patients increased to a median of 2500 U/ml (range: 2-2500 U/ml; <0.4 U/ml: 0/26 patients, 0.0%; p<.001, Wilcoxon signed-rank test, Figure 1A ).

In the Meran cohort, median anti-S levels after the first vaccination were 222.8 AU/ml tumors. In the latter, no difference according to applied treatments could be detected overall (p=.199, Kruskal Wallis); still, in uncorrected pairwise comparisons, patients receiving chemotherapy had somewhat lower levels (median 13383 AU/ml, range: 32-40000, <50 AU/ml: 1/98, 1.0%) than those with no active antineoplastic treatment (median: 34288 AU/ml, range: 7112-40000, <50 AU/ml: 0/17, 0.0%, p=.022, Figure   1D ). There were no differences in time between third vaccination and blood sampling between these subgroups (pairwise p>.999, Mann-Whitney-U). previously [9] . In these patients, the third vaccination dose did not result in higher antibody levels (median: 3.6 AU/ml, range: 0-40000, p=.325). In contrast, antibody levels increased in solid tumors (median: 23074 AU/ml, range: 7.2-40000, p<.001) and

hematological patients without B cell-targeted therapy (median: 11261 AU/ml, range: 1.7-39998) after the third vaccination. In both patient cohorts with and without prior SARS-CoV-2 infection, antibody levels decreased over time and increased after the third shot. In addition, patients with prior COVID-19 overall reached higher antibody levels after the third dose (median: 37620.9 AU/ml; range: 1969.6-40000; <50 AU/ml: 0/58, 0.0%) than patients with no documented COVID-19 (median: 7426.95, range: 0-40000; <50 AU/ml: 24/120, 20.0%, Figure 2B ). There was no difference in time between third vaccination and blood sampling between patients with and without prior COVID-19 (p=.494, Mann-Whitney-U).

In 149/178 (83.7%) patients of the longitudinal cohort, lymphocyte subtype analyses were performed 6 months after the second vaccination dose. Of these patients, 56/149 (37.6%) had previously been infected with SARS-CoV-2. These individuals had higher total lymphocyte counts (p = 0.031, Mann-Whitney-U) and CD19+ counts (p < 0.001) as well as numerically higher CD56+ cell counts (p = 0.051) than their counterparts who had not previously been tested positive for SARS-CoV-2 ( Figure 4) . Overall, antibody levels 6 months after second vaccination weakly correlated with CD19+ counts (Spearman's r = 0.325, p < 0.001) and CD56+ counts (Spearman's r = 0.220, p = 0.008), as did antibody levels after the third dose (CD19+: r = 0.288, p < 0.001; CD56+: r = 0.250, p = 0.002). This association remained significant in patients who were not infected, while there was no correlation in previously infected patients (detailed correlation plots are given in Supplementary Figure 1 ).

Possibly vaccination-associated adverse events (AE) were reported in 413 patients of the Meran cohort after the first dose as well as in 361/413 (87.4%) patients after the second and 160/461 (34.7%) patients after the third dose. Higher-grade adverse events were overall rare, with only 9 grade 3 AE after the first, 6 Table 2 . [9] [10] [11] [12] [13] [14] . However, with the emergence of novel SARS-CoV-2 variants such as the recent B.1.1.529 ("Omikron") variant, efficacy of the available vaccines may be further impaired [28, 29] . Early reports suggest that three doses of the BNT162b2 vaccine result in sufficient amounts of antibodies neutralizing the B.1.1.529 variant, whereas levels of these antibodies are markedly lower in individuals who had only received two doses [29, 30] . These results underscore the importance of a third "booster" vaccination dose. However, respective data after the third dose are limited in immunosuppressed patients such as those undergoing cancer treatment.

Here, we show in one of the largest published cohorts so far that anti-S levels increased after the third vaccination in patients with cancer. These results are in line with recent publications of smaller cohorts, where a meaningfully strengthened humoral immune response after the third vaccination dose was suggested [20] [21] [22] 31, 32] . However, and in accordance with other published data, patients with hemato-oncological diseases still show lower antibody levels as compared to controls [20] . In fact, antibody concentrations after the third dose in our cancer patient cohort were comparable to those after the second vaccination in the HCW control group.

Further follow-up studies will be needed to evaluate whether the administration of additional vaccination doses will be necessary in the further course of the pandemic, especially in immunocompromised patients at high risk for severe clinical courses of COVID-19.

Once again, hematological patients receiving anti-B cell therapies showed impaired humoral immune responses compared to patients undergoing other treatment modalities. This is particularly concerning, as one patient in our study experienced a J o u r n a l P r e -p r o o f breakthrough infection with severe course and ICU admission 6 months after prior rituximab treatment for follicular lymphoma. Of note, no anti-S antibodies were observed in this patient after the third vaccination. Still, cellular immunity may be preserved in many of these patients receiving anti-B cell agents. Reassuringly, previous reports have shown that T cell priming and reactivity are still preserved in patients with rheumatic diseases or multiple sclerosis [33, 34] . In a recent publication, the correlation between neutralizing antibodies and spike protein-specific T cell responses was weak in patients with cancer, indicating that cellular immunity against SARS-CoV-2 may mount independently from humoral immune responses [20] . In addition, a correlation between cellular immunity and disease severity was postulated in individuals who recovered from COVID-19 [35] . These results suggest that patients under anti-CD20 treatment may still be protected to some extent from severe courses of COVID-19, even if SARS-CoV-2 infection per se cannot be prevented in the absence of neutralizing antibodies. Moreover, although our data and a previously published report [36] showed that antibody levels diminished in the months after the second dose in patients with cancer, data from other vaccinations imply that vaccination-specific memory T cells may persist for decades after immunization [37] . Along these lines, a long-acting monoclonal antibody was recently approved for use as pre-exposure prophylaxis in patients with compromised immune systems due to immunosuppressive medications who are not able to mount an adequate immune response to COVID-19 vaccination [38] . Further monoclonal antibodies for post-exposure prophylaxis and early outpatient treatment in patients with risk factors have been studied [39, 40] .

Notably, we found correlations between antibody levels and lymphocyte subset counts, specifically CD19+ B cells and CD56+ natural killer (NK) cells. In line, patients with prior SARS-CoV-2 infection (and subsequently higher antibody levels) had higher B cell and NK cell counts. Still, the correlation of CD19+ and CD56+ cell counts with J o u r n a l P r e -p r o o f antibody levels remained also significant in patients who had not been infected. These data confirm previous findings where a link between CD56+ NK cell counts and immune responses towards mRNA-based vaccines has been postulated [41, 42] .

However, the underlying immunological mechanisms remain elusive, and further investigations including antigen-specific T cell assays would be needed to obtain deeper insights into vaccination-induced immunity in patients with cancer.

In our cohorts, 14% of the included patients were infected before vaccination, while breakthrough infections were seen in only ~1%. Although the inferences of our retrospective observations on vaccine efficacy have to be taken cautiously, these data underline that the combination of SARS-CoV-2 vaccination with general protective measures (including personal, institutional and governmental safety measures [23,43]) helps to limit uncontrolled viral spread in patients with cancer. To add further, AE rates after vaccine administration in our cohort were comparable to those observed in the general population [7] . Overall, our data corroborate the favorable safety profile of the available vaccines in patients with hemato-oncological diseases, confirming the beneficial risk-benefit ratio of SARS-CoV-2 vaccination.

Clearly, our study has important limitations. As in our previous report [9] , pooled analyses between both patient cohorts were not feasible due to different assays used in both centers. Again, only patients who were seronegative after the first dose underwent antibody measurements after the second dose in the Meran cohort. 

1 (0.2%) 0 (0.0%) 0 (0.0%) 2 (0.6%) 0 (0.0%) 0 (0.0%) 0 (0.0%) 0 (0.0%) 0 (0.0%) sore throat 3 (0.7%) 0 (0.0%) 0 (0.0%) 2 (0.6%) 0 (0.0%) 0 (0.0%) 1 (0.6%) 0 (0.0%) 0 (0.0%) headache 21 3 (0.7%) 1 (0.2%) 0 (0.0%) 4 (1.1%) 0 (0.0%) 0 (0.0%) 2 (1.3%) 0 (0.0%) 0 (0.0%) decreased appetite 9 (2.2%) 0 (0.0%) 0 (0.0%) 10 (2.8%) 0 (0.0%) 0 (0.0%) 4 (2.5%) 0 (0.0%) 0 (0.0%) nausea 6 (1.4%) 2 (0.5%) 0 (0.0%) 5 (1.4%) 2 (0.5%) 0 (0.0%) 3 (1.9%) 0 (0.0%) 0 (0.0%) diarrhea 3 (0.7%) 0 (0.0%) 1 (0.2%) 3 (0.8%) 0 (0.0%) 1 (0.3%) 4 (2.5%) 0 (0.0%) 1 (0.6%) cough 1 (0.2%) 0 (0.0%) 0 (0.0%) 2 (0.6%) 0 (0.0%) 0 (0.0%) 2 (1.3%) 0 (0.0%) 0 (0.0%) dyspnea 1 (0.2%) 0 (0.0%) 0 (0.0%) 2 (0.6%) 0 (0.0%) 0 (0.0%) 1 (0.6%) 0 (0.0%) 0 (0.0%) chest pain 3 (0.7%) 0 (0.0%) 1 (0.24%) 2 (0.6%) 0 (0.0%) 0 (0.0%) 1 (0.6%) 1 (0.6%) 0 (0.0%) tachycardia 5 (1.2%) 1 (0.2%) 0 (0.0%) 2 (0.6%) 0 (0.0%) 0 (0.0%) 0 (0.0%) 0 (0.0%) 0 (0.0%) anosmia 1 (0.2%) 0 (0.0%) 0 (0.0%) 0 (0.0%) 0 (0.0%) 0 (0.0%) 0 (0.0%) 0 (0.0%) 0 (0.0%) hypesthesia 2 (0.5%) 0 (0.0%) 0 (0.0%) 2 (0.6%) 0 (0.0%) 0 (0.0%) 1 (0.6%) 0 (0.0%) 0 (0.0%)

Supplementary 

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SARS-CoV-2 vaccines for cancer patients: a call to action

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Safety and Efficacy of the BNT162b2 mRNA Covid-19 Vaccine

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Efficacy of the BNT162b2 mRNA COVID-19 vaccine in patients with chronic lymphocytic leukemia

Seroconversion rate after vaccination against COVID-19 in patients with cancer-a systematic review

Immunogenicity and Safety of COVID-19 Vaccine BNT162b2 for Patients with Solid Cancer: A Large Cohort Prospective Study from a Single Institution

Immunogenicity and risk of Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) infection after Coronavirus Disease 2019 (COVID-19) vaccination in patients with cancer: a systematic review and meta-analysis

Emerging issues related to COVID-19 vaccination in patients with cancer

Antibody Response to 2-Dose SARS-CoV-2 mRNA Vaccine Series in Solid Organ Transplant Recipients

Vaccine third dose and cancer patients: necessity or luxury?

Protection of BNT162b2 Vaccine Booster against Covid-19 in Israel

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Immune responses to two and three doses of the BNT162b2 mRNA vaccine in adults with solid tumors

mRNA Vaccine in Patients With Solid Tumors Undergoing Active Treatment

SARS-CoV-2 Antibody Response to 2 or 3 Doses of the BNT162b2 Vaccine in Patients Treated With Anticancer Agents

SARS-CoV-2 Testing in Patients With Cancer Treated at a Tertiary Care Hospital During the COVID-19 Pandemic

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release: Pfizer and BioNTech provide update on Omicron variant

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Efficacy of booster doses in augmenting waning immune responses to COVID-19 vaccine in patients with cancer

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Cellular and humoral immune responses following SARS-CoV-2 mRNA vaccination in patients with multiple sclerosis on anti-CD20 therapy

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