key: cord-278112-uqxvr77g authors: Chen, Binzhen; Xia, Rong title: Early experience with convalescent plasma as immunotherapy for COVID‐19 in China: Knowns and unknowns date: 2020-06-09 journal: Vox Sang DOI: 10.1111/vox.12968 sha: doc_id: 278112 cord_uid: uqxvr77g BACKGROUND AND OBJECTIVES: In the absence of a vaccine or specific drug treatment options for coronavirus disease (COVID‐19), attention has been shifted in China to the possible therapeutic use of convalescent plasma. COVID‐19 convalescent plasma (CCP) is currently under investigation. We summarized clinical studies and other research data available as of May 5, 2020, on CCP therapy according to the Clinical Treatment Guideline of COVID‐19 Convalescent Plasma in China, as well as clinical experience at the First Affiliated Hospital of Zhejiang University, as part of a comprehensive anti‐epidemic strategy. MATERIALS AND METHODS: As of May 5, 2020, when the epidemic was well‐controlled in China, healthcare databases and sources of English literature relating to convalescent plasma were searched and reviewed. Sources of clinical and methodological heterogeneity were identified. RESULTS: As of May 5, 2020, up to 2,000 samples of CCP had been collected across China and administered to 700 COVID‐19 patients. From donors, 200–400 mL of plasma were collected at each donation, with antibody titers >1:160. We identified three clinical studies for COVID‐19 in China. Analyses showed a statistically significant improvement in clinical outcomes compared with untreated cases (P<0.001). No adverse effects were reported. CONCLUSION: From initial studies, convalescent plasma therapy appears effective and safe for COVID‐19. However, there is clearly a need for well‐designed RCTs (randomized controlled trials) or other formal studies to further evaluate the efficacy and any potential adverse effects of CCP. This article is protected by copyright. All rights reserved The outbreak of novel coronavirus disease at the end of 2019 was caused by a novel coronavirus, SARS-CoV-2, that is highly contagious [1] and resulted in a crude mortality rate of about 2.3% for all cases [2] and 25.5% for severe cases [3] . In the absence of a vaccine for SARS-CoV-2, COVID-19 convalescent plasma (CCP) represents a form of passive immunotherapy. Although current clinical experience with CCP is encouraging, well-designed control studies are lacking. Convalescent plasma has long been used as a therapeutic tool in blood transfusion and this procedure involves extracting plasma containing anti-viral antibodies from patients who have recovered, and then injecting this convalescent plasma into other patients to treat diseases. It has become an emergency or empirical therapy, and has been successfully applied in the field of cell therapy and immunotherapy [4] . The application of convalescent plasma in the treatment of Ebola [5, 6], SARS [7] [8] [9] , pandemic influenza [10] [11] [12] and MERS [13, 14] , has suggested that CCP may be an effective, timely and widely available treatment option in the case of COVID-19. This article discusses the application of convalescent plasma as an immunotherapy in the treatment of emerging infectious diseases, focusing on the recent COVID-19 outbreak in China. The PubMed, Embase and Medline databases were searched from December 8, 2019 (first breakout) to May 5, 2020, for newly published literature using the search term "convalescent plasma AND COVID-19". All search records were imported into EndNote X9 software and screened manually using paper records. This article is protected by copyright. All rights reserved Publications were selected that involved the assessment of CCP treatment for COVID-19 patients in China including clinical trials such as randomized controlled trials, controlled clinical trials, case series, case reports, prospective and retrospective comparative cohort studies, and case-control studies. Clinical information for the 19 [15, 16] . Adverse events and serious adverse events were assessed as part of the CCP safety evaluation. Improvements in clinical outcomes were defined as symptomatic relief, oxygen saturation normalization, a decrease in lymphocytes and C-reactive protein levels, and decreased mortality rates. Continuous variables (i.e., age, laboratory test parameters) were presented as median (IQR) values and were compared using a Mann-Whitney U test. Categorical variables were expressed as n (%), where n was the total number of This article is protected by copyright. All rights reserved patients with available data, and were compared by Fisher's exact test between the convalescent plasma treatment group and the control group. Statistical software used was SPSS 24.0. For all analyses, P<0.05 was considered statistically significant. All data relevant to this manuscript and available to the authors at the time of publication are included in the main text or Supplementary file S1.  The treatment of COVID-19 in China has focused on preventing patients with mild disease from progressing to severe or critical illness. Therapeutic agents including steroids, antivirals and traditional Chinese medicines that have been considered are summarized in Table I [17] [18] [19] [20] [21] [22] [23] . Indications, directions and clinical trials located in China up until May 5, 2020, are listed for each agent respectively. For severely and critically ill individuals, besides the agents above, patients have been treated with CCP, tocilizumab, stem cells and artificial liver therapy. A recent publication reporting a trial of lopinavir-ritonavir showed no clinical benefits beyond standard care in adults hospitalized with severe COVID-19 [24] . While most of the agents listed above remain under investigation (status: recruiting, suspended, terminated or not yet recruiting), none have been proven to be specific for COVID-19. Thus, CCP may be a promising immunotherapy for COVID-19, especially among severely and critically ill patients. This article is protected by copyright. All rights reserved The first dose of convalescent plasma from a COVID-19 patient was collected on February 1, 2020, and the first severely ill patient received CCP treatment at a hospital in Wuhan, the capital of Hubei Province that was first to be hit by the virus, on February 9, 2020. China has established expert task forces at the provincial and national levels to analyze and improve the use of plasma therapy, and the National Health Commission (NHC) has worked with other authorities to motivate more recovered patients to donate plasma. According to a press conference by the Joint Prevention and Control Mechanism of the State Council on May 5, 2020, as many as 2,000 doses of CCP had been collected across China and 700 COVID-19 patients had received this therapy. Patients had shown improvements in clinical indicators and symptoms, and the median length of hospital stays in ICU for patients receiving CCP was also significantly shorter than for those in the control group. Recent studies have reported that plasma therapy is safe and effective [15, 16, 25] and the details from these three studies (one pilot study, one case series and one case report) are presented in Table II ( Donors are eligible for CCP donation at least 2 weeks after recovery and discharge from hospital. Donors should be between 18 and 55 years of age, with a body weight of ≥50 kg (male) or ≥45 kg (female). Donors should present at least 14 days after their last blood donation and must qualify as donors after a comprehensive clinical assessment. Plasma is collected by plasmapheresis (200-400 mL) at each donation (based on medical consultation). In addition to routine tests for transfusion-transmitted diseases, CCP samples are subjected to: (1) nucleic acid testing for COVID-19; (2) a qualitative test of COVID-19-specific IgG and IgM detection (involving a 160-fold dilution) or a qualitative test for whole antibody detection (involving a 320-fold dilution). If possible, a test for viral neutralizing antibodies is also performed. This article is protected by copyright. All rights reserved CCP is being used for severe or critically ill COVID-19 patients or those with rapidly progressive disease who test positive in a virus nucleic acid test. As a general rule, CCP is not used on COVID-19 patients whose disease course exceeds three weeks. Specific contraindications include: (1) a history of allergy to plasma or methylene blue; (2) a history of autoimmune system disease or selective IgA deficiency, in which case, the application of CCP should be evaluated cautiously by clinicians. The dose of CCP is based on a patient's weight and clinical condition. In general, 200-500 mL (4-5 mL/kg) is recommended. It is recommended to transfuse recipients with plasma obtained from donors on the day of collection (fresh collection) to maximize plasma activity [15] . In the absence of specific drugs and vaccines, convalescent plasma therapy is an alternative treatment option. However, there are a number of issues to be considered when determining the suitability of a large-scale convalescent plasma infusion strategy. The neutralizing antibodies present in CCP do not appear to influence the cytokine storm that can result from COVID-19 infection. A cytokine storm is an important pathophysiological event in the transformation of COVID-19 from a mild to severe disease and from single organ damage to multiple organ dysfunction syndrome (MODS) [28] . It has been reported that SARS-CoV-2 can induce the excessive release of inflammatory cytokines, causing diffuse alveolar injury, hyaline membrane formation and the exudation of proteins into This article is protected by copyright. All rights reserved the lungs. In severe cases, shock, disseminated intravascular coagulation and MODS may occur [29] . The levels of IL-6 in patients with severe COVID-19 were found to have increased significantly [30] , which may be related to the poor prognosis of patients and suggests the need for alternative treatments [31] . From this perspective, it is not expected that neutralizing antibodies in the CCP will be able to inhibit an inflammatory storm. In China, as of May 5, 2020, at least 95 clinical trials were registered at www.clinicaltrials.gov [32] to investigate potential treatments for the novel coronavirus, and the number of registered trials is increasing every day. Studies are testing both new and traditional medicines. However, at present, the trials lack a unified framework of standards, are disordered, and have weaknesses in terms of the levels of evidence, ethical standards, and the efficient use of research resources. Similarly, studies on CCP in China have lacked appropriate control groups and display a moderate to high risk of bias, making it difficult to provide evidence that any clinical improvement in patients is due to CCP, to other therapies, or to the natural process of recovery. A large randomized, double-blind, controlled study is therefore needed for CCP therapy before it can be approved as a routine clinical treatment. The rationale for CCP as a therapeutic tool is to provide higher levels of neutralizing antibodies against the virus [33] . It is anticipated that the higher the titer of neutralizing antibody, the better the therapeutic effect [34] . In China, the Accepted Article difficulty in obtaining high titer CCP has limited its clinical application. Immune dysfunction may also hinder the efficacy of CCP therapy [33] . In addition, there is a great disparity between plasma demand and supply, as well as the limitations imposed by the need to obtain plasma during the eligible recovery period. Regional imbalances in plasma resources also exist in developed and developing countries. Alternative strategies should be explored to identify CCP with sufficient antibody titers, including serum collection from patients with more severe disease and collection at earlier time points during disease. Regarding the antibodies present in CCP, antibody-dependent enhancement is another potential limitation [35] . One previous study in China by Duan and colleagues [16] reported a minor side effect of an evanescent, red facial spot in one patient administered convalescent plasma. This minor side effect was not linked to any adverse events, and two other trials showed no side effects. No complications such as TRALI or TACO, or any other adverse events, were reported. However, appropriate risk assessment is still required for high-risk donors before considering relaxing the inclusion criteria. The effective treatment of disease following infection with SARS-CoV-2 presents an unprecedented challenge in China and worldwide. Studies to date suggested that CCP therapy, performed according to Chinese recommendations, was a promising and safe treatment option with no adverse events. However, there are several points to consider to aid the future success of CCP therapy. This article is protected by copyright. All rights reserved  Attention should be paid to the evaluation of both RDB (reverse dot blot) antibody and neutralization antibody levels in plasma. Convalescent plasma must be collected at the optimum time to ensure a high titer of neutralizing antibodies.  Large-scale screening is needed for the inclusion of a large number of COVID-19 patients.  Pathogen reduction and frozen stockpile technology should be applied to ensure safe infusion and to avoid transfusion-related adverse events.  Further evaluation of the efficacy and safety of convalescent plasma in patients should be carried out in carefully designed RCTs.  Another new methodology is to select B cells that can produce specific antibodies in convalescent blood, obtain the gene sequence of the antibody at the single-cell level, reconstruct and express it using specialized technology, and finally obtain a large number of B cells that can produce virus neutralizing antibodies. These antibodies can then be injected into patients [36] .  Recently, Zhang and colleagues isolated high efficiency anti-coronavirus antibodies from COVID-19 patients, which laid the foundation for the further development of antibodies [37] . In the long run, the immunotherapy of COVID-19 still depends on protective virus-specific monoclonal antibodies.  Blood centers and healthcare agencies should cooperate in calling upon the public to donate. Along with its research efforts into CCP therapy, China is strengthening communication with the World Health Organization, and cooperating with other countries, especially high-risk countries, to promote scientific research, drug and vaccine development, and the sharing of scientific information to address the global epidemic. 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All rights reserved descriptive study Correlation analysis between disease severity and inflammation-related parameters in patients with COVID-19 pneumonia National Library of Medicine: Studies found for: COVID-19 in China Convalescent plasma therapy for persistent hepatitis E virus infection Anti-Ebola virus antibody levels in convalescent plasma and viral load after plasma infusion in patients with Ebola virus disease Convalescent plasma in Covid-19: Possible mechanisms of action Efficient generation of monoclonal antibodies from single human B cells by single cell RT-PCR and expression vector cloning Human neutralizing antibodies elicited by SARS-CoV-2 infection The author thanks Prof. Xia for his assistance in guiding and proofreading for this manuscript and all people who work so hard to fight against COVID-19. This article is protected by copyright. All rights reserved The authors report no conflict of interest. NOTE Continuous variable (age) is expressed as median (IQR) and compared with Mann-Whitney U test. Categorical data are n (%) of patients, where n is the total number of patients with available data, and compared by Fisher's exact test between convalescent plasma treatment group and control group. SaO 2 , oxyhemoglobin saturation. Reference ranges are as follows: C-reactive protein, normal range＜8 mg/L; lymphocyte count, 1.2-3.4 ×10 9 /L; SaO 2 %, normal range≥95%. a Among the 19 patients in CP treatment group, nine patients did not have their blood lymphocyte and SaO 2 data available.