Preclinical safety and efficacy of a therapeutic antibody that targets SARS-CoV-2 at the Sotrovimab face but is escaped by Omicron

The recurrent emerging of novel viral variants of concern (VOCs) with evasion of preexisting antibody immunity upholds SARS-CoV-2 case numbers and maintain a persistent demand for updated therapies. We selected the patient-derived antibody CV38-142 based on its potency and breadth against the VOCs Alpha, Beta, Gamma and Delta for preclinical development into a therapeutic. CV38-142 showed in vivo efficacy in a Syrian hamster VOC infection model after post-exposure and therapeutic application and revealed a favorable safety profile in a human protein library screen and tissue cross-reactivity study. Although CV38-142 targets the same viral surface as Sotrovimab which maintains activity against Omicron, CV38-142 did not neutralize the Omicron lineages BA.1 and BA.2. These results highlight the contingencies of developing antibody therapeutics in the context of antigenic drift and reinforce the need to develop broadly neutralizing variant-proof antibodies against SARS-CoV-2. Graphical

Preclinical safety and efficacy of a therapeutic antibody that targets SARS-CoV-2 at the sotrovimab face but is escaped by Omicron.

The recurrent emerging of novel viral variants of concern (VOCs) with evasion of preexisting antibody immunity upholds severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) case numbers and maintains a persistent demand for updated therapies. We selected the patient-derived antibody CV38-142 based on its potency and breadth against the VOCs Alpha, Beta, Gamma, and Delta for preclinical development into a therapeutic. CV38-142 showed in vivo efficacy in a Syrian hamster VOC infection model after post-exposure and therapeutic application and revealed a favorable safety profile in a human protein library screen and tissue cross-reactivity study. Although CV38-142 targets the same viral surface as sotrovimab, which maintains activity against Omicron, CV38-142 did not neutralize the Omicron lineages BA.1 and BA.2. These results highlight the contingencies of developing antibody therapeutics in the context of antigenic drift and reinforce the need to develop broadly neutralizing variant-proof antibodies against SARS-CoV-2.

Humoral immune responses remain quantitatively impaired but improve qualitatively in anti-CD20-treated patients with multiple sclerosis after three or four COVID-19 vaccinations.

OBJECTIVE: To analyze anti-SARS-CoV-2-S1-IgG levels, avidity, Omicron BA.2 variant neutralizing capacity, and SARS-CoV-2-specific T cells in anti-CD20-treated patients with multiple sclerosis (aCD20pwMS) after two, three, or four COVID-19 vaccinations. RESULTS: Frequencies of aCD20pwMS with detectable SARS-CoV-2-S1-IgG increased moderately between two (31/61 (51%)), three (31/57 (54%)), and four (17/26 (65%)) vaccinations. However, among patients with detectable SARS-CoV-2-S1-IgG, frequencies of high avidity (6/31 (19%) vs 11/17 (65%)) and Omicron neutralizing antibodies (0/10 (0%) vs 6/10 (60%)) increased strongly between two and four vaccinations. SARS-CoV-2-specific T cells were detectable in >92% after two or more vaccinations. CONCLUSION: Additional vaccinations qualitatively improve SARS-CoV-2 antibody responses.

Impaired B Cell Recall Memory and Reduced Antibody Avidity but Robust T Cell Response in CVID Patients After COVID-19 Vaccination.

PURPOSE: Humoral and cellular immune responses were described after COVID-19 vaccination in patients with common variable immunodeficiency disorder (CVID). This study aimed to investigate SARS-CoV-2-specific antibody quality and memory function of B cell immunity as well as T cell responses after COVID-19 vaccination in seroresponding and non-responding CVID patients. METHODS: We evaluated antibody avidity and applied a memory B cell ELSPOT assay for functional B cell recall memory response to SARS-CoV-2 after COVID-19 vaccination in CVID seroresponders. We comparatively analyzed SARS-CoV-2 spike reactive polyfunctional T cell response and reactive peripheral follicular T helper cells (pTFH) by flow cytometry in seroresponding and non-seroresponding CVID patients. All CVID patients had previously failed to mount a humoral response to pneumococcal conjugate vaccine. RESULTS: SARS-CoV-2 spike antibody avidity of seroresponding CVID patients was significantly lower than in healthy controls. Only 30% of seroresponding CVID patients showed a minimal memory B cell recall response in ELISPOT assay. One hundred percent of CVID seroresponders and 83% of non-seroresponders had a detectable polyfunctional T cell response. Induction of antigen-specific CD4+CD154+CD137+CXCR5+ pTFH cells by the COVID-19 vaccine was higher in CVID seroresponder than in non-seroresponder. Levels of pTFH did not correlate with antibody response or avidity. CONCLUSION: Reduced avidity and significantly impaired recall memory formation after COVID-19 vaccination in seroresponding CVID patients stress the importance of a more differentiated analysis of humoral immune response in CVID patients. Our observations challenge the clinical implications that follow the binary categorization into seroresponder and non-seroresponder.

Type of vaccine and immunosuppressive therapy but not diagnosis critically influence antibody response after COVID-19 vaccination in patients with rheumatic disease

Objective The development of sufficient COVID-19 vaccines has been a big breakthrough in fighting the global SARS-CoV-2 pandemic. However, vaccination effectiveness can be reduced in patients with autoimmune rheumatic diseases (AIRD). The aim of this study was to identify factors that lead to a diminished humoral vaccination response in patients with AIRD. Methods Vaccination response was measured with a surrogate virus neutralisation test and by testing for antibodies directed against the receptor-binding-domain (RBD) of SARS-CoV-2 in 308 fully vaccinated patients with AIRD. In addition, 296 immunocompetent participants were investigated as a control group. Statistical adjusted analysis included covariates with a possible influence on antibody response. Results Patients with AIRD showed lower antibody responses compared with immunocompetent individuals (median neutralising capacity 90.8% vs 96.5%, p<0.001;median anti-RBD-IgG 5.6 S/CO vs 6.7 S/CO, p<0.001). Lower antibody response was significantly influenced by type of immunosuppressive therapy, but not by rheumatic diagnosis, with patients under rituximab therapy developing the lowest antibody levels. Patients receiving mycophenolate, methotrexate or janus kinase inhibitors also showed reduced vaccination responses. Additional negative influencing factors were vaccination with AZD1222, old age and shorter intervals between the first two vaccinations. Conclusion Certain immunosuppressive therapies are associated with lower antibody responses after vaccination. Additional factors such as vaccine type, age and vaccination interval should be taken into account. We recommend antibody testing in at-risk patients with AIRD and emphasise the importance of booster vaccinations in these patients.

Preserved T cell responses to SARS-CoV-2 in anti-CD20 treated multiple sclerosis.

BACKGROUND: Optimal management of anti-CD20-treated patients with multiple sclerosis (pwMS) is an important clinical task during the current severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) pandemic. OBJECTIVES: To characterize humoral and cellular immune responses to SARS-CoV-2 vaccinations/infections in a longitudinal cohort of anti-CD20 treated (n = 175) and anti-CD20 therapy-naïve (n = 41) pwMS. METHODS: Anti-SARS-CoV-2 spike protein immunoglobulin G (IgG) and IgA, virus neutralizing capacity, IgG avidity and SARS-CoV-2-specific T cells were determined. RESULTS: Following two SARS-CoV-2 vaccinations, not only SARS-CoV-2 spike protein IgG and IgA, but also neutralizing capacity and avidity of SARS-CoV-2 IgG were lower in anti-CD20-treated (n = 51) than in anti-CD20 therapy-naïve pwMS (n = 14) and in healthy controls (HC, n = 19). However, in all anti-CD20-treated pwMS vaccinated twice (n = 26) or infected with SARS-CoV-2 (n = 2), in whom SARS-CoV-2-specific T cells were measured, SARS-CoV-2-specific T cells were detectable, at levels similar to those of twice-vaccinated anti-CD20 therapy-naïve pwMS (n = 7) and HC (n = 19). SARS-CoV-2-S1 IgG levels (r = 0.42, p = 0.002), antibody avidity (r = 0.7, p < 0.001), and neutralizing capacity (r = 0.44, p = 0.03) increased with time between anti-CD20 infusion and second vaccination. Based on detection of SARS-CoV-2 antibodies, SARS-CoV-2 infections occurred in 4 out of 175 (2.3%) anti-CD20-treated pwMS, all of whom recovered fully. CONCLUSIONS: These findings should inform treatment decisions and SARS-CoV-2 vaccination management in pwMS.

SARS-CoV-2 T Cell Response in Severe and Fatal COVID-19 in Primary Antibody Deficiency Patients Without Specific Humoral Immunity.

Morbidity and mortality of COVID-19 is increased in patients with inborn errors of immunity (IEI). Age and comorbidities and also impaired type I interferon immunity were identified as relevant risk factors. In patients with primary antibody deficiency (PAD) and lack of specific humoral immune response to SARS-CoV-2, clinical disease outcome is very heterogeneous. Despite extensive clinical reports, underlying immunological mechanisms are poorly characterized and levels of T cellular and innate immunity in severe cases remain to be determined. In the present study, we report clinical and immunological findings of 5 PAD patients with severe and fatal COVID-19 and undetectable specific humoral immune response to SARS-CoV-2. Reactive T cells to SARS-CoV-2 spike (S) and nucleocapsid (NCAP) peptide pools were analyzed comparatively by flow cytometry in PAD patients, convalescents and naïve healthy individuals. All examined PAD patients developed a robust T cell response. The presence of polyfunctional cytokine producing activated CD4+ T cells indicates a memory-like phenotype. An analysis of innate immune response revealed elevated CD169 (SIGLEC1) expression on monocytes, a surrogate marker for type I interferon response, and presence of type I interferon autoantibodies was excluded. SARS-CoV-2 RNA was detectable in peripheral blood in three severe COVID-19 patients with PAD. Viral clearance in blood was observed after treatment with COVID-19 convalescent plasma/monoclonal antibody administration. However, prolonged mucosal viral shedding was observed in all patients (median 67 days) with maximum duration of 127 days. PAD patients without specific humoral SARS-CoV-2 immunity may suffer from severe or fatal COVID-19 despite robust T cell and normal innate immune response. Intensified monitoring for long persistence of SARS-CoV-2 viral shedding and (prophylactic) convalescent plasma/specific IgG as beneficial treatment option in severe cases with RNAemia should be considered in seronegative PAD patients.

Pausing methotrexate improves immunogenicity of COVID-19 vaccination in elderly patients with rheumatic diseases.

OBJECTIVE: To study the effect of methotrexate (MTX) and its discontinuation on the humoral immune response after COVID-19 vaccination in patients with autoimmune rheumatic diseases (AIRD). METHODS: In this retrospective study, neutralising SARS-CoV-2 antibodies were measured after second vaccination in 64 patients with AIRD on MTX therapy, 31 of whom temporarily paused medication without a fixed regimen. The control group consisted of 21 patients with AIRD without immunosuppressive medication. RESULTS: Patients on MTX showed a significantly lower mean antibody response compared with patients with AIRD without immunosuppressive therapy (71.8% vs 92.4%, p<0.001). For patients taking MTX, age correlated negatively with immune response (r=-0.49; p<0.001). All nine patients with antibody levels below the cut-off were older than 60 years. Patients who held MTX during at least one vaccination showed significantly higher mean neutralising antibody levels after second vaccination, compared with patients who continued MTX therapy during both vaccinations (83.1% vs 61.2%, p=0.001). This effect was particularly pronounced in patients older than 60 years (80.8% vs 51.9%, p=0.001). The impact of the time period after vaccination was greater than of the time before vaccination with the critical cut-off being 10 days. CONCLUSION: MTX reduces the immunogenicity of SARS-CoV-2 vaccination in an age-dependent manner. Our data further suggest that holding MTX for at least 10 days after vaccination significantly improves the antibody response in patients over 60 years of age.

Cutting Edge: Serum but Not Mucosal Antibody Responses Are Associated with Pre-Existing SARS-CoV-2 Spike Cross-Reactive CD4+ T Cells following BNT162b2 Vaccination in the Elderly.

Advanced age is a main risk factor for severe COVID-19. However, low vaccination efficacy and accelerated waning immunity have been reported in this age group. To elucidate age-related differences in immunogenicity, we analyzed human cellular, serological, and salivary SARS-CoV-2 spike glycoprotein-specific immune responses to the BNT162b2 COVID-19 vaccine in old (69-92 y) and middle-aged (24-57 y) vaccinees compared with natural infection (COVID-19 convalescents, 21-55 y of age). Serological humoral responses to vaccination excee-ded those of convalescents, but salivary anti-spike subunit 1 (S1) IgA and neutralizing capacity were less durable in vaccinees. In old vaccinees, we observed that pre-existing spike-specific CD4+ T cells are associated with efficient induction of anti-S1 IgG and neutralizing capacity in serum but not saliva. Our results suggest pre-existing SARS-CoV-2 cross-reactive CD4+ T cells as a predictor of an efficient COVID-19 vaccine-induced humoral immune response in old individuals.

SARS-CoV-2 Beta variant infection elicits potent lineage-specific and cross-reactive antibodies.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) Beta variant of concern (VOC) resists neutralization by major classes of antibodies from COVID-19 patients and vaccinated individuals. In this study, serum of Beta-infected patients revealed reduced cross-neutralization of wild-type virus. From these patients, we isolated Beta-specific and cross-reactive receptor-binding domain (RBD) antibodies. The Beta-specificity results from recruitment of VOC-specific clonotypes and accommodation of mutations present in Beta and Omicron into a major antibody class that is normally sensitive to these mutations. The Beta-elicited cross-reactive antibodies share genetic and structural features with wild type-elicited antibodies, including a public VH1-58 clonotype that targets the RBD ridge. These findings advance our understanding of the antibody response to SARS-CoV-2 shaped by antigenic drift, with implications for design of next-generation vaccines and therapeutics.

Safety, reactogenicity, and immunogenicity of homologous and heterologous prime-boost immunisation with ChAdOx1 nCoV-19 and BNT162b2: a prospective cohort study.

BACKGROUND: Heterologous vaccine regimens have been widely discussed as a way to mitigate intermittent supply shortages and to improve immunogenicity and safety of COVID-19 vaccines. We aimed to assess the reactogenicity and immunogenicity of heterologous immunisations with ChAdOx1 nCov-19 (AstraZeneca, Cambridge, UK) and BNT162b2 (Pfizer-BioNtech, Mainz, Germany) compared with homologous BNT162b2 and ChAdOx1 nCov-19 immunisation. METHODS: This is an interim analysis of a prospective observational cohort study enrolling health-care workers in Berlin (Germany) who received either homologous ChAdOx1 nCov-19 or heterologous ChAdOx1 nCov-19-BNT162b2 vaccination with a 10-12-week vaccine interval or homologous BNT162b2 vaccination with a 3-week vaccine interval. We assessed reactogenicity after the first and second vaccination by use of electronic questionnaires on days 1, 3, 5, and 7. Immunogenicity was measured by the presence of SARS-CoV-2-specific antibodies (full spike-IgG, S1-IgG, and RBD-IgG), by an RBD-ACE2 binding inhibition assay (surrogate SARS-CoV-2 virus neutralisation test), a pseudovirus neutralisation assay against two variants of concerns (alpha [B.1.1.7] and beta [B.1.351]), and anti-S1-IgG avidity. T-cell reactivity was measured by IFN-γ release assay. FINDINGS: Between Dec 27, 2020, and June 14, 2021, 380 participants were enrolled in the study, with 174 receiving homologous BNT162b2 vaccination, 38 receiving homologous ChAdOx1 nCov-19 vaccination, and 104 receiving ChAdOx1 nCov-19-BNT162b2 vaccination. Systemic symptoms were reported by 103 (65%, 95% CI 57·1-71·8) of 159 recipients of homologous BNT162b2, 14 (39%, 24·8-55·1) of 36 recipients of homologous ChAdOx1 nCov-19, and 51 (49%, 39·6-58·5) of 104 recipients of ChAdOx1 nCov-19-BNT162b2 after the booster immunisation. Median anti-RBD IgG levels 3 weeks after boost immunisation were 5·4 signal to cutoff ratio (S/co; IQR 4·8-5·9) in recipients of homologous BNT162b2, 4·9 S/co (4·3-5·6) in recipients of homologous ChAdOx1 nCov-19, and 5·6 S/co (5·1-6·1) in recipients of ChAdOx1 nCov-19- BNT162b2. Geometric mean of 50% inhibitory dose against alpha and beta variants were highest in recipients of ChAdOx1 nCov-19-BNT162b2 (956·6, 95% CI 835·6-1095, against alpha and 417·1, 349·3-498·2, against beta) compared with those in recipients of homologous ChAdOx1 nCov-19 (212·5, 131·2-344·4, against alpha and 48·5, 28·4-82·8, against beta; both p<0·0001) or homologous BNT162b2 (369·2, 310·7-438·6, against alpha and 72·4, 60·5-86·5, against beta; both p<0·0001). SARS-CoV-2 S1 T-cell reactivity 3 weeks after boost immunisation was highest in recipients of ChAdOx1 nCov-19-BNT162b2 (median IFN-γ concentration 4762 mIU/mL, IQR 2723-8403) compared with that in recipients of homologous ChAdOx1 nCov-19 (1061 mIU/mL, 599-2274, p<0·0001) and homologous BNT162b2 (2026 mIU/mL, 1459-4621, p=0·0008) vaccination. INTERPRETATION: The heterologous ChAdOx1 nCov-19-BNT162b2 immunisation with 10-12-week interval, recommended in Germany, is well tolerated and improves immunogenicity compared with homologous ChAdOx1 nCov-19 vaccination with 10-12-week interval and BNT162b2 vaccination with 3-week interval. Heterologous prime-boost immunisation strategies for COVID-19 might be generally applicable. FUNDING: Forschungsnetzwerk der Universitätsmedizin zu COVID-19, the German Ministry of Education and Research, Zalando SE.

Cross-reactive CD4+ T cells enhance SARS-CoV-2 immune responses upon infection and vaccination.

The functional relevance of preexisting cross-immunity to severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is a subject of intense debate. Here, we show that human endemic coronavirus (HCoV)­reactive and SARS-CoV-2­cross-reactive CD4+ T cells are ubiquitous but decrease with age. We identified a universal immunodominant coronavirus-specific spike peptide (S816-830) and demonstrate that preexisting spike- and S816-830­reactive T cells were recruited into immune responses to SARS-CoV-2 infection and their frequency correlated with anti­SARS-CoV-2-S1-IgG antibodies. Spike­cross-reactive T cells were also activated after primary BNT162b2 COVID-19 messenger RNA vaccination and displayed kinetics similar to those of secondary immune responses. Our results highlight the functional contribution of preexisting spike­cross-reactive T cells in SARS-CoV-2 infection and vaccination. Cross-reactive immunity may account for the unexpectedly rapid induction of immunity after primary SARS-CoV-2 immunization and the high rate of asymptomatic or mild COVID-19 disease courses.

Reactive T Cells in Convalescent COVID-19 Patients With Negative SARS-CoV-2 Antibody Serology.

Despite RT-PCR confirmed COVID-19, specific antibodies to SARS-CoV-2 spike are undetectable in serum in approximately 10% of convalescent patients after mild disease course. This raises the question of induction and persistence of SARS-CoV-2-reactive T cells in these convalescent individuals. Using flow cytometry, we assessed specific SARS-CoV-2 and human endemic coronaviruses (HCoV-229E, -OC43) reactive T cells after stimulation with spike and nucleocapsid peptide pools and analyzed cytokine polyfunctionality (IFNγ, TNFα, and IL-2) in seropositive and seronegative convalescent COVID-19 patients as well as in unexposed healthy controls. Stimulation with SARS-CoV-2 spike and nucleocapsid (NCAP) as well as HCoV spike peptide pools elicited a similar T cell response in seropositive and seronegative post COVID-19 patients. Significantly higher frequencies of polyfunctional cytokine nucleocapsid reactive CD4+ T cells (triple positive for IFNγ, TNFα, and IL-2) were observed in both, seropositive (p = 0.008) and seronegative (p = 0.04), COVID-19 convalescent compared to healthy controls and were detectable up to day 162 post RT-PCR positivity in seronegative convalescents. Our data indicate an important role of NCAP-specific T cells for viral control.

Immunogenicity of COVID-19 Tozinameran Vaccination in Patients on Chronic Dialysis.

Patients with kidney failure have notoriously weak responses to common vaccines. Thus, immunogenicity of novel SARS-CoV-2 vaccines might be impaired in this group. To determine immunogenicity of SARS-CoV-2 vaccination in patients with chronic dialysis, we analyzed the humoral and T-cell response after two doses of mRNA vaccine Tozinameran (BNT162b2 BioNTech/Pfizer). This observational study included 43 patients on dialysis before vaccination with two doses of Tozinameran 21 days apart. Overall, 36 patients completed the observation period until three weeks after the second dose and 32 patients were further analyzed at week 10. Serum samples were analyzed by SARS-CoV-2 specific IgG and IgA antibodies ~1, ~3-4 and ~10 weeks after the second vaccination. In addition, SARS-CoV-2-specific T-cell responses were assessed at ~3-4 weeks by an interferon-gamma release assay (IGRA). Antibody and T cell outcomes at this timepoint were compared to a group of 44 elderly patients not on dialysis, after immunization with Tozinameran. Median age of patients on chronic dialysis was 74.0 years (IQR 66.0, 82.0). The proportion of males was higher (69.4%) than females. Only 20/36 patients (55.6%, 95%CI: 38.29-71.67) developed SARS-CoV-2-IgG antibodies at the first sampling, whereas 32/36 patients (88.9%, 95%CI: 73.00-96.38) demonstrated IgG detection at the second sampling. In a longitudinal follow-up at ~10 weeks after the second dose, the proportion of dialysis patients reactive for anti-SARS-CoV-2-IgG decreased to 27/32 (84.37%, 95%CI: 66.46-94.10) The proportion of anti-SARS-CoV-2 S1 IgA decreased from 33/36 (91.67%; 95%CI: 76.41-97.82) at weeks 3-4 down to 19/32 (59.38; 95%CI: 40.79-75.78). Compared to a cohort of vaccinees with similar age but not on chronic dialysis seroconversion rates and antibody titers were significantly lower. SARS-CoV-2-specific T-cell responses 3 weeks after second vaccination were detected in 21/31 vaccinated dialysis patients (67.7%, 95%CI: 48.53-82.68) compared to 42/44 (93.3%, 95%CI: 76.49-98.84) in controls of similar age. Patients on dialysis demonstrate a delayed, but robust immune response three to four weeks after the second dose, which indicates effective vaccination of this vulnerable group. However, the lower immunogenicity of Tozinameran in these patients needs further attention to develop potential countermeasures such as an additional booster vaccination.

Impact of dexamethasone on SARS-CoV-2 concentration kinetics and antibody response in hospitalized COVID-19 patients: results from a prospective observational study.

OBJECTIVES: Dexamethasone has become the standard of care for severe coronavirus disease 2019 (COVID-19), but its virological impact is poorly understood. The objectives of this work were to characterize the kinetics of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) concentration in the upper respiratory tract (URT) and the antibody response in patients with (D+) and without (D-) dexamethasone treatment. METHODS: Data and biosamples from hospitalized patients with severe COVID-19, enrolled between 4th March and 11th December 2020 in a prospective observational study, were analysed. SARS-CoV-2 virus concentration in serial URT samples was measured using RT-PCR. SARS-CoV-2-specific immunoglobulins A and G (IgA and IgG) were measured in serum samples using S1-ELISA. RESULTS: We compared 101 immunocompetent patients who received dexamethasone (according to the inclusion criteria and dosage determined in the RECOVERY trial) to 93 immunocompetent patients with comparable disease severity from the first months of the pandemic, who had not been treated with dexamethasone or other glucocorticoids. We found no inter-group differences in virus concentration kinetics, duration of presence of viral loads >106 viral copies/mL (D+ median 17 days (IQR 13-24), D- 19 days (IQR 13-29)), or time from symptom onset until seroconversion (IgA: D+ median 11.5 days (IQR 11-12), D- 14 days (IQR 11.5-15.75); IgG: D+ 13 days (IQR 12-14.5), D- 12 days (IQR 11-15)). CONCLUSION: Dexamethasone does not appear to lead to a change in virus clearance or a delay in antibody response in immunocompetent patients hospitalized with severe COVID-19.

Delayed Antibody and T-Cell Response to BNT162b2 Vaccination in the Elderly, Germany.

We detected delayed and reduced antibody and T-cell responses after BNT162b2 vaccination in 71 elderly persons (median age 81 years) compared with 123 healthcare workers (median age 34 years) in Germany. These data emphasize that nonpharmaceutical interventions for coronavirus disease remain crucial and that additional immunizations for the elderly might become necessary.

Outbreak of SARS-CoV-2 B.1.1.7 Lineage after Vaccination in Long-Term Care Facility, Germany, February-March 2021.

One week after second vaccinations were administered, an outbreak of B.1.1.7 lineage severe acute respiratory syndrome coronavirus 2 infections occurred in a long-term care facility in Berlin, Germany, affecting 16/20 vaccinated and 4/4 unvaccinated residents. Despite considerable viral loads, vaccinated residents experienced mild symptoms and faster time to negative test results.

Characterization of the SARS-CoV-2 Neutralization Potential of COVID-19-Convalescent Donors.

The current SARS-CoV-2 pandemic has triggered the development of various SARS-CoV-2 neutralization tests. A wild-type virus (using African green monkey VeroE6 cells), a pseudovirus (using human Caco-2 cells), and a surrogate neutralization test platform were applied to characterize the SARS-CoV-2 neutralization potential of a cohort of 111 convalescent plasma donors over a period of seven months after diagnosis. This allowed an in-depth validation and assay performance analysis of these platforms. More importantly, we found that SARS-CoV-2 neutralization titers were stable or even increased within the observation period, which contradicts earlier studies reporting a rapid waning of Ab titers after three to four months. Moreover, we observed a positive correlation of neutralization titers with increasing age, number of symptoms reported, and the presence of the Rhesus Ag RhD. Validation of the platforms revealed that highest assay performances were obtained with the wild-type virus and the surrogate neutralization platforms. However, our data also suggested that selection of cutoff titers had a strong impact on the evaluation of neutralization potency. When taking strong neutralization potency, as demonstrated by the wild-type virus platform as the gold standard, up to 55% of plasma products had low neutralization titers. However, a significant portion of these products were overrated in their potency when using the surrogate assay with the recommended cutoff titer. In summary, our study demonstrates that SARS-CoV-2 neutralization titers are stable for at least seven months after diagnosis and offers a testing strategy for rapid selection of high-titer convalescent plasma products in a biosafety level 1 environment.

Donors for SARS-CoV-2 Convalescent Plasma for a Controlled Clinical Trial: Donor Characteristics, Content and Time Course of SARS-CoV-2 Neutralizing Antibodies.

BACKGROUND: Convalescent plasma is one of the treatment options for COVID-19 which is currently being investigated in many clinical trials. Understanding of donor and product characteristics is important for optimization of convalescent plasma. METHODS: Patients who had recovered from CO-VID-19 were recruited as donors for COVID-19 convalescent plasma (CCP) for a randomized clinical trial of CCP for treatment of severe COVID-19 (CAPSID Trial). Titers of neutralizing antibodies were measured by a plaque-reduction neutralization test (PRNT). Correlation of antibody titers with host factors and evolution of neutralizing antibody titers over time in repeat donors were analysed. RESULTS: A series of 144 donors (41% females, 59% males; median age 40 years) underwent 319 plasmapheresis procedures providing a median collection volume of 850 mL and a mean number of 2.7 therapeutic units per plasmapheresis. The majority of donors had a mild or moderate course of COVID-19. The titers of neutralizing antibodies varied greatly between CCP donors (from <1:20 to >1:640). Donor factors (gender, age, ABO type, body weight) did not correlate significantly with the titer of neutralizing antibodies. We observed a significant positive correlation of neutralization titers with the number of reported COVID-19 symptoms and with the time from SARS-CoV-2 diagnosis to plasmapheresis. Neutralizing antibody levels were stable or increased over time in 58% of repeat CCP donors. Mean titers of neutralizing antibodies of first donation and last donation of repeat CCP donors did not differ significantly (1:86 at first compared to 1:87 at the last donation). There was a significant correlation of neutralizing antibodies measured by PRNT and anti-SARS-CoV-2 IgG and IgA antibodies which were measured by ELISA. CCP donations with an anti-SARS-CoV-2 IgG antibody content above the 25th percentile were substantially enriched for CCP donations with higher neutralizing antibody levels. CONCLUSION: We demonstrate the feasibility of collection of a large number of CCP products under a harmonized protocol for a randomized clinical trial. Titers of neutralizing antibodies were stable or increased over time in a subgroup of repeat donors. A history of higher number of COVID-19 symptoms and higher levels of anti-SARS-CoV-2 IgG and IgA antibodies in immunoassays can preselect donations with higher neutralizing capacity.