Molecular basis of SARS-CoV-2 Omicron variant evasion from shared neutralizing antibody response.

Understanding the molecular features of neutralizing epitopes is important for developing vaccines/therapeutics against emerging SARS-CoV-2 variants. We describe three monoclonal antibodies (mAbs) generated from COVID-19 recovered individuals during the first wave of the pandemic in India. These mAbs had publicly shared near germline gene usage and potently neutralized Alpha and Delta, poorly neutralized Beta, and failed to neutralize Omicron BA.1 SARS-CoV-2 variants. Structural analysis of these mAbs in complex with trimeric spike protein showed that all three mAbs bivalently bind spike with two mAbs targeting class 1 and one targeting a class 4 receptor binding domain epitope. The immunogenetic makeup, structure, and function of these mAbs revealed specific molecular interactions associated with the potent multi-variant binding/neutralization efficacy. This knowledge shows how mutational combinations can affect the binding or neutralization of an antibody, which in turn relates to the efficacy of immune responses to emerging SARS-CoV-2 escape variants.

Vaccine models predict rules for updating vaccines against evolving pathogens such as SARS-CoV-2 and influenza in the context of pre-existing immunity.

Currently, vaccines for SARS-CoV-2 and influenza viruses are updated if the new vaccine induces higher antibody-titers to circulating variants than current vaccines. This approach does not account for complex dynamics of how prior immunity skews recall responses to the updated vaccine. We: (i) use computational models to mechanistically dissect how prior immunity influences recall responses; (ii) explore how this affects the rules for evaluating and deploying updated vaccines; and (iii) apply this to SARS-CoV-2. Our analysis of existing data suggests that there is a strong benefit to updating the current SARS-CoV-2 vaccines to match the currently circulating variants. We propose a general two-dose strategy for determining if vaccines need updating as well as for vaccinating high-risk individuals. Finally, we directly validate our model by reanalysis of earlier human H5N1 influenza vaccine studies.

Structural insights for neutralization of Omicron variants BA.1, BA.2, BA.4, and BA.5 by a broadly neutralizing SARS-CoV-2 antibody.

In this study, by characterizing several human monoclonal antibodies (mAbs) isolated from single B cells of the COVID-19-recovered individuals in India who experienced ancestral Wuhan strain (WA.1) of SARS-CoV-2 during early stages of the pandemic, we found a receptor binding domain (RBD)-specific mAb 002-S21F2 that has rare gene usage and potently neutralized live viral isolates of SARS-CoV-2 variants including Alpha, Beta, Gamma, Delta, and Omicron sublineages (BA.1, BA.2, BA.2.12.1, BA.4, and BA.5) with IC50 ranging from 0.02 to 0.13 µg/ml. Structural studies of 002-S21F2 in complex with spike trimers of Omicron and WA.1 showed that it targets a conformationally conserved epitope on the outer face of RBD (class 3 surface) outside the ACE2-binding motif, thereby providing a mechanistic insights for its broad neutralization activity. The discovery of 002-S21F2 and the broadly neutralizing epitope it targets have timely implications for developing a broad range of therapeutic and vaccine interventions against SARS-CoV-2 variants including Omicron sublineages.

Humoral Responses Against SARS-CoV-2 and Variants of Concern After mRNA Vaccines in Patients With Non-Hodgkin Lymphoma and Chronic Lymphocytic Leukemia.

PURPOSE: Patients with non-Hodgkin lymphoma including chronic lymphocytic leukemia (NHL/CLL) are at higher risk of severe SARS-CoV-2 infection. We investigated vaccine-induced antibody responses in patients with NHL/CLL against the original SARS-CoV-2 strain and variants of concern including B.1.167.2 (Delta) and B.1.1.529 (Omicron). MATERIALS AND METHODS: Blood from 121 patients with NHL/CLL receiving two doses of vaccine were collected longitudinally. Antibody binding against the full-length spike protein, the receptor-binding, and N-terminal domains of the original strain and of variants was measured using a multiplex assay. Live-virus neutralization against Delta, Omicron, and the early WA1/2020 strains was measured using a focus reduction neutralization test. B cells were measured by flow cytometry. Correlation between vaccine response and clinical factors was determined. RESULTS: Mean anti-SARS-CoV-2 spike immunoglobulin G-binding titers were 85-fold lower in patients with NHL/CLL compared with healthy controls, with seroconversion occurring in only 67% of patients. Neutralization titers were also lower and correlated with binding titers (P < .0001). Treatment with anti-CD20-directed therapies within 1 year resulted in 136-fold lower binding titers. Peripheral blood B-cell count also correlated with vaccine response. At 3 months from last anti-CD20-directed therapy, B-cell count ≥ 4.31/µL blood around the time of vaccination predicted response (OR 7.46, P = .04). Antibody responses also correlated with age. Importantly, neutralization titers against Delta and Omicron were reduced six- and 42-fold, respectively, with 67% of patients seropositive for WA1/2020 exhibiting seronegativity for Omicron. CONCLUSION: Antibody binding and live-virus neutralization against SARS-CoV-2 and its variants of concern including Delta and Omicron were substantially lower in patients with NHL/CLL compared with healthy vaccinees. Anti-CD20-directed therapy < 1 year before vaccination and number of circulating B cells strongly predict vaccine response.

Persistence of Virus-Specific Antibody after Depletion of Memory B Cells.

Humoral immunity is a major component of the adaptive immune response against viruses and other pathogens with pathogen-specific antibody acting as the first line of defense against infection. Virus-specific antibody levels are maintained by continual secretion of antibody by plasma cells residing in the bone marrow. This raises the important question of how the virus-specific plasma cell population is stably maintained and whether memory B cells are required to replenish plasma cells, balancing their loss arising from their intrinsic death rate. In this study, we examined the longevity of virus-specific antibody responses in the serum of mice following acute viral infection with three different viruses: lymphocytic choriomeningitis virus (LCMV), influenza virus, and vesicular stomatitis virus (VSV). To investigate the contribution of memory B cells to the maintenance of virus-specific antibody levels, we employed human CD20 transgenic mice, which allow for the efficient depletion of B cells with rituximab, a human CD20-specific monoclonal antibody. Mice that had resolved an acute infection with LCMV, influenza virus, or VSV were treated with rituximab starting at 2 months after infection, and the treatment was continued for up to a year postinfection. This treatment regimen with rituximab resulted in efficient depletion of B cells (>95%), with virus-specific memory B cells being undetectable. There was an early transient drop in the antibody levels after rituximab treatment followed by a plateauing of the curve with virus-specific antibody levels remaining relatively stable (half-life of 372 days) for up to a year after infection in the absence of memory B cells. The number of virus-specific plasma cells in the bone marrow were consistent with the changes seen in serum antibody levels. Overall, our data show that virus-specific plasma cells in the bone marrow are intrinsically long-lived and can maintain serum antibody titers for extended periods of time without requiring significant replenishment from memory B cells. These results provide insight into plasma cell longevity and have implications for B cell depletion regimens in cancer and autoimmune patients in the context of vaccination in general and especially for COVID-19 vaccines. IMPORTANCE Following vaccination or primary virus infection, virus-specific antibodies provide the first line of defense against reinfection. Plasma cells residing in the bone marrow constitutively secrete antibodies, are long-lived, and can thus maintain serum antibody levels over extended periods of time in the absence of antigen. Our data, in the murine model system, show that virus-specific plasma cells are intrinsically long-lived but that some reseeding by memory B cells might occur. Our findings demonstrate that, due to the longevity of plasma cells, virus-specific antibody levels remain relatively stable in the absence of memory B cells and have implications for vaccination.

Longitudinal analysis shows durable and broad immune memory after SARS-CoV-2 infection with persisting antibody responses and memory B and T cells.

Ending the COVID-19 pandemic will require long-lived immunity to SARS-CoV-2. Here, we evaluate 254 COVID-19 patients longitudinally up to 8 months and find durable broad-based immune responses. SARS-CoV-2 spike binding and neutralizing antibodies exhibit a bi-phasic decay with an extended half-life of >200 days suggesting the generation of longer-lived plasma cells. SARS-CoV-2 infection also boosts antibody titers to SARS-CoV-1 and common betacoronaviruses. In addition, spike-specific IgG+ memory B cells persist, which bodes well for a rapid antibody response upon virus re-exposure or vaccination. Virus-specific CD4+ and CD8+ T cells are polyfunctional and maintained with an estimated half-life of 200 days. Interestingly, CD4+ T cell responses equally target several SARS-CoV-2 proteins, whereas the CD8+ T cell responses preferentially target the nucleoprotein, highlighting the potential importance of including the nucleoprotein in future vaccines. Taken together, these results suggest that broad and effective immunity may persist long-term in recovered COVID-19 patients.

Characterization of neutralizing versus binding antibodies and memory B cells in COVID-19 recovered individuals from India.

India is one of the most affected countries by COVID-19 pandemic; but little is understood regarding immune responses to SARS-CoV-2 in this region. Herein we examined SARS-CoV-2 neutralizing antibodies, IgG, IgM, IgA and memory B cells in COVID-19 recovered individual from India. While a vast majority of COVID-19 recovered individuals showed SARS-CoV-2 RBD-specific IgG, IgA and IgM antibodies (38/42, 90.47%; 21/42, 50%; 33/42, 78.57% respectively), only half of them had appreciable neutralizing antibody titers. RBD-specific IgG, but not IgA or IgM titers, correlated with neutralizing antibody titers and RBD-specific memory B cell frequencies. These findings have timely significance for identifying potential donors for plasma therapy using RBD-specific IgG assays as surrogate measurement for neutralizing antibodies in India. Further, this study provides useful information needed for designing large-scale studies towards understanding of inter-individual variation in immune memory to SARS CoV-2 natural infection for future vaccine evaluation and implementation efforts.

Rapid Generation of Neutralizing Antibody Responses in COVID-19 Patients.

SARS-CoV-2, the virus responsible for COVID-19, is causing a devastating worldwide pandemic, and there is a pressing need to understand the development, specificity, and neutralizing potency of humoral immune responses during acute infection. We report a cross-sectional study of antibody responses to the receptor-binding domain (RBD) of the spike protein and virus neutralization activity in a cohort of 44 hospitalized COVID-19 patients. RBD-specific IgG responses are detectable in all patients 6 days after PCR confirmation. Isotype switching to IgG occurs rapidly, primarily to IgG1 and IgG3. Using a clinical SARS-CoV-2 isolate, neutralizing antibody titers are detectable in all patients by 6 days after PCR confirmation and correlate with RBD-specific binding IgG titers. The RBD-specific binding data were further validated in a clinical setting with 231 PCR-confirmed COVID-19 patient samples. These findings have implications for understanding protective immunity against SARS-CoV-2, therapeutic use of immune plasma, and development of much-needed vaccines.