Structural and functional dissection of neutralisation differences among SARS-CoV-2 variants using antigenicity prediction and CR3022 binding analysis

Variations in cross-neutralizing reactivity is dependent on mutations on immunogenic epitopes in the spike protein of SARS-CoV-2 variants.

PURPOSE The COVID-19 pandemic has witnessed the rapid evolution of the SARS-CoV-2 virus, resulting in the emergence of multiple variants with mutations in critical regions of the spike protein, notably in the receptor binding domain. These mutations can lead to immune evasion and breakthrough (BT) infections, even in the vaccinated individuals. While previous studies have documented various mutations, there is a limited understanding of these mutations in different epitopes and their influence on the vaccine elicited immune response. METHODS Neutralizing antibodies were detected using Plaque reduction neutralization test and peptide enzyme linked immunosorbent assay was standardized to identify possible epitopes contributing to immune escape. We used GraphPad Prism 9.5.0 (525) for analysis. Results were significant at p < 0.05. RESULTS In this study, we observed that individuals with hybrid immunity, defined as immunity derived from both the natural infection and vaccination, exhibit significantly higher levels of neutralizing antibodies than individuals who had been solely vaccinated with COVISHIELD. Consequently, we highlight the substantial differences in the reactivity of neutralizing antibodies against the Wuhan, Delta, and Omicron variants. Our findings indicate that individuals who have recovered from COVID-19 and /or who experienced BT infections, showed stronger reactivity toward the conserved peptides than individuals vaccinated with no prior exposure. CONCLUSIONS COVISHIELD induced antibodies effectively neutralized original Wuhan strain but showed reduced neutralization against Delta and Omicron due to mutations in specific epitopes within their spike proteins. Mutations in variant specific epitopes can lead to evasion of the virus, emphasizing the importance of targeting linear region epitopes as critical binding sites.

Anti-SARS-CoV-2 receptor-binding domain IgG titers and associated factors after heterologous mRNA booster vaccination in individuals primed with the inactivated vaccine.

Coronavirus disease 2019 (COVID-19), caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), continues to circulate globally, highlighting the need to understand immune responses after heterologous booster vaccination. This cross-sectional study evaluated anti-SARS-CoV-2 receptor-binding domain (RBD) IgG titers and associated factors among adults who had completed a two-dose CoronaVac primary series and received a mRNA-based booster dose (BNT162b2 or mRNA-1273) 1-13 months before sampling, with no documented or self-reported history of COVID-19. Anti-SARS-CoV-2 RBD IgG was measured using the Abbott SARS-CoV-2 IgG II Quant assay. Antibody titers were ln-transformed and analyzed using independent t-tests, one-way analysis of variance, and log-linear regression, with results reported as geometric mean titers (GMTs) and geometric mean ratios (GMRs). A total of 158 participants were included. Anti-SARS-CoV-2 RBD IgG GMTs were not significantly different between the two booster groups. In the combined analysis, the highest GMT was observed in the ≤3-month post-booster interval group, followed by lower GMTs at longer intervals. After adjustment for age, sex, and body mass index, participants sampled at 4-6 months had significantly lower titers than those sampled at ≤3 months (adjusted GMR=0.42; 95%CI: 0.22-0.82). Overweight participants had higher titers than those with normal body mass index in the combined analysis (adjusted GMR=1.73; 95%CI: 1.04-2.86). These findings suggest lower humoral immune responses with increasing time after heterologous mRNA booster vaccination in individuals primed with CoronaVac and may inform future booster strategies.

Structural and mutational analyses define distinct molecular routes to broad SARS-CoV-2 receptor-binding domain recognition

Broadly reactive antibodies elicited by SARS-CoV-2 infection or vaccination can reveal conserved viral vulnerabilities and inform vaccines with broad coronavirus coverage. Here, we characterize two human-derived monoclonal antibodies, B2014 and C5078, that recognize conserved epitopes on the SARS-CoV-2 RBD and retain activity across antigenically distinct variants. Notably, C5078 also recognizes diverse sarbecoviruses and remains active against currently circulating variants, including XFG and NB.1.8.1. Cryo-EM structures reveal that B2014 recognizes an epitope adjacent to the class 3 antibody site, whereas C5078 targets the highly conserved, cryptic site V epitope. Structural analysis defines how C5078 uses affinity-matured interactions to engage conserved RBD residues, providing a molecular basis for its exceptional breadth. Deep mutational scanning across multiple SARS-CoV-2 variant backgrounds further defines potential pathways of antibody escape, explaining the loss of B2014 activity against antigenically evolved variants while revealing a high barrier to escape from C5078. Together, these findings define distinct structural solutions for broad RBD recognition and highlight conserved, mutationally constrained epitopes that may serve as targets for vaccines designed to elicit antibody responses resilient to ongoing SARS-CoV-2 evolution and future sarbecovirus emergence.

Antibody evasion and receptor binding of SARS-CoV-2 variants PQ.16.1.1 and RK.1

Evaluations utilizing surface plasmon resonance and pseudovirus assays demonstrate that these sublineages exhibit significantly reduced human ACE2 receptor engagement compared to their parental strain, which suggests these variants will soon spread globally and emphasize the critical need for ongoing surveillance to monitor D420N-carrying lineages.

Structure-Based Adaptation of a SARS-CoV-2 Neutralizing Peptide to New Virus Variants.

The starting point of this work was a SARS-CoV-2 neutralizing peptide (LW25.13), which binds to the receptor-binding domain of the viral spike protein and inhibits the attachment of the virus to its cellular receptor ACE2. As LW25.13 is unable to neutralize later SARS-CoV-2 variants, such as omicron, we have extended the neutralization breadth of LW25.13 through structural and bioinformatic analysis. This involved the systematic variation of a range of positions and yielded peptides neutralizing SARS-CoV-2 beta and omicron at low nanomolar concentrations, while preserving the strong neutralizing capacity against earlier virus variants (wild-type, alpha, delta), as well as the proteolytic stability and α-helical conformation of the peptide. This gain in neutralizing breadth illustrates the utility of the peptide as a scaffold that can be adapted to different virus variants, which may prove useful for the development of peptides against new coronavirus variants of concern in the future.

Immunoinformatics-guided design of a conserved multi-epitope vaccine candidate against pathogenic human coronaviruses.