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.
Abstract
The recent global expansion of the SARS-CoV-2 variant NB.1.8.1 has driven the emergence of sublineages PQ.16.1.1 and RK.1, which independently acquired the D420N mutation in their receptor-binding domains and and now dominate the Asia-Pacific region. Evaluations utilizing surface plasmon resonance and pseudovirus assays demonstrate that these sublineages exhibit significantly reduced human ACE2 receptor engagement compared to their parental strain. However, this functional cost is offset by a marked ability to evade humoral immunity, specifically demonstrating profound resistance to Class 1 neutralizing monoclonal antibodies and convalescent plasma from Wuhan-Hu-1-primed individuals. This convergent evolution exemplifies a classical viral trade-off, sacrificing receptor binding efficiency to escape population-level immune pressure. Consequently, these findings suggest these variants will soon spread globally and emphasize the critical need for ongoing surveillance to monitor D420N-carrying lineages.
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.
Nina Raasch, L. Weissenborn, Elie Richel et al.· Journal of Medicinal Chemist...· 0 citations
A combined experimental and computational characterization of SARS-CoV-2 variant-specific neutralisation across infection, vaccination and hybrid immunity-driven cohorts is provided, offering a hypothesis-generating framework to contextualize observed differences in antibody responses and epitope recognition across variants.
Jyoti Sawant, Ajit Patil, Madhuri Thakar et al.· Frontiers in Immunology· 0 citations
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.
M. Abernathy, William B. Foreman, Jasmyn A. Lopez et al.· bioRxiv· 0 citations
ABSTRACT SARS-CoV-2 antigenic evolution continues to erode the activity of first-generation monoclonal antibodies, underscoring the value of antibodies that recognize conserved features within the spike receptor-binding domain (RBD). As a model for breadth-oriented engineering, we assessed the RBD-directed antibody XG83, which has a CDRH3-dominated paratope. Utilizing the XG83-Wuhan RBD crystal structure as a reference, we integrated MOE alanine scanning and residue scanning with CDRH3 (A116-Y132) to identify chemically reasonable replacements and test interaction to an Omicron BA.1 RBD. The parental CDRH3-centric pose in comparative docking had a better score (−280.18) than the mutant MuXG83 (−254.2), along with 100-ns molecular dynamics showed that MuXG83 was less stable (with higher RMSD/RMSF with less favorable interaction energy). The ELISA results against Omicron BA.1 RBD demonstrated that XG83 had 40% stronger binding than MuXG83; convergence happened only at the highest concentration, concordant with SPR studies and the fact that mutation increased dissociation. According to this binding gap, BA.1 pseudovirus neutralization demonstrated that XG83 was much more powerful than MuXG83, showing that the E118L/F130H CDR-H3 alterations decreased functional activity against Omicron BA.1. These results imply that allosteric influences on interface stability and conformational dynamics by non-epitope CDR-H3 residues can affect antibody performance. Functional testing was confined to Omicron BA.1; therefore, larger variant-panel studies are needed to ascertain if such mutations affect antibody breadth. Our findings highlight a structure-guided approach for optimizing paratopes and indicate that non-epitope (potentially allosteric) changes to CDRH3 are also important while investigating potential development and neutralization before advancement. IMPORTANCE This study shows how modest allosteric characteristics in CDR-H3 control the delicate balance between neutralizing potency and breadth, making a timely and significant addition to SARS-CoV-2 antibody engineering. Using crystallography, alanine scanning, residue scanning, docking, molecular dynamics, and experimental ELISA and neutralization assays, this study offers a structure-guided framework for rational paratope optimization. The discovery of CDR-H3 residues that regulate long-range stability rather than just direct epitope contacts reveals an unappreciated aspect of antibody design and explains why some alterations improve anticipated interactions but degrade functional performance. Importantly, the comparison of wild-type XG83 and modified MuXG83 shows how allosteric tuning affects antibody-antigen compatibility in developing variations like Omicron. These findings illuminate conserved RBD recognition and offer strategies for building next-generation therapeutic antibodies that are more resistant to viral evolution. This study shows how modest allosteric characteristics in CDR-H3 control the delicate balance between neutralizing potency and breadth, making a timely and significant addition to SARS-CoV-2 antibody engineering. Using crystallography, alanine scanning, residue scanning, docking, molecular dynamics, and experimental ELISA and neutralization assays, this study offers a structure-guided framework for rational paratope optimization. The discovery of CDR-H3 residues that regulate long-range stability rather than just direct epitope contacts reveals an unappreciated aspect of antibody design and explains why some alterations improve anticipated interactions but degrade functional performance. Importantly, the comparison of wild-type XG83 and modified MuXG83 shows how allosteric tuning affects antibody-antigen compatibility in developing variations like Omicron. These findings illuminate conserved RBD recognition and offer strategies for building next-generation therapeutic antibodies that are more resistant to viral evolution.
Muhammad Waqas Nasir, Qiyun Liang, Jun He et al.· Microbiology spectrum· 0 citations
HRBD demonstrated potent and broad-spectrum inhibition against Pangolin-CoV, SARS-CoV, SARS-CoV, SARS-CoV-2, and its variants, lowering the half-maximal inhibitory concentration (IC50) by approximately 1000-fold compared to the monomeric RBD.
Jintao Zou, Lingyu Su, Jiansheng Lu et al.· Antiviral Research· 0 citations
The continued emergence of SARS-CoV-2 variants worldwide underscores the need for effective measures to combat the current and future outbreaks. The receptor-binding domain (RBD) of the spike glycoprotein plays a critical role in viral entry and represents a highly immunogenic target. In this study, we generated a mouse-derived single-chain variable fragment (scFv) phage display library against the RBD of the SARS-CoV-2 spike protein. Peptide ELISA-based epitope mapping demonstrated that selected monoclonal scFv phages exhibited broad epitope recognition and reactivity across multiple recently reported variants. In a SARS-CoV-2 pseudovirus neutralization assay, scFv C4 showed potent neutralizing activity against the wild-type, Gamma, and Delta variants, with IC50 values below 4 μg/mL. These findings highlight the potential utility of these scFvs as candidate reagents for the development of improved diagnostic platforms and as scaffolds for next-generation monoclonal antibody-based therapeutics.
M. Alam, Saurabh Sharma, Romisa Razvi et al.· ACS Infectious Diseases· 0 citations