Identification, structures and ontogenies of SARS-CoV-2 antibodies

National Institute Of Allergy And Infectious Diseases

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Abstract

The continued emergence of SARS-CoV-2 Omicron sub-variants evaded neutralization by antibodies elicited against the viral spike protein of early variants. To map or evaluate the antibody responses and antibody repertoire elicited by current vaccines or infection, we developed molecular probes based on new variants of concern. Utilizing these probes, our study characterized the repertoire and epitope specificity of antibodies elicited by infection with the Beta, Gamma and WA1 ancestral variants and assessed their cross-reactivity to these and the more recent Delta and Omicron variants. A technique was created for acquiring antibody sequences capable of quickly generating and evaluating monoclonal antibodies from numerous probes sorted B cells isolated through flow cytometry. In response to infection by any variant, comparable cross-binding antibody reactions were triggered, demonstrating a consistent ranking of dominant epitopes. Moreover, there was a consistent employment of convergent V genes and similar B cell clones visible across all infecting variants. These shared responses, even amid variations in the antigen, might explain the persistent effectiveness of vaccines relying on an initial single variant. Our collaborator selected two synthetic DARPin-derived proteins-FSR16m and FSR22-for the possible treatment of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection. FSR16m and FSR22 are trimeric proteins composed of DARPin SR16m or SR22 fused with a T4 foldon. Despite being selected using a spike protein derived from a now historical SARS-CoV-2 strain, both FSR16m and FSR22 demonstrate a wide-ranging ability to neutralize various SARS-CoV-2 strains. They effectively inhibit the activity of authentic B.1.351, B.1.617.2, and BA.1.1 viruses. To elucidate the neutralization mechanism, we performed cryo-EM structural analysis which revealed that these DARPins (Designed Ankyrin Repeat Proteins) recognize a specific segment within the receptor-binding domain (residues 456, 475, 486, 487, and 489) that overlaps a critical region of the angiotensin-converting enzyme 2 (ACE2)-binding surface. Intranasal administration of FSR16m protected K18-hACE2 transgenic mice exposed to B.1.617.2. Given the potent and extensive neutralization capabilities of FSR16m and FSR22, these agents hold substantial promise as candidates for both preventing and treating SARS-CoV-2 infections. In a third study, we employed a yeast display technology that maintains natural antibody pairing, combined with next-generation sequencing (NGS) and extensive bioinformatic analysis. The goal is to comprehensively investigate the subdomain specificity of natural human antibodies from two convalescent donors. Utilizing this advanced screening approach, the study maps the cross-reactive responses of antibodies generated by the two donors against various SARS-CoV-2 variants and other betacoronaviruses. The neutralization potency of a subset of cross-reactive antibodies generated in this study is tested, revealing that most antibodies from these patients do not possess neutralizing capabilities. A comparison of specific and non-specific antibodies is conducted by analyzing somatic hypermutation on a repertoire-wide scale for the two individuals. Remarkably, the extent of somatic hypermutation is found to be unique for each patient. The insights gained from this research provide functional understanding of cross-reactive antibodies, which could prove invaluable in devising strategies to combat emerging SARS-CoV-2 variants and divergent betacoronaviruses.