Researchers identify 55 broadly reactive monoclonal antibodies to betacoronaviruses

By Neha MathurNov 8 2022Reviewed by Danielle Ellis, B.Sc.

In a recent study published in the journal Cell Host & Microbe, researchers pursued novel approaches that could help find monoclonal antibodies (mAbs) broadly targeting coronaviruses (CoVs), especially genera beta CoVs.

Study: Rare, convergent antibodies targeting the stem helix broadly neutralize diverse betacoronaviruses. Image Credit: Naeblys / Shutterstock

Background

Three subgenera of betacoronaviruses cause respiratory diseases in humans. Lineage A betacoronaviruses, including HCoV-OC43 and HCoV-HKU1, cause mild upper respiratory infections. Middle Eastern respiratory syndrome CoV (MERS-CoV), SARS-CoV, and severe acute respiratory syndrome CoV-2 (SARS-CoV-2) are lineage C and B betaCoVs, respectively, that caused three recent outbreaks in the past two decades and claimed millions of lives globally.

Currently, the most concerning is the emergence of SARS-CoV-2 Omicron subvariant BA.5, which is resistant to most clinically available mAbs. Thus, there is an urgent need for therapeutic mAbs that broadly target beta-CoVs.

About the study

The present study enrolled 19 COVID-19 convalescent donors who displayed plasma reactivity to diverse beta CoVs. They obtained 673,671 immunoglobulin G (IgG+) and 305,142 IgA+ memory B cells (MBCs) from these donors, which they screened using a two-step workflow. This workflow utilized sequential oligoclonal and monoclonal B cell cultures to identify B cells of interest.

The researchers tested the mAb panel comprising  55 broadly reactive mAbs for binding to SARS-CoV-2 receptor-binding domain (RBD), N-terminal domain (NTD), S1, and S2. Flow cytometry analyses revealed the SARS-CoV-2 spike S2 subunit as the target of the majority of the mAbs. Subsequent surface plasmon resonance (SPR)-based epitope binning analysis demonstrated that the mAbs could be divided into two groups targeting different fusion peptide regions. Further, the team assessed the breadth and potency of the neutralization of the study's mAb panel.

First, the team performed a sequence alignment of 28 isolates representing the five betacoronavirus subgenera to determine the degree of conservation of the stem helix sequence among beta-CoVs. Next, they investigated the genetic profile of the stem helix-specific mAbs. So they examined their heavy, light chain V gene usage and their complementarity-determining region 3 (CDR3) amino acid sequences. Further, they compared the potency and breadth of the IGHV1-46/IGKV3-20 mAbs to the other broadly reactive mAbs.

The team conducted an alanine scan on the stem helix peptide to determine whether the IGHV1-46/IGKV3-20 mAbs preferentially formed contacts with a distinct set of amino acids from the other mAbs targeting this site. In addition, they used the S shotgun mutagenesis assay to examine the binding profile of COV89-22, the most potent stem helix-targeting mAb of the study panel, since it had the least susceptibility to mutations in the stem helix region.

Study findings

The researchers identified 55 monoclonal antibodies from COVID-19 convalescent donors that bound diverse beta CoV S proteins. Amino acids, F1148, E1151, K1157, and N1158 within the stem helix are highly conserved (>90%) within the betacoronavirus subgenera. However, the human alphaCoVs, HCoV-229E, and HCoV-NL63 have divergent sequences at this location. All SARS-CoV-2 VOCs identified to date, including the Omicron subvariant BA.5, have identical sequences in this region.

Thus, most mAbs targeted an S2 epitope that included the K814 residue and was non-neutralizing. In addition, 11 antibodies targeting the stem helix of the S2 subunit neutralized beta CoVs from different lineages. Yet, none of the 55 mAbs could neutralize HCoV-NL63 (an alphacoronavirus), even at the highest tested concentration of 100 μg/mL.

Eight antibodies in this group, including the six broadest and most potent neutralizers, were encoded by IGHV1-46 and IGKV3-20. The IGHV1-46/IGKV3-20 combination was used very frequently (72.7%) by mAbs targeting the stem helix, suggesting positive selection due to favorable binding to the stem helix. Also, the germinal center reaction enhanced the antibody response to this site.

Conclusions

These findings identified a class of IGHV1-46/IGKV3-20 antibodies that broadly neutralized beta-CoVs by targeting the stem helix. However, these antibodies constituted only a small fraction of the broadly reactive antibodies to beta-CoVs after SARS-CoV-2 infection. The S2 subunit, which is more conserved than S1, is currently being explored as a candidate for next-generation coronavirus vaccines. The study findings showed that a targeted vaccine construct that triggered an immune response on the stem helix and avoided the immunodominant K814+ site that masked this site might be promising against diverse betaCoVs.

Crystal structures of three antibodies of this class at 1.5–1.75-Å resolution and mutagenesis data revealed a conserved mode of binding of potent stem helix-specific mAbs. This interaction mode could serve as a template for the design of a vaccine construct that elicits the desired antibody response, i.e., germline-targeting immunogens that activate B cell lineages.

The mAbs targeting the stem helix showed lower in vitro neutralization potency relative to the RBD-specific mAbs, such as sotrovimab. It does not always reflect efficacy in humans, as other factors, such as Fc activity, also contribute to protection. In fact, COV89-22, a stem helix-specific mAb was effective in preventing disease mediated by SARS-CoV-2 in a hamster model; thus, studies should further explore its usage.