Neutralization of SARS‐CoV‐2 requires antibodies against conformational receptor‐binding domain epitopes

Abstract

Background The determinants of successful humoral immune response to the severe acute respiratory syndrome coronavirus 2 (SARS‐CoV‐2) are of critical importance for the design of effective vaccines and the evaluation of the degree of protective immunity conferred by exposure to the virus. As novel variants emerge, understanding their likelihood of suppression by population antibody repertoires has become increasingly important. Methods In this study, we analyzed the SARS‐CoV‐2 polyclonal antibody response in a large population of clinically well‐characterized patients after mild and severe COVID‐19 using a panel of microarrayed structurally folded and unfolded SARS‐CoV‐2 proteins, as well as sequential peptides, spanning the surface spike protein (S) and the receptor‐binding domain (RBD) of the virus. Results S‐ and RBD‐specific antibody responses were dominated by immunoglobulin G (IgG), mainly IgG 1 , and directed against structurally folded S and RBD and three distinct peptide epitopes in S2. The virus neutralization activity of patients´ sera was highly correlated with IgG antibodies specific for conformational but not sequential RBD epitopes and their ability to prevent RBD binding to its human receptor angiotensin‐converting enzyme 2 (ACE2). Twenty percent of patients selectively lacked RBD‐specific IgG. Only immunization with folded, but not with unfolded RBD, induced antibodies against conformational epitopes with high virus‐neutralizing activity. Conformational RBD epitopes required for protection do not seem to be altered in the currently emerging virus variants. Conclusion These results are fundamental for estimating the protective activity of antibody responses after natural infection or vaccination and for the design of vaccines, which can induce high levels of SARS‐CoV‐2–neutralizing antibodies conferring sterilizing immunity.

Evaluation of a commercial ELISA as alternative to plaque reduction neutralization test to detect neutralizing antibodies against SARS-CoV-2

“…In contrast, neutralizing antibodies that are present after natural infections target also virus epitopes other than the S-protein RBD, including other virus proteins. Indeed, a study by Gattinger showed that 20.9% of 253 COVID patients did not develop detectable IgGs to RBD, but did in most cases (83% of the RBD-non-responders) develop IgGs to other S-protein epitopes or NCP 16 . Hence this could result in an underestimation of neutralizing antibodies in convalescent individuals by the GenScript sVNT (which targets the RBD of the S-protein), and it can be conceived that the GenScript sVNT can more accurately quantify neutralizing antibodies in vaccinees compared to convalescent patients.…”

Characterization of the antibody response to SARS‐CoV‐2 in a mildly affected pediatric population

“…Anti‐RBD IgG antibodies were found in 84.6% of COVID‐19 convalescent children, whereas in 15.4% of children, IgG antibodies against RBD were not detected. Using the same assay, approximately 20% of adult patients were negative for RBD‐specific IgG, with females being affected more often 20 . Whether these discrepancies confer to the previously observed sex‐related susceptibility of COVID‐19 remains open 30 .…”

“…coli ‐expressed, unfolded S1 20 ; HEK cell‐expressed, folded S2, Native Antigen Company, Oxford, UK; E . coli ‐expressed, unfolded S2 20 ; HEK cell‐expressed, folded RBD, Genscript; E . coli ‐expressed, unfolded RBD 20 ) were spotted onto pre‐activated glass slides as described 20,27,28 .…”

“…In majority of the SARS‐CoV‐2 convalescent adult patients, antibodies to S, RBD, and NCP were detected 2,17‐20 . Further investigation revealed that neutralizing antibodies recognize conformational, but neither linear epitopes nor S‐derived peptides 20 .…”

“…Additionally, we investigated the virus‐neutralizing activity of antibodies and their ability to inhibit the interaction of RBD and ACE2 in a recently developed molecular interaction assay 24 . Moreover, we compared results obtained in children with data from a SARS‐CoV‐2‐affected adult population collected at the same time period 20,25 …”