Comparative Immunomodulatory Evaluation of the Receptor Binding Domain of the SARS-CoV-2 Spike Protein; a Potential Vaccine Candidate Which Imparts Potent Humoral and Th1 Type Immune Response in a Mouse Model

Abstract

The newly emerged novel coronavirus, SARS-CoV-2, the causative agent of COVID-19 has proven to be a threat to the human race globally, thus, vaccine development against SARS-CoV-2 is an unmet need driving mass vaccination efforts. The receptor binding domain of the spike protein of this coronavirus has multiple neutralizing epitopes and is associated with viral entry. Here we have designed and characterized the SARS-CoV-2 spike protein fragment 330-526 as receptor binding domain 330-526 (RBD330-526) with two native glycosylation sites (N331 and N343); as a potential subunit vaccine candidate. We initially characterized RBD330-526 biochemically and investigated its thermal stability, humoral and T cell immune response of various RBD protein formulations (with or without adjuvant) to evaluate the inherent immunogenicity and immunomodulatory effect. Our result showed that the purified RBD immunogen is stable up to 72 h, without any apparent loss in affinity or specificity of interaction with the ACE2 receptor. Upon immunization in mice, RBD generates a high titer humoral response, elevated IFN-γ producing CD4+ cells, cytotoxic T cells, and robust neutralizing antibodies against live SARS-CoV-2 virus. Our results collectively support the potential of RBD330-526 as a promising vaccine candidate against SARS-CoV-2.

Designed SARS‐CoV‐2 receptor binding domain variants form stable monomers

“…Other studies of RBD mutants with N‐terminal truncations that removed T323 also report monomer loss upon incubation at room temperature. [ 24 ] However, the capacity of RBD to interact with ACE2 was thereby not compromised, which is consistent with structural data demonstrating that T323 is not located in the proximity of the ACE2‐binding site. [ 12 ]…”

A Novel Bacterial Protease Inhibitor Adjuvant in RBD-Based COVID-19 Vaccine Formulations Containing Alum Increases Neutralizing Antibodies, Specific Germinal Center B Cells and Confers Protection Against SARS-CoV-2 Infection in Mice

“…However, only alum containing formulations resulted in detectable antibody titers after the first immunization. This result is in line with literature reporting low immunogenicity in mice after a single dose when RBD is formulated with AddaVax ( 51 – 54 ), but significant seroconversion titers after single dose when formulated with alum ( 41 , 54 ) or after two doses when formulated with AddaVax or AddaS03 ( 40 , 47 , 51 , 52 , 54 ). The overall response was dominated by the IgG1 subclass in all immunized groups.…”

A novel bacterial protease inhibitor adjuvant in RBD-based COVID-19 vaccine formulations increases neutralizing antibodies, specific germinal center B cells and confers protection against SARS-CoV-2 infection

“…However, only alum containing formulations resulted in detectable antibody titers after the first immunization. This result is in line with literature reporting low immunogenicity in mice after a single dose when RBD is formulated with AddaVax [50][51][52][53] , but significant seroconversion titers after single dose when formulated with alum 40,53 or after two doses when formulated with AddaVax or AddaS03 39,46,50,51,53 . The overall response was dominated by the IgG1 subclass in all immunized groups.…”