Neutralizing epitopes of the SARS-CoV S-protein cluster independent of repertoire, antigen structure or mAb technology

Abstract

Neutralizing antibody responses to the surface glycoproteins of enveloped viruses play an important role in immunity. Many of these glycoproteins, including the severe acute respiratory syndrome-coronavirus (SARS-CoV) spike (S) protein form trimeric units in the membrane of the native virion. There is substantial experimental and pre-clinical evidence showing that the S protein is a promising lead for vaccines and therapeutics. Previously we generated a panel of monoclonal antibodies (mAbs) to whole inactivated SARS-CoV which neutralize the virus in vitro. Here, we define their specificity and affinity, map several of their epitopes and lastly characterise chimeric versions of them. Our data show that the neutralizing mAbs bind to the angiotensin-converting enzyme 2 (ACE2) receptor-binding domain (RBD) of the SARS S protein. Three of the chimeric mAbs retain their binding specificity while one conformational mAb, F26G19, lost its ability to bind the S protein despite high level expression. The affinity for recombinant S is maintained in all of the functional chimeric versions of the parental mAbs. Both parental mAb F26G18 and the chimeric version neutralize the TO R2 strain of SARS-CoV with essentially identical titres (2.07 and 2.47 nM, respectively). Lastly, a comparison with other neutralizing mAbs to SARS-CoV clearly shows that the dominance of a 33 amino acid residue loop of the SARS-CoV RBD is independent of repertoire, species, quaternary structure, and importantly, the technology used to derive the mAbs. In cases like this, the dominance of a compact RBD antigenic domain and the central role of the S protein in pathogenesis may inherently create immunoselection pressure on viruses to evolve more complex evasion strategies or die out of a host species. The apparent simplicity of the mechanism of SARS-CoV neutralization is in stark contrast to the complexity shown by other enveloped viruses.

Functional analysis of the receptor binding domain of SARS coronavirus S1 region and its monoclonal antibody

“…It seems that this RBD region plays a critical role attaching the host ACE2 molecule. This phenomenon was also supported by chimeric monoclonal antibodies that bind to the ACE2 RBD of the SARS S protein (Berry et al 2010). Experimental support for ACE2-RBD binding was provided from a study of SARS antibody fragments competing with ACE2 for binding to the RBD (Prabakaran et al 2006;Sui et al 2004;Hwang et al 2006).…”

Perspectives on monoclonal antibody therapy as potential therapeutic intervention for Coronavirus disease-19 (COVID-19)

“…• Blocking the interaction of S subunit protein using CDR loops H1, H2, H3, and L3 with cellular receptor ACE2 in vitro. 42,46 1A9…”

“…• Blocking the interaction of S1 subunit protein with cellular receptor ACE2 using 6 complementary determining region (CDR) in vitro and in vivo (Mouse). 11,20,41,42 CR3014…”

Quantitative and sensitive detection of the SARS-CoV spike protein using bispecific monoclonal antibody-based enzyme-linked immunoassay

“…Anti-HRPO YP4 is a well-characterized rat hybridoma that was selected previously for drug resistance to 8-azaguanine, making it sensitive to aminopterine in HAT medium. Anti-SARS-CoV S1 F26G18 is a mouse hybridoma that can bind the receptor binding domain in S1 region of SARS CoV (Berry et al, 2010). Two cell lines of YP4 and F26G18 were chosen for developing quadromas (hybridoma × hybridoma) (Kammila et al, 2008).…”

“…The profiles of antibody responses to SARS-CoV have been well established . Reports indicate that specific viral proteins might be better markers for serological detection of infection by SARS-CoV (Tan et al, 2004;Lu et al, 2004;Berry et al, 2010).…”