Monoclonal Antibodies Directed toward the Hepatitis C Virus Glycoprotein E2 Detect Antigenic Differences Modulated by the N-Terminal Hypervariable Region 1 (HVR1), HVR2, and Intergenotypic Variable Region

Abstract

Hepatitis C virus (HCV) envelope glycoproteins E1 and E2 form a heterodimer and mediate receptor interactions and viral fusion. Both E1 and E2 are targets of the neutralizing antibody (NAb) response and are candidates for the production of vaccines that generate humoral immunity. Previous studies demonstrated that N-terminal hypervariable region 1 (HVR1) can modulate the neutralization potential of monoclonal antibodies (MAbs), but no information is available on the influence of HVR2 or the intergenotypic variable region (igVR) on antigenicity. In this study, we examined how the variable regions influence the antigenicity of the receptor binding domain of E2 spanning HCV polyprotein residues 384 to 661 (E2661) using a panel of MAbs raised against E2661 and E2661 lacking HVR1, HVR2, and the igVR (Δ123) and well-characterized MAbs isolated from infected humans. We show for a subset of both neutralizing and nonneutralizing MAbs that all three variable regions decrease the ability of MAbs to bind E2661 and reduce the ability of MAbs to inhibit E2-CD81 interactions. In addition, we describe a new MAb directed toward the region spanning residues 411 to 428 of E2 (MAb24) that demonstrates broad neutralization against all 7 genotypes of HCV. The ability of MAb24 to inhibit E2-CD81 interactions is strongly influenced by the three variable regions. Our data suggest that HVR1, HVR2, and the igVR modulate exposure of epitopes on the core domain of E2 and their ability to prevent E2-CD81 interactions. These studies suggest that the function of HVR2 and the igVR is to modulate antibody recognition of glycoprotein E2 and may contribute to immune evasion.IMPORTANCE This study reveals conformational and antigenic differences between the Δ123 and intact E2661 glycoproteins and provides new structural and functional data about the three variable regions and their role in occluding neutralizing and nonneutralizing epitopes on the E2 core domain. The variable regions may therefore function to reduce the ability of HCV to elicit NAbs directed toward the conserved core domain. Future studies aimed at generating a three-dimensional structure for intact E2 containing HVR1, and the adjoining NAb epitope at residues 412 to 428, together with HVR2, will reveal how the variable regions modulate antigenic structure.

An Optimized Hepatitis C Virus E2 Glycoprotein Core Adopts a Functional Homodimer That Efficiently Blocks Virus Entry

“…Our analysis clearly demonstrated that simultaneous replacement of HVR2 and the igVR cooperatively stabilized the CD81-binding site within E2, despite these individual mutations resulting in reduced CD81-LEL binding. We have recently reported that the ability of both a neutralizing antibody and an apparently nonneutralizing antibody to inhibit E2-CD81 binding was significantly improved when HVR2 and the igVR were deleted, with or without HVR1, suggesting that these variable regions can modulate the CD81-LEL binding site and epitopes therein (52).…”

Computational Prediction of the Heterodimeric and Higher-Order Structure of gpE1/gpE2 Envelope Glycoproteins Encoded by Hepatitis C Virus

“…7), HVR2 from one E2 monomer is positioned close to HVR1 from another monomer and is flanked on another side by IgVR. This observation is supported by experimental data suggesting that HVR1, HVR2, and IgVR are structurally linked in the virion (64). For example, antibodies that bind epitope I in E2 have been shown to exhibit improved binding to E2 and an improved ability to inhibit E2-CD81 interactions when any one of these hypervariable regions was deleted.…”

“…For example, antibodies that bind epitope I in E2 have been shown to exhibit improved binding to E2 and an improved ability to inhibit E2-CD81 interactions when any one of these hypervariable regions was deleted. Furthermore, inhibition of the E2-CD81 interaction was additive if two or more of the hypervariable regions were deleted (64). In addition, antibodies that recognize an epitope that included HVR1 residues were shown to have a reduced affinity for E1/E2 when IgVR was deleted but not when IgVR and HVR2 were deleted simultaneously (64).…”

“…Indeed, previous studies have shown that HVR1 can interfere with the binding of antibodies to E2 (64,71), and HCV pseudoparticles lacking HVR1 have been reported to exhibit increased sensitivity to a range of neutralizing monoclonal antibodies that are directed to diverse E2 epitopes (72). This observation is surprising since these antibody binding sites are not all clustered into one region.…”

“…Furthermore, inhibition of the E2-CD81 interaction was additive if two or more of the hypervariable regions were deleted (64). In addition, antibodies that recognize an epitope that included HVR1 residues were shown to have a reduced affinity for E1/E2 when IgVR was deleted but not when IgVR and HVR2 were deleted simultaneously (64). A possible explanation for this observation is that IgVR may be important for constraining the position of HVR2 in wild-type E2.…”