Broadly neutralizing human monoclonal antibodies to the hepatitis C virus E2 glycoprotein

Abstract

The humoral response to hepatitis C virus (HCV) may contribute to controlling infection. We previously isolated human monoclonal antibodies to conformational epitopes on the HCV E2 glycoprotein. Here, we report on their ability to inhibit infection by retroviral pseudoparticles incorporating a panel of full-length E1E2 clones representing the full spectrum of genotypes 1–6. We identified one antibody, CBH-5, that was capable of neutralizing every genotype tested. It also potently inhibited chimeric cell culture-infectious HCV, which had genotype 2b envelope proteins in a genotype 2a (JFH-1) background. Analysis using a panel of alanine-substitution mutants of HCV E2 revealed that the epitope of CBH-5 includes amino acid residues that are required for binding of E2 to CD81, a cellular receptor essential for virus entry. This suggests that CBH-5 inhibits HCV infection by competing directly with CD81 for a binding site on E2.

In vitro selection of a neutralization-resistant hepatitis C virus escape mutant

“…These mutations were not found in virus passaged in parallel without AP33 neutralization. Although N415 is within a domain identified previously as comprising the AP33 epitope 413-420 (22,23), E655 is distant in the linear E2 sequence and has not been implicated previously in recognition of HCV by AP33. To validate the role of these mutations in conferring neutralization escape, they were engineered both singly and together into pHJ3-5 plasmid DNA, and the cognate viruses rescued after RNA transfection of naive cells.…”

“…HC-1, Ϫ2, -11, -12, -13, and CBH-5 recognize residues comprising ''domain B'' and neutralize both genotype 1a and 2a virus, whereas CBH-8C and -11 recognize overlapping epitopes but neutralize only the 2a virus (23,28). CBH-7 recognizes an epitope in domain C that does not overlap those bound by domain B antibodies and that is capable of neutralizing both (B) AP33-selected viruses from the third, fifth, and sixth rounds of neutralization were tested for neutralization with AP33 (100 g/ml) by FFU-reduction assay.…”

“…One such mAb, AP33, is of particular interest because it demonstrates broad and potent, cross-genotypic neutralizing activity. Prior studies using synthetic peptides, recombinant antigens, and HCVpp suggest that AP33 binds a highly conserved, linear epitope spanning residues 413-420 (22,23). Here, we describe the use of a cell culture-infectious, intergenotypic chimeric HCV with genotype 1a structural proteins (24,25) to further define the AP33 epitope by analyzing mutants selected for the ability to escape neutralization by this mAb.…”

Low pH-dependent Hepatitis C Virus Membrane Fusion Depends on E2 Integrity, Target Lipid Composition, and Density of Virus Particles

“…For this series of experiments, we chose the human monoclonal antibody CBH-5, which neutralizes infectivity of HCVpp carrying diverse HCV genotypes (17,55), via binding to an epitope within E2. The Jc1 virus was preincubated with increasing concentrations of CBH-5 at neutral pH, and the fusion of such mixtures was then assessed after acidification in our lipid mixing assay.…”

“…Both HCV envelope glycoproteins are the targets for virusneutralizing antibodies (7,(15)(16)(17)(18)(19). In E2, one important neutralizing epitope is the so-called hypervariable region 1, which includes the 27 amino-terminal residues of E2 (20 -22).…”