Hepatitis C virus resistance to broadly neutralizing antibodies measured using replication-competent virus and pseudoparticles

Abstract

A better understanding of natural variation in neutralization resistance and fitness of diverse hepatitis C virus (HCV) envelope (E1E2) variants will be critical to guide rational development of an HCV vaccine. This work has been hindered by inadequate genetic diversity in viral panels and by a lack of standardization of HCV entry assays. Neutralization assays generally use lentiviral pseudoparticles expressing HCV envelope proteins (HCVpp) or chimeric full-length viruses that are replication competent in cell culture (HCVcc). There have been few systematic comparisons of specific infectivities of E1E2-matched HCVcc and HCVpp, and to our knowledge, neutralization of E1E2-matched HCVpp and HCVcc has never been compared using a diverse panel of human broadly neutralizing monoclonal antibodies (bNAbs) targeting distinct epitopes. Here, we describe an efficient method for introduction of naturally occurring E1E2 genes into a full-length HCV genome, producing replication-competent chimeric HCVcc. We generated diverse panels of E1E2-matched HCVcc and HCVpp and measured the entry-mediating fitness of E1E2 variants using the two systems. We also compared neutralization of E1E2-matched HCVcc and HCVpp by a diverse panel of human bNAbs targeting epitopes across E1E2. We found no correlation between specific infectivities of E1E2-matched HCVcc versus HCVpp, but found a very strong positive correlation between relative neutralization resistance of these same E1E2-matched HCVcc and HCVpp variants. These results suggest that quantitative comparisons of neutralization resistance of E1E2 variants can be made with confidence using either HCVcc or HCVpp, allowing the use of either or both systems to maximize diversity of neutralization panels.

Cholesterol sensing by CD81 is important for hepatitis C virus entry

“…Direct comparison of the data acquired using the J6 HCVpp and J6/JFH HCVcc indicates excellent agreement (R 2 = 0.96, S3 Fig. ); this further demonstrates that HCVpp are a good model for HCV entry [56].…”

Synergistic anti-HCV broadly neutralizing human monoclonal antibodies with independent mechanisms

“…6). Experimental neutralization of 1a53 HCVcc exceeded Loewe-predicted neutralization at 7 of 10 concentrations (P < 0.05 for each), while it differed from Bliss-predicted virus (22,24). Experimental neutralization of strain 1a53 HCVpp significantly exceeded the neutralization predicted by the Loewe additivity model at six of nine concentrations (P < 0.05 for each), while it differed significantly from the Bliss prediction at only two of nine concentrations.…”

“…1A and Table S1). We analyzed the neutralizing breadth of each of these NAbs in prior studies using a diverse panel of 19 genotype 1 HCVpp (20,(22)(23)(24). We previously showed that these quantitative neutralization data across diverse strains comprise a neutralization profile for each NAb, which can be compared among NAbs to understand functional relationships (22).…”

“…Generation of an HCVcc chimera expressing the 1a53 E1E2 proteins was previously described (24,25). HCVcc neutralization assays were performed in triplicate as previously described (24,25).…”

“…While some human bNAbs show impressive neutralizing breadth (5,9,(20)(21)(22), HCV is an extraordinarily diverse virus, so no single bNAb neutralizes all viral strains (21)(22)(23)(24)(25). This resistance makes it less likely that vaccine induction of any single bNAb would provide adequate protection against a diverse HCV quasispecies challenge.…”