Mapping a Region of Hepatitis C Virus E2 That Is Responsible for Escape from Neutralizing Antibodies and a Core CD81-Binding Region That Does Not Tolerate Neutralization Escape Mutations

Abstract

Understanding the interaction between broadly neutralizing antibodies and their epitopes provides a basis for the rational design of a preventive hepatitis C virus (HCV) vaccine. CBH-2, HC-11, and HC-1 are representatives of antibodies to overlapping epitopes on E2 that mediate neutralization by blocking virus binding to CD81. To obtain insights into escape mechanisms, infectious cell culture virus, 2a HCVcc, was propagated under increasing concentrations of a neutralizing antibody to isolate escape mutants. Three escape patterns were observed with these antibodies. First, CBH-2 escape mutants that contained mutations at D431G or A439E, which did not compromise viral fitness, were isolated. Second, under the selective pressure of HC-11, escape mutations progressed from a single L438F substitution at a low antibody concentration to double substitutions, L438F and N434D or L438F and T435A, at higher antibody concentrations. Escape from HC-11 was associated with a loss of viral fitness. An HCV pseudoparticle (HCVpp) containing the L438F mutation bound to CD81 half as efficiently as did wild-type (wt) HCVpp. Third, for HC-1, the antibody at a critical concentration completely suppressed viral replication and generated no escape mutants. Epitope mapping revealed contact residues for CBH-2 and HC-11 in two regions of the E2 glycoprotein, amino acids (aa) 425 to 443 and aa 529 to 535. Interestingly, contact residues for HC-1 were identified only in the region encompassing aa 529 to 535 and not in aa 425 to 443. Taken together, these findings point to a region of variability, aa 425 to 443, that is responsible primarily for viral escape from neutralization, with or without compromising viral fitness. Moreover, the region aa 529 to 535 is a core CD81 binding region that does not tolerate neutralization escape mutations.Up to 170 million people worldwide are chronically infected with hepatitis C virus (HCV), with many at significant risk for liver failure and hepatocellular carcinoma (http://www.who.int /vaccine_research/diseases/viral_cancers/en/index2.html). The virus is transmitted primarily by parenteral routes and injection drug use in developed countries, whereas contaminated injection equipment appears to be the major risk factor for HCV infection in developing countries. From unsafe needle injections alone, the World Health Organization estimates an annual increase in the global burden by 2 million new infections (35). Current therapy with combined pegylated interferon and ribavirin has led to clinical improvement for some patients, but treatment is associated with adverse side effects and a high relapse rate off therapy. Clearly, additional approaches are needed for treatment and prevention of infection. However, an effective HCV vaccine has yet to be achieved, despite considerable effort. A major impediment is the genetic diversity of the virus. The phylogenetic tree of HCV contains seven major genotypes with more than 30% divergence between genotypes, and each genotype contains a large number of related subtype...

Broadly neutralizing antibodies with few somatic mutations and hepatitis C virus clearance

“…Broadly neutralizing human mAbs (bNAbs) capable of neutralizing diverse HCV strains have been isolated from HCV-infected individuals, proving that antibodies can target relatively conserved regions of the two HCV envelope glycoproteins (E1 and E2), despite the enormous genetic diversity of HCV (6)(7)(8)(9)(10)(11)(12)(13)(14)(15)(16)(17). Infusion of bNAbs is protective against infection in animal models of HCV (17,18), and a recent study also showed that bNAbs could abrogate established HCV infection in a humanized transgenic mouse model (19).…”

Naturally selected hepatitis C virus polymorphisms confer broad neutralizing antibody resistance

“…However, isolation of human mAbs capable of neutralizing multiple diverse HCV isolates has shown that nAbs may also target more conserved regions of the E1 and E2 proteins (12)(13)(14)(15)(16)(17)(18)(19)(20)(21)(22)(23)(24)(25). Discovery of these broadly neutralizing mAbs has raised hope that a vaccine inducing similar nAbs could prevent HCV infection.…”

Cooperativity in Virus Neutralization by Human Monoclonal Antibodies to Two Adjacent Regions Located at the Amino Terminus of Hepatitis C Virus E2 Glycoprotein

“…Antibodies to antigenic domain B share two contact residues at G530 and D535 (30,33,35), which are universally conserved and have been shown to participate in the interaction of E2 with CD81 (19,36). Some of the antigenic domain B epitopes also contain residues located within aa 434 to 446, e.g., an epitope identified by an antibody designated HC-11 (35,37 425 ) were synthesized using a C-terminal biotin residue with a Gly-Ser-Gly linker (American Peptide, Sunnyvale, CA).…”