Broadly neutralizing antibodies from an individual that naturally cleared multiple hepatitis C virus infections uncover molecular determinants for E2 targeting and vaccine design

Hepatitis C virus (HCV) is a major cause of liver disease, affecting over 2% of the world’s population. The HCV envelope glycoproteins E1 and E2 mediate viral entry, with E2 being the main target of neutralizing antibody responses. Structural investigations of E2 have produced templates for vaccine design, including the conserved CD81 receptor-binding site (CD81bs) that is a key target of broadly neutralizing antibodies (bNAbs). Unfortunately, immunization with recombinant E2 and E1E2 rarely elicits sufficient levels of bNAbs for protection. To understand the challenges for eliciting bNAb responses against the CD81bs, we investigated the E2 CD81bs by electron microscopy (EM), hydrogen–deuterium exchange (HDX), molecular dynamics (MD), and calorimetry. By EM, we observed that HCV1, a bNAb recognizing the N-terminal region of the CD81bs, bound a soluble E2 core construct from multiple angles of approach, suggesting components of the CD81bs are flexible. HDX of multiple E2 constructs consistently indicated the entire CD81bs was flexible relative to the rest of the E2 protein, which was further confirmed by MD simulations. However, E2 has a high melting temperature of 84.8 °C, which is more akin to proteins from thermophilic organisms. Thus, recombinant E2 is a highly stable protein overall, but with an exceptionally flexible CD81bs. Such flexibility may promote induction of nonneutralizing antibodies over bNAbs to E2 CD81bs, underscoring the necessity of rigidifying this antigenic region as a target for rational vaccine design.

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Broadly neutralizing antibodies protect against hepatitis C virus quasispecies challenge

Nature Medicine ◽

10.1038/nm1698 ◽

2007 ◽

Vol 14 (1) ◽

pp. 25-27 ◽

Cited By ~ 416

Author(s):

Mansun Law ◽

Toshiaki Maruyama ◽

Jamie Lewis ◽

Erick Giang ◽

Alexander W Tarr ◽

...

Keyword(s):

Hepatitis C Virus ◽

Hepatitis C ◽

Neutralizing Antibodies ◽

Broadly Neutralizing Antibodies

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Antibody Response to Hypervariable Region 1 Interferes with Broadly Neutralizing Antibodies to Hepatitis C Virus

Journal of Virology ◽

10.1128/jvi.02458-15 ◽

2016 ◽

Vol 90 (6) ◽

pp. 3112-3122 ◽

Cited By ~ 35

Author(s):

Zhen-yong Keck ◽

Christine Girard-Blanc ◽

Wenyan Wang ◽

Patrick Lau ◽

Adam Zuiani ◽

...

Keyword(s):

Hepatitis C Virus ◽

Hepatitis C ◽

Persistent Infection ◽

Neutralizing Antibodies ◽

Hypervariable Region ◽

Humoral Response ◽

Broadly Neutralizing Antibodies ◽

Alanine Scanning Mutagenesis ◽

Effect Analysis ◽

Hypervariable Region 1

ABSTRACTHypervariable region 1 (HVR1) (amino acids [aa] 384 to 410) on the E2 glycoprotein of hepatitis C virus contributes to persistent infection by evolving escape mutations that attenuate binding of inhibitory antibodies and by blocking access of broadly neutralizing antibodies to their epitopes. A third proposed mechanism of immune antagonism is that poorly neutralizing antibodies binding to HVR1 interfere with binding of other superior neutralizing antibodies. Epitope mapping of human monoclonal antibodies (HMAbs) that bind to an adjacent, conserved domain on E2 encompassing aa 412 to 423 revealed two subsets, designated HC33 HMAbs. While both subsets have contact residues within aa 412 to 423, alanine-scanning mutagenesis suggested that one subset, which includes HC33.8, has an additional contact residue within HVR1. To test for interference of anti-HVR1 antibodies with binding of antibodies to aa 412 to 423 and other E2 determinants recognized by broadly neutralizing HMAbs, two murine MAbs against HVR1 (H77.16) and aa 412 to 423 (H77.39) were studied. As expected, H77.39 inhibited the binding of all HC33 HMAbs. Unexpectedly, H77.16 also inhibited the binding of both subsets of HC33 HMAbs. This inhibition also was observed against other broadly neutralizing HMAbs to epitopes outside aa 412 to 423. Combination antibody neutralization studies by the median-effect analysis method with H77.16 and broadly reactive HMAbs revealed antagonism between these antibodies. Structural studies demonstrated conformational flexibility in this antigenic region, which supports the possibility of anti-HVR1 antibodies hindering the binding of broadly neutralizing MAbs. These findings support the hypothesis that anti-HVR1 antibodies can interfere with a protective humoral response against HCV infection.IMPORTANCEHVR1 contributes to persistent infection by evolving mutations that escape from neutralizing antibodies to HVR1 and by shielding broadly neutralizing antibodies from their epitopes. This study provides insight into a new immune antagonism mechanism by which the binding of antibodies to HVR1 blocks the binding and activity of broadly neutralizing antibodies to HCV. Immunization strategies that avoid the induction of HVR1 antibodies should increase the inhibitory activity of broadly neutralizing anti-HCV antibodies elicited by candidate vaccines.

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Extra-epitopic hepatitis C virus polymorphisms confer resistance to broadly neutralizing antibodies by modulating binding to scavenger receptor B1

PLoS Pathogens ◽

10.1371/journal.ppat.1006235 ◽

2017 ◽

Vol 13 (2) ◽

pp. e1006235 ◽

Cited By ~ 22

Author(s):

Ramy El-Diwany ◽

Valerie J. Cohen ◽

Madeleine C. Mankowski ◽

Lisa N. Wasilewski ◽

Jillian K. Brady ◽

...

Keyword(s):

Hepatitis C Virus ◽

Hepatitis C ◽

Neutralizing Antibodies ◽

Scavenger Receptor ◽

Broadly Neutralizing Antibodies

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Faculty Opinions recommendation of Broadly neutralizing antibodies protect against hepatitis C virus quasispecies challenge.

Faculty Opinions – Post-Publication Peer Review of the Biomedical Literature ◽

10.3410/f.1112205.568197 ◽

2008 ◽

Author(s):

Ray Chung

Keyword(s):

Hepatitis C Virus ◽

Hepatitis C ◽

Neutralizing Antibodies ◽

Broadly Neutralizing Antibodies

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A Hepatitis C Virus Envelope Polymorphism Confers Resistance to Neutralization by Polyclonal Sera and Broadly Neutralizing Monoclonal Antibodies

Journal of Virology ◽

10.1128/jvi.02837-15 ◽

2016 ◽

Vol 90 (7) ◽

pp. 3773-3782 ◽

Cited By ~ 16

Author(s):

Lisa N. Wasilewski ◽

Ramy El-Diwany ◽

Supriya Munshaw ◽

Anna E. Snider ◽

Jillian K. Brady ◽

...

Keyword(s):

Hepatocellular Carcinoma ◽

Monoclonal Antibodies ◽

Hepatitis C Virus ◽

Hepatitis C ◽

Neutralizing Antibodies ◽

Vaccine Development ◽

Resistance Mutation ◽

The United States ◽

Broadly Neutralizing Antibodies ◽

Access To Treatment

ABSTRACTHepatitis C virus (HCV) infection is a global health problem, with millions of chronically infected individuals at risk for cirrhosis and hepatocellular carcinoma. HCV vaccine development is vital in the effort toward disease control and eradication, an undertaking aided by an increased understanding of the mechanisms of resistance to broadly neutralizing antibodies (bNAbs). In this study, we identified HCV codons that vary deep in a phylogenetic tree of HCV sequences and showed that a polymorphism at one of these positions renders Bole1a, a computationally derived, ancestral genotype 1a HCV strain, resistant to neutralization by both polyclonal-HCV-infected plasma and multiple broadly neutralizing monoclonal antibodies with unique binding epitopes. This bNAb resistance mutation reduces replicative fitness, which may explain the persistence of both neutralization-sensitive and neutralization-resistant variants in circulating viral strains. This work identifies an important determinant of bNAb resistance in an ancestral, representative HCV genome, which may inform HCV vaccine development.IMPORTANCEWorldwide, more than 170 million people are infected with hepatitis C virus (HCV), the leading cause of hepatocellular carcinoma and liver transplantation in the United States. Despite recent significant advances in HCV treatment, a vaccine is needed. Control of the HCV pandemic with drug treatment alone is likely to fail due to limited access to treatment, reinfections in high-risk individuals, and the potential for resistance to direct-acting antivirals (DAAs). Broadly neutralizing antibodies (bNAbs) block infection by diverse HCV variants and therefore serve as a useful guide for vaccine development, but our understanding of resistance to bNAbs is incomplete. In this report, we identify a viral polymorphism conferring resistance to neutralization by both polyclonal plasma and broadly neutralizing monoclonal antibodies, which may inform HCV vaccine development.

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Broadly Neutralizing Antibodies Targeting New Sites of Vulnerability in Hepatitis C Virus E1E2

Journal of Virology ◽

10.1128/jvi.02070-18 ◽

2019 ◽

Vol 93 (14) ◽

Cited By ~ 5

Author(s):

Michelle D. Colbert ◽

Andrew I. Flyak ◽

Clinton O. Ogega ◽

Valerie J. Kinchen ◽

Guido Massaccesi ◽

...

Keyword(s):

Monoclonal Antibodies ◽

Hepatitis C Virus ◽

Hepatitis C ◽

Neutralizing Antibodies ◽

Vaccine Development ◽

Infected Individual ◽

Hcv Infection ◽

Broadly Neutralizing Antibodies ◽

Genotype 1A ◽

Antigenic Sites

ABSTRACTIncreasing evidence indicates that broadly neutralizing antibodies (bNAbs) play an important role in immune-mediated control of hepatitis C virus (HCV) infection, but the relative contribution of neutralizing antibodies targeting antigenic sites across the HCV envelope (E1 and E2) proteins is unclear. Here, we isolated thirteen E1E2-specific monoclonal antibodies (MAbs) from B cells of a single HCV-infected individual who cleared one genotype 1a infection and then became persistently infected with a second genotype 1a strain. These MAbs bound six distinct discontinuous antigenic sites on the E1 protein, the E2 protein, or the E1E2 heterodimer. Three antigenic sites, designated AS108, AS112 (an N-terminal E1 site), and AS146, were distinct from previously described antigenic regions (ARs) 1 to 5 and E1 sites. Antibodies targeting four sites (AR3, AR4-5, AS108, and AS146) were broadly neutralizing. These MAbs also displayed distinct patterns of relative neutralizing potency (i.e., neutralization profiles) across a panel of diverse HCV strains, which led to complementary neutralizing breadth when they were tested in combination. Overall, this study demonstrates that HCV bNAb epitopes are not restricted to previously described antigenic sites, expanding the number of sites that could be targeted for vaccine development.IMPORTANCEWorldwide, more than 70 million people are infected with hepatitis C virus (HCV), which is a leading cause of hepatocellular carcinoma and liver transplantation. Despite the development of potent direct acting antivirals (DAAs) for HCV treatment, a vaccine is urgently needed due to the high cost of treatment and the possibility of reinfection after cure. Induction of multiple broadly neutralizing antibodies (bNAbs) that target distinct epitopes on the HCV envelope proteins is one approach to vaccine development. However, antigenic sites targeted by bNAbs in individuals with spontaneous control of HCV have not been fully defined. In this study, we characterize 13 monoclonal antibodies (MAbs) from a single person who cleared an HCV infection without treatment, and we identify 3 new sites targeted by neutralizing antibodies. The sites targeted by these MAbs could inform HCV vaccine development.

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