Glycan Shifting on Hepatitis C Virus (HCV) E2 Glycoprotein Is a Mechanism for Escape from Broadly Neutralizing Antibodies

Pantua, Homer; Diao, Jun; Ultsch, Mark; Hazen, Meredith; Mathieu, Mary; McCutcheon, Krista; Takeda, Kazuhisa; Date, Shailesh V.; Cheung, Tommy K.; Phung, Qui; Hass, Phil; Arnott, David; Hongo, Jo-Anne; Matthews, David J.; Brown, Alex; Patel, Arvind H.; Kelley, Robert F.; Eigenbrot, Charles; Kapadia, Sharookh B.

doi:10.1016/j.jmb.2013.02.025

Cited by 100 publications

(146 citation statements)

References 60 publications

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“…Our results extend and confirm previous observations and speculations made from crystal structures showing that the AS412 region is disordered when unbound (14,41) and adopts a hairpin or open-loop conformation when complexed with different NAbs (17)(18)(19)(20)(21)(22). Such alternate conformations appear to be critical for virus neutralization.…”

Section: Resultssupporting

confidence: 90%

“…The first structural investigations of HCV E2 revealed that AS412 within the E2 CD81bs, when presented as a synthetic peptide, forms a β-hairpin when bound to bNAbs HCV1 and AP33 (17)(18)(19)(20). Both HCV1 and AP33 interact with one face of the AS412 β-hairpin, whereas the opposite face contains an N-linked glycan.…”

Section: Resultsmentioning

confidence: 99%

“…The first antigenic site (AS412) consists of the conserved N-terminal end of the CD81bs spanning residues 412-423. The bNAbs AP33, HCV1, HC33.1, and 3/11 primarily bind to E2 at AS412, and their interactions have been structurally characterized (17)(18)(19)(20)(21)(22). The second antigenic site (AS434) consists of a short α-helix in the front layer residues 434-446.…”

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confidence: 99%

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Structural flexibility at a major conserved antibody target on hepatitis C virus E2 antigen

Kong

Lee

Kadam

et al. 2016

Proc. Natl. Acad. Sci. U.S.A.

102

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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.hepatitis C virus | E2 | CD81-binding site | conformational flexibility | protein dynamics H epatitis C virus (HCV) is a leading cause of liver cirrhosis and hepatocellular carcinoma, infecting more than 2% of the world's population (1). HCV infection is highly prevalent in developing countries and among marginalized populations, such as injection drug users (IDUs) and prisoners in developed countries (2). Effective direct-acting antiviral (DAA) drugs have been developed recently to curb the advance of HCV (3, 4). Nevertheless, new infections are on the rise among young IDUs in developed countries and along drug-trafficking routes (2, 5, 6). Because DAA treatment is prohibitively expensive and does not prevent reinfection (7,8), an effective vaccine is essential for management of the global HCV epidemic (9-11).Despite the significant public health burden, no effective prophylactic vaccine against HCV has been developed. High genetic variability of the virus is a major barrier, particularly in the E1 and E2 envelope glycoproteins that are the primary neutralizing antibody (NAb) targets. E2, the receptor-binding protein, mediates cell entry by interacting with the tetraspanin CD81 and several other cell surface molecules (12, 13). The crystal structure of the E2 core domain (E2c) consists of a central β-sandwich flanked by "front" and "back" layers (14) (Fig. 1). The CD81-binding site (CD81bs) has been mapped by mutagenesis and electron microscopy (EM) to various elements: a conserved N-terminal...

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Section: Resultssupporting

confidence: 90%

Section: Resultsmentioning

confidence: 99%

mentioning

confidence: 99%

See 1 more Smart Citation

Structural flexibility at a major conserved antibody target on hepatitis C virus E2 antigen

Kong

Lee

Kadam

et al. 2016

Proc. Natl. Acad. Sci. U.S.A.

102

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“…In addition, a number of studies have highlighted the role of N-linked glycosylation in shielding E2 epitopes from recognition by broadly neutralizing antibodies (31)(32)(33), as observed for HIV and influenza virus (34). In a particularly revealing recent example, deep sequencing analysis of HCV resistance to neutralization by mAbs HCV1 and AP33 identified mutations at asparagine 417 of E2 (N417S and N417T), an N-glycosylation site within the E2 412-423 epitope recognized by these mAbs, as responsible for virus escape (35). Comparison of the glycosylation status of E2 containing these resistance mutations revealed a glycosylation shift from Asn-417 to Asn-415 in the N417S and N417T E2 proteins.…”

Section: Hepatitis C Virus (Hcv)mentioning

confidence: 99%

“…Crystal structures of the wild-type E2 412-423 epitope bound to HCV1 and AP33 showed that, whereas the side chain of Asn-417 points away from the antibody, Asn-415 is buried within the peptide-antibody interface (35)(36)(37)(38). Consequently, attachment of a glycan at Asn-415, but not at Asn-417, would create major steric clashes with HCV1 and AP33, resulting in abrogation of antibody binding and HCV escape, as observed in resistance selection studies (29,35,39). Notably, liver transplantation patients with HCV who were treated with HCV1 showed rebounds in viral load at various times following transplantation (39).…”

Section: Hepatitis C Virus (Hcv)mentioning

confidence: 99%

Structural Basis for Penetration of the Glycan Shield of Hepatitis C Virus E2 Glycoprotein by a Broadly Neutralizing Human Antibody

Pierce

Wang

et al. 2015

Journal of Biological Chemistry

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Background: HCV uses glycan shielding to mask epitopes recognized by neutralizing antibodies. Results: The structure of a human antibody bound to an HCV E2 epitope revealed how it penetrates a shield created by glycosylation shifting. Conclusion: Antibody binding induces an epitope conformation that accommodates multiple glycans. Significance: The structure provides a template for engineering E2 to elicit protective antibodies able to overcome glycosylation shifting.

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Monoclonal anti‐envelope antibody AP33 protects humanized mice against a patient‐derived hepatitis C virus challenge

et al. 2016

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End-stage liver disease (ESLD) caused by hepatitis C virus (HCV) infection is a major indication for liver transplantation. However, immediately after transplantation, the liver graft of viremic patients universally becomes infected by circulating virus, resulting in accelerated liver disease progression. Currently available direct-acting antiviral therapies have reduced efficacy in patients with ESLD and prophylactic strategies to prevent HCV recurrence are still highly needed. In this study, we compared the ability of two broadly reactive monoclonal antibodies (mAbs), designated 3/11 and AP33, recognizing a distinct, but overlapping, epitope in the viral E2 glycoprotein to protect humanized mice from a patient-derived HCV challenge. Their neutralizing activity was assessed using the HCV pseudoparticles and cell-culture-derived HCV systems expressing multiple patient-derived envelopes and a human-liver chimeric mouse model. HCV RNA was readily detected in all control mice challenged with a patient-derived HCV genotype 1b isolate, whereas 3 of 4 AP33-treated mice were completely protected. In contrast, only one of four 3/11-treated mice remained HCV-RNA negative throughout the observation period, whereas the other 3 had a viral load that was indistinguishable from that in the control group. The increased in vivo efficacy of AP33 was in line with its higher affinity and neutralizing capacity observed in vitro. Conclusions: Although mAbs AP33 and 3/11 target the same region in E2, only mAb AP33 can efficiently protect from challenge with a heterologous HCV population in vivo. Given that mAb AP33 efficiently neutralizes viral variants that escaped the humoral immune response and reinfected the liver graft of transplant patients, it may be a valuable candidate to prevent HCV recurrence. In addition, our data are valuable for the design of a prophylactic vaccine. (HEPATOLOGY 2016;63:1120-1134

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Glycan Shifting on Hepatitis C Virus (HCV) E2 Glycoprotein Is a Mechanism for Escape from Broadly Neutralizing Antibodies

Cited by 100 publications

References 60 publications

Structural flexibility at a major conserved antibody target on hepatitis C virus E2 antigen

Structural flexibility at a major conserved antibody target on hepatitis C virus E2 antigen

Structural Basis for Penetration of the Glycan Shield of Hepatitis C Virus E2 Glycoprotein by a Broadly Neutralizing Human Antibody

Monoclonal anti‐envelope antibody AP33 protects humanized mice against a patient‐derived hepatitis C virus challenge

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