scholarly journals Genetic and structural insights into broad neutralization of hepatitis C virus by human VH1-69 antibodies

2019 ◽  
Vol 5 (1) ◽  
pp. eaav1882 ◽  
Author(s):  
Netanel Tzarum ◽  
Erick Giang ◽  
Leopold Kong ◽  
Linling He ◽  
Jannick Prentoe ◽  
...  
Keyword(s):  
Hepatitis C Virus ◽  
Hepatitis C ◽  
Gene Family ◽  
Neutralizing Antibodies ◽  
Germ Line ◽  
Affinity Maturation ◽  
Effective Vaccine ◽  
Immune Recognition ◽  
Sequencing Analysis ◽  
Cross Neutralization

An effective vaccine to the antigenically diverse hepatitis C virus (HCV) must target conserved immune epitopes. Here, we investigate cross-neutralization of HCV genotypes by broadly neutralizing antibodies (bNAbs) encoded by the relatively abundant human gene familyVH1-69. We have deciphered the molecular requirements for cross-neutralization by this unique class of human antibodies from crystal structures of HCV E2 in complex with bNAbs. An unusually high binding affinity is found for germ line–reverted versions of VH1-69 precursor antibodies, and neutralization breadth is acquired during affinity maturation. Deep sequencing analysis of an HCV-immune B cell repertoire further demonstrates the importance of theVH1-69gene family in the generation of HCV bNAbs. This study therefore provides critical insights into immune recognition of HCV with important implications for rational vaccine design.

scholarly journals Structure-Based Design of Hepatitis C Virus E2 Glycoprotein Improves Serum Binding and Cross-Neutralization

2020 ◽  
Vol 94 (22) ◽  
Author(s):  
Brian G. Pierce ◽  
Zhen-Yong Keck ◽  
Ruixue Wang ◽  
Patrick Lau ◽  
Kyle Garagusi ◽  
...  
Keyword(s):  
Hepatitis C Virus ◽  
Hepatitis C ◽  
Envelope Glycoprotein ◽  
Neutralizing Antibodies ◽  
Viral Escape ◽  
Envelope Glycoproteins ◽  
Effective Vaccine ◽  
Immunogenic Region ◽  
Antigenic Properties ◽  
Cross Neutralization

ABSTRACT An effective vaccine for hepatitis C virus (HCV) is a major unmet need, and it requires an antigen that elicits immune responses to key conserved epitopes. Based on structures of antibodies targeting HCV envelope glycoprotein E2, we designed immunogens to modulate the structure and dynamics of E2 and favor induction of broadly neutralizing antibodies (bNAbs) in the context of a vaccine. These designs include a point mutation in a key conserved antigenic site to stabilize its conformation, as well as redesigns of an immunogenic region to add a new N-glycosylation site and mask it from antibody binding. Designs were experimentally characterized for binding to a panel of human monoclonal antibodies (HMAbs) and the coreceptor CD81 to confirm preservation of epitope structure and preferred antigenicity profile. Selected E2 designs were tested for immunogenicity in mice, with and without hypervariable region 1, which is an immunogenic region associated with viral escape. One of these designs showed improvement in polyclonal immune serum binding to HCV pseudoparticles and neutralization of isolates associated with antibody resistance. These results indicate that antigen optimization through structure-based design of the envelope glycoproteins is a promising route to an effective vaccine for HCV. IMPORTANCE Hepatitis C virus infects approximately 1% of the world’s population, and no vaccine is currently available. Due to the high variability of HCV and its ability to actively escape the immune response, a goal of HCV vaccine design is to induce neutralizing antibodies that target conserved epitopes. Here, we performed structure-based design of several epitopes of the HCV E2 envelope glycoprotein to engineer its antigenic properties. Designs were tested in vitro and in vivo, demonstrating alteration of the E2 antigenic profile in several cases, and one design led to improvement of cross-neutralization of heterologous viruses. This represents a proof of concept that rational engineering of HCV envelope glycoproteins can be used to modulate E2 antigenicity and optimize a vaccine for this challenging viral target.

scholarly journals Production and Characterization of Monoclonal Antibodies Specific for a Conserved Epitope within Hepatitis C Virus Hypervariable Region 1

2001 ◽  
Vol 75 (24) ◽  
pp. 12412-12420 ◽  
Author(s):  
Chengyao Li ◽  
Daniel Candotti ◽  
Jean-Pierre Allain
Keyword(s):  
Monoclonal Antibodies ◽  
Hepatitis C Virus ◽  
Hepatitis C ◽  
Neutralizing Antibodies ◽  
Hypervariable Region ◽  
Passive Immunization ◽  
Immune Recognition ◽  
Hypervariable Region 1 ◽  
Neutralizing Capacity ◽  
Conserved Epitope

ABSTRACT Frequent mutations in hypervariable region 1 (HVR1) of the main envelope protein of hepatitis C virus (HCV) is a major mechanism of persistence by escaping the host immune recognition. HVR1 contains an epitope eliciting neutralizing antibodies. This study was aimed to prepare broadly cross-reacting, high-affinity, monoclonal antibodies (MAb) to the HVR1 C terminus of HCV with potential therapeutic neutralizing capacity. A conserved amino residue group of glycine (G) at position 23 and glutamic acid (Q) at position 26 in HVR1 was confirmed as a key epitope against which two MAbs were selected and characterized. MAbs 2P24 and 15H4 were immunoglobulin G1 kappa chain [IgG1(κ)], cross-reacted with 32 and 30 of 39 random C-terminal HVR1 peptides, respectively, and did not react with other HCV peptides. The VH of 2P24 and 15H4 heavy chains originated from Igh germ line v gene family 1 and 8, respectively. In contrast, the VL κ sequences were highly homologous. The affinity (K d ) of 2P24 and 15H4 (10−9 or 10−8 M with two immunizing peptides and 10−8 M with two nonimmunizing HVR1 peptides) paralleled the reactivity obtained with peptide enzyme immunoassay. MAbs 2P24 and 15H4 captured 25 of 31 (81%) HCV in unselected patients' plasmas. These antibodies also blocked HCV binding to Molt-4 cells in a dose-dependent fashion. The data presented suggest that broadly cross-reactive MAbs to a conserved epitope within HCV HVR1 can be produced. Clinical application for passive immunization in HCV-related chronic liver disease and after liver transplantation is considered.

scholarly journals Conformational Flexibility in the Immunoglobulin-Like Domain of the Hepatitis C Virus Glycoprotein E2

mBio ◽  
2017 ◽  
Vol 8 (3) ◽  
Author(s):  
Ieva Vasiliauskaite ◽  
Ania Owsianka ◽  
Patrick England ◽  
Abdul Ghafoor Khan ◽  
Sarah Cole ◽  
...  
Keyword(s):  
Hepatitis C Virus ◽  
Hepatitis C ◽  
Binding Site ◽  
Neutralizing Antibodies ◽  
Vaccine Development ◽  
Asymptomatic Infection ◽  
Effective Vaccine ◽  
Viral Glycoprotein ◽  
Direct Acting Antiviral ◽  
Glycoprotein E2

ABSTRACT The hepatitis C virus (HCV) glycoprotein E2 is the major target of neutralizing antibodies and is therefore highly relevant for vaccine design. Its structure features a central immunoglobulin (Ig)-like β-sandwich that contributes to the binding site for the cellular receptor CD81. We show that a synthetic peptide corresponding to a β-strand of this Ig-like domain forms an α-helix in complex with the anti-E2 antibody DAO5, demonstrating an inside-out flip of hydrophobic residues and a secondary structure change in the composite CD81 binding site. A detailed interaction analysis of DAO5 and cross-competing neutralizing antibodies with soluble E2 revealed that the Ig-like domain is trapped by different antibodies in at least two distinct conformations. DAO5 specifically captures retrovirus particles bearing HCV glycoproteins (HCVpp) and infectious cell culture-derived HCV particles (HCVcc). Infection of cells by DAO5-captured HCVpp can be blocked by a cross-competing neutralizing antibody, indicating that a single virus particle simultaneously displays E2 molecules in more than one conformation on its surface. Such conformational plasticity of the HCV E2 receptor binding site has important implications for immunogen design. IMPORTANCE Recent advances in the treatment of hepatitis C virus (HCV) infection with direct-acting antiviral drugs have enabled the control of this major human pathogen. However, due to their high costs and limited accessibility in combination with the lack of awareness of the mostly asymptomatic infection, there is an unchanged urgent need for an effective vaccine. The viral glycoprotein E2 contains regions that are crucial for virus entry into the host cell, and antibodies that bind to these regions can neutralize infection. One of the major targets of neutralizing antibodies is the central immunoglobulin (Ig)-like domain within E2. We show here that this Ig-like domain is conformationally flexible at the surface of infectious HCV particles and pseudoparticles. Our study provides novel insights into the interactions of HCV E2 with the humoral immune system that should aid future vaccine development. IMPORTANCE Recent advances in the treatment of hepatitis C virus (HCV) infection with direct-acting antiviral drugs have enabled the control of this major human pathogen. However, due to their high costs and limited accessibility in combination with the lack of awareness of the mostly asymptomatic infection, there is an unchanged urgent need for an effective vaccine. The viral glycoprotein E2 contains regions that are crucial for virus entry into the host cell, and antibodies that bind to these regions can neutralize infection. One of the major targets of neutralizing antibodies is the central immunoglobulin (Ig)-like domain within E2. We show here that this Ig-like domain is conformationally flexible at the surface of infectious HCV particles and pseudoparticles. Our study provides novel insights into the interactions of HCV E2 with the humoral immune system that should aid future vaccine development.

scholarly journals Structure-Based Design of Hepatitis C Virus Vaccines That Elicit Neutralizing Antibody Responses to a Conserved Epitope

2017 ◽  
Vol 91 (20) ◽  
Author(s):  
Brian G. Pierce ◽  
Elisabeth N. Boucher ◽  
Kurt H. Piepenbrink ◽  
Monir Ejemel ◽  
Chelsea A. Rapp ◽  
...  
Keyword(s):  
Hepatitis C Virus ◽  
Hepatitis C ◽  
Neutralizing Antibodies ◽  
Neutralizing Antibody ◽  
Antibody Responses ◽  
Therapeutic Options ◽  
Effective Vaccine ◽  
Sequence Variability ◽  
Peptide Epitope ◽  
Conserved Epitope

ABSTRACT Despite recent advances in therapeutic options, hepatitis C virus (HCV) remains a severe global disease burden, and a vaccine can substantially reduce its incidence. Due to its extremely high sequence variability, HCV can readily escape the immune response; thus, an effective vaccine must target conserved, functionally important epitopes. Using the structure of a broadly neutralizing antibody in complex with a conserved linear epitope from the HCV E2 envelope glycoprotein (residues 412 to 423; epitope I), we performed structure-based design of immunogens to induce antibody responses to this epitope. This resulted in epitope-based immunogens based on a cyclic defensin protein, as well as a bivalent immunogen with two copies of the epitope on the E2 surface. We solved the X-ray structure of a cyclic immunogen in complex with the HCV1 antibody and confirmed preservation of the epitope conformation and the HCV1 interface. Mice vaccinated with our designed immunogens produced robust antibody responses to epitope I, and their serum could neutralize HCV. Notably, the cyclic designs induced greater epitope-specific responses and neutralization than the native peptide epitope. Beyond successfully designing several novel HCV immunogens, this study demonstrates the principle that neutralizing anti-HCV antibodies can be induced by epitope-based, engineered vaccines and provides the basis for further efforts in structure-based design of HCV vaccines. IMPORTANCE Hepatitis C virus is a leading cause of liver disease and liver cancer, with approximately 3% of the world's population infected. To combat this virus, an effective vaccine would have distinct advantages over current therapeutic options, yet experimental vaccines have not been successful to date, due in part to the virus's high sequence variability leading to immune escape. In this study, we rationally designed several vaccine immunogens based on the structure of a conserved epitope that is the target of broadly neutralizing antibodies. In vivo results in mice indicated that these antigens elicited epitope-specific neutralizing antibodies, with various degrees of potency and breadth. These promising results suggest that a rational design approach can be used to generate an effective vaccine for this virus.

scholarly journals A Diverse Panel of Hepatitis C Virus Glycoproteins for Use in Vaccine Research Reveals Extremes of Monoclonal Antibody Neutralization Resistance

2015 ◽  
Vol 90 (7) ◽  
pp. 3288-3301 ◽  
Author(s):  
Richard A. Urbanowicz ◽  
C. Patrick McClure ◽  
Richard J. P. Brown ◽  
Theocharis Tsoleridis ◽  
Mats A. A. Persson ◽  
...  
Keyword(s):  
Monoclonal Antibodies ◽  
Hepatitis C Virus ◽  
Hepatitis C ◽  
Neutralizing Antibodies ◽  
Therapeutic Potential ◽  
Effective Vaccine ◽  
Antibody Neutralization ◽  
Neutralization Sensitivity ◽  
Vaccine Candidates ◽  
Intermediate Phenotypes

ABSTRACTDespite significant advances in the treatment of hepatitis C virus (HCV) infection, the need to develop preventative vaccines remains. Identification of the best vaccine candidates and evaluation of their performance in preclinical and clinical development will require appropriate neutralization assays utilizing diverse HCV isolates. We aimed to generate and characterize a panel of HCV E1E2 glycoproteins suitable for subsequent use in vaccine and therapeutic antibody testing. Full-length E1E2 clones were PCR amplified from patient-derived serum samples, cloned into an expression vector, and used to generate viral pseudoparticles (HCVpp). In addition, some of these clones were used to generate cell culture infectious (HCVcc) clones. The infectivity and neutralization sensitivity of these viruses were then determined. Bioinformatic and HCVpp infectivity screening of approximately 900 E1E2 clones resulted in the assembly of a panel of 78 functional E1E2 proteins representing distinct HCV genotypes and different stages of infection. These HCV glycoproteins differed markedly in their sensitivity to neutralizing antibodies. We used this panel to predict antibody efficacy against circulating HCV strains, highlighting the likely reason why some monoclonal antibodies failed in previous clinical trials. This study provides the first objective categorization of cross-genotype patient-derived HCV E1E2 clones according to their sensitivity to antibody neutralization. It has shown that HCV isolates have clearly distinguishable neutralization-sensitive, -resistant, or -intermediate phenotypes, which are independent of genotype. The panel provides a systematic means for characterization of the neutralizing response elicited by candidate vaccines and for defining the therapeutic potential of monoclonal antibodies.IMPORTANCEHepatitis C virus (HCV) has a global burden of more than 170 million people, many of whom cannot attain the new, expensive, direct-acting antiviral therapies. A safe and effective vaccine that generates both T cell responses and neutralizing antibodies is required to eradicate the disease. Regions within the HCV surface glycoproteins E1 and E2 are essential for virus entry and are targets for neutralizing antibodies. Screening of vaccine candidates requires suitable panels of glycoproteins that represent the breadth of neutralization resistance. Use of a standard reference panel for vaccine studies will ensure comparability of data sets, as has become routine for HIV-1. Here, we describe a large panel of patient-derived HCV glycoproteins with an assessment of their neutralization sensitivity to defined monoclonal antibodies, which has enabled us to predict their likely efficacy in the wider HCV-infected population. The panel could also be important for future selection of additional therapeutic antibodies and for vaccine design.

scholarly journals Identification of a Broadly Cross-Reacting and Neutralizing Human Monoclonal Antibody Directed against the Hepatitis C Virus E2 Protein

2007 ◽  
Vol 82 (2) ◽  
pp. 1047-1052 ◽  
Author(s):  
Mario Perotti ◽  
Nicasio Mancini ◽  
Roberta A. Diotti ◽  
Alexander W. Tarr ◽  
Jonathan K. Ball ◽  
...  
Keyword(s):  
Hepatitis C Virus ◽  
Hepatitis C ◽  
Neutralizing Antibodies ◽  
Human Monoclonal Antibody ◽  
Effective Vaccine ◽  
Human Antibody ◽  
Hcv Genotypes ◽  
Genotype 1A ◽  
E2 Glycoprotein ◽  
Antibody Competition

ABSTRACT Identification of anti-hepatitis C virus (anti-HCV) human antibody clones with broad neutralizing activity is important for a better understanding of the interplay between the virus and host and for the design of an effective passive immunotherapy and an effective vaccine. We report the identification of a human monoclonal Fab (e137) able to bind the HCV E2 glycoprotein of all HCV genotypes but genotype 5. The results of antibody competition assays and testing the reactivity to alanine mutant E2 proteins confirmed that the e137 epitope includes residues (T416, W420, W529, G530, and D535) highly conserved across all HCV genotypes. Fab e137 neutralized HCV pseudoparticles bearing genotype 1a, 1b, and 4 E1-E2 proteins and to a lesser extent, genotype 2b. Fab e137 was also able to inhibit cell culture-grown HCV (genotype 2a). These data indicate that broadly cross-reacting and cross-neutralizing antibodies are generated during HCV infection.

Neutralizing Antibodies Induced by Cell Culture–Derived Hepatitis C Virus Protect Against Infection in Mice

2013 ◽  
Vol 145 (2) ◽  
pp. 447-455.e4 ◽  
Author(s):  
Daisuke Akazawa ◽  
Masaki Moriyama ◽  
Hiroshi Yokokawa ◽  
Noriaki Omi ◽  
Noriyuki Watanabe ◽  
...  
Keyword(s):  
Cell Culture ◽  
Hepatitis C Virus ◽  
Hepatitis C ◽  
Neutralizing Antibodies

scholarly journals Genotypic and Phenotypic Analyses of Hepatitis C Virus from Patients Treated with JTK-853 in a Three-Day Monotherapy

2012 ◽  
Vol 57 (1) ◽  
pp. 436-444 ◽  
Author(s):  
Naoki Ogura ◽  
Yukiyo Toyonaga ◽  
Izuru Ando ◽  
Kunihiro Hirahara ◽  
Tsutomu Shibata ◽  
...  
Keyword(s):  
Hepatitis C Virus ◽  
Hepatitis C ◽  
Amino Acid ◽  
Viral Resistance ◽  
Hcv Rna ◽  
Amino Acid Substitutions ◽  
Sequencing Analysis ◽  
Resistance Mutations ◽  
In The Wild

ABSTRACTJTK-853, a palm site-binding NS5B nonnucleoside polymerase inhibitor, shows antiviral activityin vitroand in hepatitis C virus (HCV)-infected patients. Here, we report the results of genotypic and phenotypic analyses of resistant variants in 24 HCV genotype 1-infected patients who received JTK-853 (800, 1,200, or 1,600 mg twice daily or 1,200 mg three times daily) in a 3-day monotherapy. Viral resistance in NS5B was investigated using HCV RNA isolated from serum specimens from the patients. At the end of treatment (EOT) with JTK-853, the amino acid substitutions M414T (methionine [M] in position 414 at baseline was replaced with threonine [T] at EOT), C445R (cysteine [C] in position 445 at baseline was replaced with arginine [R] at EOT), Y448C/H (tyrosine [Y] in position 448 at baseline was replaced with cysteine [C] or histidine [H] at EOT), and L466F (leucine [L] in position 466 at baseline was replaced with phenylalanine [F] at EOT), which are known to be typical resistant variants of nonnucleoside polymerase inhibitors, were observed in a clonal sequencing analysis. These substitutions were also selected by a treatment with JTK-853in vitro, and the 50% effective concentration of JTK-853 in the M414T-, C445F-, Y448H-, and L466V-harboring replicons attenuated the susceptibility by 44-, 5-, 6-, and 21-fold, respectively, compared with that in the wild-type replicon (Con1). These findings suggest that amino acid substitutions of M414T, C445R, Y448C/H, and L466F are thought to be viral resistance mutations in HCV-infected patients receiving JTK-853 in a 3-day monotherapy.

scholarly journals Hepatitis C virus cell-cell transmission in hepatoma cells in the presence of neutralizing antibodies

Hepatology ◽  
2007 ◽  
Vol 47 (1) ◽  
pp. 17-24 ◽  
Author(s):  
Jennifer M. Timpe ◽  
Zania Stamataki ◽  
Adam Jennings ◽  
Ke Hu ◽  
Michelle J. Farquhar ◽  
...  
Keyword(s):  
Hepatitis C Virus ◽  
Hepatitis C ◽  
Neutralizing Antibodies ◽  
Hepatoma Cells ◽  
Cell Transmission ◽  
Cell Cell

scholarly journals Dissecting Human Antibody Responses: Useful, Basic, and Surprising Findings

Blood ◽  
2017 ◽  
Vol 130 (Suppl_1) ◽  
pp. SCI-49-SCI-49
Author(s):  
Antonio Lanzavecchia
Keyword(s):  
B Cell ◽  
Dna Sequences ◽  
Neutralizing Antibodies ◽  
Influenza A ◽  
Somatic Mutations ◽  
Vaccine Design ◽  
Human Memory ◽  
Affinity Maturation ◽  
Effective Vaccine ◽  
Human Antibody

Abstract We use cell culture-based high-throughput methods to interrogate human memory B cell and plasma cell repertoires and to isolate antibodies selected on the basis of their neutralizing potency and breadth. Relevant examples are antibodies that neutralize all influenza A viruses or even four paramyxoviruses. By targeting conserved structures, these broadly neutralizing antibodies are less prone to select escape mutants and are promising candidates for prophylaxis and therapy of infections, as well as tools for vaccine design. The value of a target-agnostic approach to vaccine design is illustrated by our discovery of extremely potent antibodies that neutralize human cytomegalovirus, which led to the identification of their viral ligand, a pentameric complex that was then produced and tested as an effective vaccine. By reconstructing the genealogy trees of specific B cell clones, we investigate the role of somatic mutations in affinity maturation and in generation of antibody variants with broader or different specificity. Somatic mutations can also generate autoantibodies, as found in patients with pemphigus and autoimmune pulmonary alveolar proteinosis. Recently, while searching for antibodies that broadly react with malaria variant antigens, we discovered a new mechanism of antibody diversification, which relies on templated insertions of genomic DNA sequences into immunoglobulin genes, followed by somatic mutations. Disclosures Lanzavecchia: Humabs SA: Equity Ownership, Research Funding.

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