scholarly journals Hepatitis C Virus (HCV)-Induced Immunoglobulin Hypermutation Reduces the Affinity and Neutralizing Activities of Antibodies against HCV Envelope Protein

2008 ◽  
Vol 82 (13) ◽  
pp. 6711-6720 ◽  
Author(s):  
Keigo Machida ◽  
Yasuteru Kondo ◽  
Jeffrey Y. Huang ◽  
Yung-Chia Chen ◽  
Kevin T.-H. Cheng ◽  
...  
Keyword(s):  
Hepatitis C Virus ◽  
Hepatitis C ◽  
B Cells ◽  
Neutralizing Antibodies ◽  
Immune Surveillance ◽  
Neutralizing Antibody ◽  
Viral Receptor ◽  
Complement Dependent Cytotoxicity ◽  
Protective Antibodies ◽  
Hcv Antibody

ABSTRACT Hepatitis C virus (HCV) often causes persistent infection despite the presence of neutralizing antibodies against the virus in the sera of hepatitis C patients. HCV infects both hepatocytes and B cells through the binding of its envelope glycoprotein E2 to CD81, the putative viral receptor. Previously, we have shown that E2-CD81 interaction induces hypermutation of heavy-chain immunoglobulin (V H ) in B cells. We hypothesize that if HCV infects antibody-producing B cells, the resultant hypermutation of VH may lower the affinity and specificity of the HCV-specific antibodies, enabling HCV to escape from immune surveillance. To test this hypothesis, we infected human hybridoma clones producing either neutralizing or non-neutralizing anti-E2 or anti-E1 antibodies with a lymphotropic HCV (SB strain). All of the hybridoma clones, except for a neutralizing antibody-producing hybridoma, could be infected with HCV and support virus replication for at least 8 weeks after infection. The VH sequences in the infected hybridomas had a significantly higher mutation frequency than those in the uninfected hybridomas, with mutations concentrating in complementarity-determining region 3. These mutations lowered the antibody affinity against the targeting protein and also lowered the virus-neutralizing activity of anti-E2 antibodies. Furthermore, antibody-mediated complement-dependent cytotoxicity with the antibodies secreted from the HCV-infected hybridomas was impaired. These results suggest that HCV infection could cause some anti-HCV-antibody-producing hybridoma B cells to make less-protective antibodies.

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

2016 ◽  
Vol 113 (45) ◽  
pp. 12768-12773 ◽  
Author(s):  
Leopold Kong ◽  
David E. Lee ◽  
Rameshwar U. Kadam ◽  
Tong Liu ◽  
Erick Giang ◽  
...  
Keyword(s):  
Hepatitis C Virus ◽  
Hepatitis C ◽  
Viral Entry ◽  
Neutralizing Antibodies ◽  
Vaccine Design ◽  
Neutralizing Antibody ◽  
Md Simulations ◽  
Receptor Binding Site ◽  
Structural Investigations ◽  
Thermophilic Organisms

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.

scholarly journals A role for B cells to transmit hepatitis C virus infection

2020 ◽  
Author(s):  
Isabelle Desombere ◽  
Freya Van Houtte ◽  
Ali Farhoudi ◽  
Lieven Verhoye ◽  
Caroline Buysschaert ◽  
...  
Keyword(s):  
Hepatitis C Virus ◽  
Hepatitis C ◽  
B Cells ◽  
Neutralizing Antibody ◽  
Hcv Rna ◽  
Serum Samples ◽  
Cell Culture Systems ◽  
Hcv Transmission

Abstract Hepatitis C virus (HCV) is highly variable and transmits through infected blood to establish a chronic liver infection in the majority of patients. Our knowledge of the infectivity of clinical HCV strains is hampered by the lack of in vitro cell culture systems that support efficient viral replication. We previously reported that laboratory strains of HCV associated with non-permissive B cells could trans-infect hepatocytes and thereby evade host neutralizing antibody responses, suggesting a role for B cells in HCV transmission. To evaluate this hypothesis, we assessed the ability of B cells and sera from recent (<2 years) or chronic (≥ 2 years) infections to infect humanized liver chimeric mice. HCV was efficiently transmitted by B cells from chronically infected patients whereas the sera were non-infectious. In contrast, we noted that B cells from recently infected patients failed to transmit HCV to the mice, whereas all serum samples were infectious. Only patients with circulating anti-glycoprotein antibodies harbored genomic HCV-RNA in B cells. Taken together, our studies provide direct in vivo evidence for HCV transmission by B cells and these findings may have clinical implications for prophylactic and therapeutic antibody-based vaccine design.

scholarly journals Binding of the Hepatitis C Virus E2 Glycoprotein to CD81 Is Strain Specific and Is Modulated by a Complex Interplay between Hypervariable Regions 1 and 2

2003 ◽  
Vol 77 (3) ◽  
pp. 1856-1867 ◽  
Author(s):  
RosaMaria Roccasecca ◽  
Helenia Ansuini ◽  
Alessandra Vitelli ◽  
Annalisa Meola ◽  
Elisa Scarselli ◽  
...  
Keyword(s):  
Hepatitis C Virus ◽  
Hepatitis C ◽  
Receptor Binding ◽  
Neutralizing Antibodies ◽  
Intramolecular Interactions ◽  
Amino Acid Residues ◽  
Complex Interplay ◽  
Viral Receptor ◽  
Hypervariable Regions

ABSTRACT The envelope glycoprotein E2 of hepatitis C virus (HCV) is the target of neutralizing antibodies and is presently being evaluated as an HCV vaccine candidate. HCV binds to human cells through the interaction of E2 with the tetraspanin CD81, a putative viral receptor component. We have analyzed four different E2 proteins from 1a and 1b viral isolates for their ability to bind to recombinant CD81 in vitro and to the native receptor displayed on the surface of Molt-4 cells. A substantial difference in binding efficiency between these E2 variants was observed, with proteins derived from 1b subtypes showing significantly lower binding than the 1a protein. To elucidate the mechanism of E2-CD81 interaction and to identify critical regions responsible for the different binding efficiencies of the E2 variants, several mutants were generated in E2 protein regions predicted by computer modeling to be exposed on the protein surface. Functional analysis of these E2 derivatives revealed that at least two distinct domains are responsible for interaction with CD81. A first segment centered around amino acid residues 613 to 618 is essential for recognition, while a second element including the two hypervariable regions (HVRs) modulates E2 receptor binding. Binding inhibition experiments with anti-HVR monoclonal antibodies confirmed this mapping and supported the hypothesis that a complex interplay between the two HVRs of E2 is responsible for modulating receptor binding, possibly through intramolecular interactions. Finally, E2 proteins from different isolates displayed a profile of binding to human hepatic cells different from that observed on Molt-4 cells or isolated recombinant CD81, indicating that additional factors are involved in viral recognition by target liver cells.

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 Broad Anti-Hepatitis C Virus (HCV) Antibody Responses Are Associated with Improved Clinical Disease Parameters in Chronic HCV Infection

2016 ◽  
Vol 90 (9) ◽  
pp. 4530-4543 ◽  
Author(s):  
Rachael E. Swann ◽  
Vanessa M. Cowton ◽  
Mark W. Robinson ◽  
Sarah J. Cole ◽  
Stephen T. Barclay ◽  
...  
Keyword(s):  
Hepatitis C Virus ◽  
Hepatitis C ◽  
Chronic Infection ◽  
Neutralizing Antibodies ◽  
Neutralizing Antibody ◽  
Hcv Infection ◽  
Neutralization Activity ◽  
Broadly Neutralizing Antibody ◽  
Chronic Hcv Infection ◽  
Chronic Hcv

ABSTRACTDuring hepatitis C virus (HCV) infection, broadly neutralizing antibody (bNAb) responses targeting E1E2 envelope glycoproteins are generated in many individuals. It is unclear if these antibodies play a protective or a pathogenic role during chronic infection. In this study, we investigated whether bNAb responses in individuals with chronic infection were associated with differences in clinical presentation. Patient-derived purified serum IgG was used to assess the breadth of HCV E1E2 binding and the neutralization activity of HCV pseudoparticles. The binding and neutralization activity results for two panels bearing viral envelope proteins representing either an intergenotype or an intragenotype 1 group were compared. We found that the HCV load was negatively associated with strong cross-genotypic E1E2 binding (P= 0.03). Overall, we observed only a modest correlation between total E1E2 binding and neutralization ability. The breadth of intergenotype neutralization did not correlate with any clinical parameters; however, analysis of individuals with genotype 1 (gt1) HCV infection (n= 20), using an intragenotype pseudoparticle panel, found a strong association between neutralization breadth and reduced liver fibrosis (P= 0.006). A broad bNAb response in our cohort with chronic infection was associated with a single nucleotide polymorphism (SNP) in theHLA-DQB1gene (P= 0.038), as previously reported in a cohort with acute disease. Furthermore, the bNAbs in these individuals targeted more than one region of E2-neutralizing epitopes, as assessed through cross-competition of patient bNAbs with well-characterized E2 antibodies. We conclude that the bNAb responses in patients with chronic gt1 infection are associated with lower rates of fibrosis and host genetics may play a role in the ability to raise such responses.IMPORTANCEGlobally, there are 130 million to 150 million people with chronic HCV infection. Typically, the disease is progressive and is a major cause of severe liver cirrhosis and hepatocellular carcinoma. While it is known that neutralizing antibodies have a role in spontaneous clearance during acute infection, little is known about their role in chronic infection. In the present work, we investigated the antibody response in a cohort of chronically infected individuals and found that a broadly neutralizing antibody response is protective and is associated with reduced levels of liver fibrosis and cirrhosis. We also found an association between SNPs in class II HLA genes and the presence of a broadly neutralizing response, indicating that antigen presentation may be important for the production of HCV-neutralizing antibodies.

scholarly journals Hepatitis C virus association with peripheral blood B lymphocytes potentiates viral infection of liver-derived hepatoma cells

Blood ◽  
2009 ◽  
Vol 113 (3) ◽  
pp. 585-593 ◽  
Author(s):  
Zania Stamataki ◽  
Claire Shannon-Lowe ◽  
Jean Shaw ◽  
David Mutimer ◽  
Alan B. Rickinson ◽  
...  
Keyword(s):  
Hepatitis C Virus ◽  
Hepatitis C ◽  
B Cells ◽  
B Cell ◽  
Neutralizing Antibodies ◽  
Cd40 Ligand ◽  
Hepatoma Cells ◽  
Productive Infection ◽  
Interleukin 4 ◽  
Extracellular Virus

Abstract Hepatitis C virus (HCV) primarily replicates within the liver, leading to hepatitis, fibrosis, and hepatocellular carcinoma. Infection is also associated with B-cell abnormalities, suggesting an association of the virus with B cells. The infectious JFH-1 strain of HCV can bind primary and immortalized B cells but fails to establish productive infection. However, B cell–associated virus readily infects hepatoma cells, showing an enhanced infectivity compared with extracellular virus. B cells express the viral receptors CD81, SR-BI, and the C-type lectins DC-SIGN and L-SIGN. Antibodies specific for SR-BI and DC-SIGN/L-SIGN reduced B-cell transinfection, supporting a role for these molecules in B-cell association with HCV. Stimulation of B cells with CD40 ligand and interleukin-4 promoted their ability to transinfect hepatoma cells. B cell–associated virus is resistant to trypsin proteolysis and HCV-specific neutralizing antibodies, consistent with particle internalization. HCV promoted the adhesion of primary B cells to Huh-7 hepatomas, providing a mechanism for B-cell retention in the infected liver. In summary, B cells may provide a vehicle for HCV to persist and transmit to the liver.

scholarly journals A Role for B Cells to Transmit Hepatitis C Virus Infection

2021 ◽  
Vol 12 ◽  
Author(s):  
Isabelle Desombere ◽  
Freya Van Houtte ◽  
Ali Farhoudi ◽  
Lieven Verhoye ◽  
Caroline Buysschaert ◽  
...  
Keyword(s):  
Hepatitis C Virus ◽  
Hepatitis C ◽  
B Cells ◽  
Immune Surveillance ◽  
Serum Samples ◽  
Infected Blood ◽  
Cell Culture Systems ◽  
Hcv Transmission ◽  
Host Immune Surveillance

Hepatitis C virus (HCV) is highly variable and transmits through infected blood to establish a chronic liver infection in the majority of patients. Our knowledge on the infectivity of clinical HCV strains is hampered by the lack of in vitro cell culture systems that support efficient viral replication. We and others have reported that HCV can associate with and infect immune cells and may thereby evade host immune surveillance and elimination. To evaluate whether B cells play a role in HCV transmission, we assessed the ability of B cells and sera from recent (&lt;2 years) or chronic (≥ 2 years) HCV patients to infect humanized liver chimeric mice. HCV was transmitted by B cells from chronic infected patients whereas the sera were non-infectious. In contrast, B cells from recently infected patients failed to transmit HCV to the mice, whereas all serum samples were infectious. We observed an association between circulating anti-glycoprotein E1E2 antibodies and B cell HCV transmission. Taken together, our studies provide evidence for HCV transmission by B cells, findings that have clinical implications for prophylactic and therapeutic antibody-based vaccine design.

scholarly journals Antigenicity and Immunogenicity of Differentially Glycosylated Hepatitis C Virus E2 Envelope Proteins Expressed in Mammalian and Insect Cells

2019 ◽  
Vol 93 (7) ◽  
Author(s):  
Richard A. Urbanowicz ◽  
Ruixue Wang ◽  
John E. Schiel ◽  
Zhen-yong Keck ◽  
Melissa C. Kerzic ◽  
...  
Keyword(s):  
Hepatitis C Virus ◽  
Hepatitis C ◽  
Insect Cell ◽  
Neutralizing Antibodies ◽  
Vaccine Development ◽  
Neutralizing Antibody ◽  
Antibody Responses ◽  
Glycosylation Sites ◽  
Complete Deletion ◽  
Global Health Challenge

ABSTRACTThe development of a prophylactic vaccine for hepatitis C virus (HCV) remains a global health challenge. Cumulative evidence supports the importance of antibodies targeting the HCV E2 envelope glycoprotein to facilitate viral clearance. However, a significant challenge for a B cell-based vaccine is focusing the immune response on conserved E2 epitopes capable of eliciting neutralizing antibodies not associated with viral escape. We hypothesized that glycosylation might influence the antigenicity and immunogenicity of E2. Accordingly, we performed head-to-head molecular, antigenic, and immunogenic comparisons of soluble E2 (sE2) produced in (i) mammalian (HEK293) cells, which confer mostly complex- and high-mannose-type glycans; and (ii) insect (Sf9) cells, which impart mainly paucimannose-type glycans. Mass spectrometry demonstrated that all 11 predictedN-glycosylation sites were utilized in both HEK293- and Sf9-derived sE2, but thatN-glycans in insect sE2 were on average smaller and less complex. Both proteins bound CD81 and were recognized by conformation-dependent antibodies. Mouse immunogenicity studies revealed that similar polyclonal antibody responses were generated against antigenic domains A to E of E2. Although neutralizing antibody titers showed that Sf9-derived sE2 induced moderately stronger responses than did HEK293-derived sE2 against the homologous HCV H77c isolate, the two proteins elicited comparable neutralization titers against heterologous isolates. Given that global alteration of HCV E2 glycosylation by expression in different hosts did not appreciably affect antigenicity or overall immunogenicity, a more productive approach to increasing the antibody response to neutralizing epitopes may be complete deletion, rather than just modification, of specificN-glycans proximal to these epitopes.IMPORTANCEThe development of a vaccine for hepatitis C virus (HCV) remains a global health challenge. A major challenge for vaccine development is focusing the immune response on conserved regions of the HCV envelope protein, E2, capable of eliciting neutralizing antibodies. Modification of E2 by glycosylation might influence the immunogenicity of E2. Accordingly, we performed molecular and immunogenic comparisons of E2 produced in mammalian and insect cells. Mass spectrometry demonstrated that the predicted glycosylation sites were utilized in both mammalian and insect cell E2, although the glycan types in insect cell E2 were smaller and less complex. Mouse immunogenicity studies revealed similar polyclonal antibody responses. However, insect cell E2 induced stronger neutralizing antibody responses against the homologous isolate used in the vaccine, albeit the two proteins elicited comparable neutralization titers against heterologous isolates. A more productive approach for vaccine development may be complete deletion of specific glycans in the E2 protein.

scholarly journals Structure of the hepatitis C virus E1E2 glycoprotein complex

2021 ◽  
Author(s):  
Alba Torrents de la Pena ◽  
Kwinten Sliepen ◽  
Lisa Eshun-Wilson ◽  
Maddy Newby ◽  
Joel D. Allen ◽  
...  
Keyword(s):  
Hepatitis C Virus ◽  
Hepatitis C ◽  
Viral Entry ◽  
Neutralizing Antibodies ◽  
Rational Design ◽  
Molecular Mechanisms ◽  
Hydrophobic Interactions ◽  
Neutralizing Antibody ◽  
Broadly Neutralizing Antibodies ◽  
Fusion Glycoprotein

Hepatitis C virus (HCV) infection is a leading cause of chronic liver disease, cirrhosis, and hepatocellular carcinoma in humans, and afflicts more than 58 million people worldwide. The HCV envelope E1 and E2 glycoproteins are essential for viral entry and infection, and comprise the primary antigenic target for neutralizing antibody responses. The molecular mechanisms of E1E2 assembly, as well as how the E1E2 heterodimer binds broadly neutralizing antibodies, remains elusive. We present the cryo-electron microscopy (cryoEM) structure of the membrane-extracted full-length E1E2 heterodimer in complex with broadly neutralizing antibodies (bNAbs) AR4A, AT12009 and IGH505 at ~3.5 Å resolution. We resolve the long sought-after interface between the E1 and E2 ectodomains and reveal how it is stabilized by hydrophobic interactions and glycans. This structure deepens our understanding of the HCV fusion glycoprotein and delivers a blueprint for the rational design of novel vaccine immunogens and anti-viral drugs.

scholarly journals 1065. Hepatitis C Virus Care Cascade Assessment–One Step Closer to Micro-Elimination

2020 ◽  
Vol 7 (Supplement_1) ◽  
pp. S561-S562
Author(s):  
Jehan F Chowdhury ◽  
Anna Winston ◽  
Tanya Zeina ◽  
Hong Gi Shim ◽  
Tine Vindenes
Keyword(s):  
Hepatitis C Virus ◽  
Hepatitis C ◽  
Drug Use ◽  
Sustained Viral Response ◽  
The United States ◽  
World Health ◽  
Viral Response ◽  
Care Cascade ◽  
Care Gaps ◽  
Hcv Antibody

Abstract Background Hepatitis C virus (HCV) is a leading cause of advanced liver disease and death. In the United States about 3.5 million people are living with HCV, but only 50% are aware of the infection, 16% are prescribed treatment, and only 9% achieve sustained viral response. The World Health Organization published an HCV elimination goal for 2030 that strives to achieve a 65% reduction in HCV-related deaths and 90% reduction in transmission. An important step toward this goal is micro-elimination at local hospitals by addressing care gaps in the HCV care cascade. Figure 1 Methods We created a retrospective cohort of patients who tested positive for HCV antibody (HCV Ab+) between 2016 and 2018 at Tufts Medical Center in Boston, Massachusetts. We assessed achievement of care cascade steps including HCV viral load (VL) testing, linkage to care, treatment initiation, and sustained viral response (SVR). We also assessed patient demographics, clinical factors and HCV risk factors. We used STATA/IC 14.1 to conduct bivariate analysis to identify factors associated with loss to follow-up across each care cascade step. Results A total of 24,308 HCV antibody tests were done during this timeframe, of which 5% (n=1,222) were HCV Ab+. After excluding duplicate tests, 1,041 unique patients with HCV Ab+ were included. This cohort had a mean age of 47 years and were 61% male, 66% white, 72% on public insurance, 12% HIV-positive, 13% HCV treatment-experienced. The most frequent HCV risk factor was injection drug use, occurring in 64% of patients. Of patients with HCV Ab+, 76% (n=791) were tested for an HCV VL, of which 50% (n=393) had detectable VL and 50% (n=398) had undetectable VL. Of the patients with a detectable VL, 58% (n=226) were linked with care. Following care linkage, 69% (n=155) initiated treatment, of which 90% (n=139) completed treatment, of which 97% (n=135) achieved SVR (Figure 1). Factors that were significantly associated with getting a VL test and linking to care included private insurance, HIV co-infection, absence of intravenous drug use and cirrhosis; however, these factors were not significantly associated with achieving subsequent steps. Conclusion Assessment of the HCV care cascade at our hospital allowed us to identify clear care gaps and areas needing improvement towards a local micro-elimination. Disclosures All Authors: No reported disclosures

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