Recent Advances in Our Understanding of Receptor Binding, Viral Fusion and Cell Entry of Hepatitis C Virus: New Targets for the Design of Antiviral Agents

2007 ◽  
Vol 18 (4) ◽  
pp. 169-189 ◽  
Author(s):  
Pantelis Poumbourios ◽  
Heidi E Drummer
Keyword(s):  
Cell Culture ◽  
Hepatitis C Virus ◽  
Hepatitis C ◽  
Receptor Binding ◽  
Viral Entry ◽  
Antiviral Agents ◽  
Genome Replication ◽  
Type I ◽  
Viral Fusion ◽  
Multiple Drugs

Improvements to antiviral therapies for the treatment of hepatitis C virus (HCV) infections will require the use of multiple drugs that target viral proteins essential for replication. The discovery of anti-HCV compounds has been severely hampered by the lack of cell culture replication systems. Since the late 1990s, the advent of sub-genomic replicons that model the intracellular events leading to HCV genome replication have enabled the discovery of HCV protease and polymerase inhibitors, but did not allow the study of HCV entry or entry inhibitors. More recently, retroviral pseudotyping of the viral glycoproteins and the development of a cell culture-based system that recapitulates the entire HCV replication cycle were achieved. These new experimental systems have enabled a rapid advance in our knowledge of how HCV glycoproteins, E1 and E2, mediate receptor binding and viral entry. These systems have facilitated the discovery of a range of viral receptors. Evidence is emerging that CD81, scavenger receptor class B type I, claudin-1 and the low-density lipoprotein receptor are involved in viral entry. In addition, DC-SIGN and L-SIGN may function to internalize virus into dendritic or endothelial cells, facilitating the transport of virions to sites of infection such as the liver. This review focuses on the interaction between the HCV glycoproteins and cellular receptors, and our current understanding of the viral entry pathway. In addition, key questions on the role that these receptors play in viral entry are raised and potential avenues for the discovery of new antiviral agents are highlighted.

Distinct roles in folding, CD81 receptor binding and viral entry for conserved histidine residues of hepatitis C virus glycoprotein E1 and E2

2012 ◽  
Vol 443 (1) ◽  
pp. 85-94 ◽  
Author(s):  
Irene Boo ◽  
Kevin teWierik ◽  
Florian Douam ◽  
Dimitri Lavillette ◽  
Pantelis Poumbourios ◽  
...  
Keyword(s):  
Hepatitis C Virus ◽  
Hepatitis C ◽  
Membrane Fusion ◽  
Receptor Binding ◽  
Viral Entry ◽  
Virus Entry ◽  
Ph Sensitive ◽  
Viral Fusion ◽  
Histidine Residues ◽  
Virus Glycoprotein

The protonation of histidine in acidic environments underpins its role in regulating the function of pH-sensitive proteins. For pH-sensitive viral fusion proteins, histidine protonation in the endosome leads to the activation of their membrane fusion function. The HCV (hepatitis C virus) glycoprotein E1–E2 heterodimer mediates membrane fusion within the endosome, but the roles of conserved histidine residues in the formation of a functional heterodimer and in sensing pH changes is unknown. We examined the functional roles of conserved histidine residues located within E1 and E2. The E1 mutations, H222A/R, H298R and H352A, disrupted E1–E2 heterodimerization and reduced virus entry. A total of five out of six histidine residues located within the E2 RBD (receptor-binding domain) were important for the E2 fold, and their substitution with arginine or alanine caused aberrant heterodimerization and/or CD81 binding. Distinct roles in E1–E2 heterodimerization and in virus entry were identified for His691 and His693 respectively within the membrane-proximal stem region. Viral entry and cell–cell fusion at neutral and low pH values were enhanced with H445R, indicating that the protonation state of His445 is a key regulator of HCV fusion. However, H445R did not overcome the block to virus entry induced by bafilomycin A1, indicating a requirement for an endosomal activation trigger in addition to acidic pH.

Inhibitory Effects of Amentoflavone and Orobol on Daclatasvir-Induced Resistance-Associated Variants of Hepatitis C Virus

2018 ◽  
Vol 46 (04) ◽  
pp. 835-852 ◽  
Author(s):  
Wei-Ping Lee ◽  
Keng-Li Lan ◽  
Shi-Xian Liao ◽  
Yi-Hsiang Huang ◽  
Ming-Chih Hou ◽  
...  
Keyword(s):  
Hepatitis C Virus ◽  
Hepatitis C ◽  
Viral Entry ◽  
Antiviral Agents ◽  
Cost Effective ◽  
Pegylated Interferon ◽  
Chinese Herbs ◽  
Inhibitory Effects ◽  
Direct Acting Antiviral ◽  
Direct Acting

Hepatitis C virus (HCV) is recognized as a major causative agent of chronic hepatitis, cirrhosis, and hepatocellular carcinoma. Despite rapid progress in the development of direct-acting antivirals (DAA) against HCV infection in recent years, cost-effective antiviral drugs with more affordable prices still need to be developed. In this study, we screened a library of natural compounds to identify natural HCV inhibitors. The library of the pure compounds extracted from Chinese herbs deposited in the chemical bank of National Research Institute of Chinese Medicine (NRICM), Taiwan was screened in the cell culture-derived HCV (HCVcc) system. We identified the flavone or flavan-based compounds amentoflavone, 7,4[Formula: see text]-dihydroxyflavanone, and orobol with the inhibition of viral entry, replication, and translation of the HCV life cycle. Amentoflavone and orobol also showed inhibitory effects on resistant-associated variants to the NS5A inhibitor daclatasvir. The results of this study have the potential to benefit patients who are intolerant to the adverse effect of pegylated interferon or who harbor resistant strains refractory to treatment by current direct-acting antiviral agents.

scholarly journals Functional Analysis of Hepatitis C Virus Envelope Proteins, Using a Cell-Cell Fusion Assay

2006 ◽  
Vol 80 (4) ◽  
pp. 1817-1825 ◽  
Author(s):  
Mariko Kobayashi ◽  
Michael C. Bennett ◽  
Theodore Bercot ◽  
Ila R. Singh
Keyword(s):  
Hepatitis C Virus ◽  
Hepatitis C ◽  
Cell Fusion ◽  
Viral Entry ◽  
Antiviral Agents ◽  
Cell Types ◽  
Envelope Proteins ◽  
Virus Envelope ◽  
Host Cell Membrane ◽  
Cell Cell

ABSTRACT Hepatitis C virus (HCV) envelope proteins mediate the entry of virus into cells by binding to cellular receptors, resulting in fusion of the viral membrane with the host cell membrane and permitting the viral genome to enter the cytoplasm. We report the development of a robust and reproducible cell-cell fusion assay using envelope proteins from commonly occurring genotypes of HCV. The assay scored HCV envelope protein-mediated fusion by the production of fluorescent green syncytia and allowed us to elucidate many aspects of HCV fusion, including the pH of fusion, cell types that permit viral entry, and the conformation of envelope proteins essential for fusion. We found that fusion could be specifically inhibited by anti-HCV antibodies and by at least one peptide. We also generated a number of insertional mutations in the envelope proteins and tested nine of these using the fusion assay. We demonstrate that this fusion assay is a powerful tool for understanding the mechanism of HCV-mediated fusion, elucidating mutant function, and testing antiviral agents.

scholarly journals Selective Inhibition of Hepatitis C Virus Infection by Hydroxyzine and Benztropine

2014 ◽  
Vol 58 (6) ◽  
pp. 3451-3460 ◽  
Author(s):  
Lidia Mingorance ◽  
Martina Friesland ◽  
Mairene Coto-Llerena ◽  
Sofía Pérez-del-Pulgar ◽  
Loreto Boix ◽  
...  
Keyword(s):  
Cell Culture ◽  
Hepatitis C Virus ◽  
Hepatitis C ◽  
Antiviral Activity ◽  
Viral Entry ◽  
Selective Inhibition ◽  
Chemical Library ◽  
Effective Components ◽  
Severe Liver Disease ◽  
Cell Culture Assay

ABSTRACTHepatitis C virus (HCV) infection is a major biomedical problem worldwide as it causes severe liver disease in millions of humans around the world. Despite the recent approval of specific drugs targeting HCV replication to be used in combination with alpha interferon (IFN-α) and ribavirin, there is still an urgent need for pangenotypic, interferon-free therapies to fight this genetically diverse group of viruses. In this study, we used an unbiased screening cell culture assay to interrogate a chemical library of compounds approved for clinical use in humans. This system enables identifying nontoxic antiviral compounds targeting every aspect of the viral life cycle, be the target viral or cellular. The aim of this study was to identify drugs approved for other therapeutic applications in humans that could be effective components of combination therapies against HCV. As a result of this analysis, we identified 12 compounds with antiviral activity in cell culture, some of which had previously been identified as HCV inhibitors with antiviral activity in cell culture and had been shown to be effective in patients. We selected two novel HCV antivirals, hydroxyzine and benztropine, to characterize them by determining their specificity and genotype spectrum as well as by defining the step of the replication cycle targeted by these compounds. We found that both compounds effectively inhibited viral entry at a postbinding step of genotypes 1, 2, 3, and 4 without affecting entry of other viruses.

scholarly journals Screening and Rational Design of Hepatitis C Virus Entry Inhibitory Peptides Derived from GB Virus A NS5A

2012 ◽  
Vol 87 (3) ◽  
pp. 1649-1657 ◽  
Author(s):  
Xiuying Liu ◽  
Yibing Huang ◽  
Min Cheng ◽  
Ling Pan ◽  
Youhui Si ◽  
...  
Keyword(s):  
Cell Culture ◽  
Hepatitis C Virus ◽  
Hepatitis C ◽  
Viral Entry ◽  
Rational Design ◽  
Antiviral Drug ◽  
Inhibitory Effect ◽  
Content Type ◽  
Peptide Helicity

ABSTRACTChronic infection by hepatitis C virus (HCV) is a cause of the global burden of liver diseases. HCV entry into hepatocytes is a complicated and multistep process that represents a promising target for antiviral intervention. The recently reported amphipathic α-helical virucidal peptide (C5A) from the HCV NS5A protein suggests a new category of antiviral drug candidates. In this study, to identify C5A-like HCV inhibitors, synthetic peptides derived from the C5A-corresponding NS5 protein region of selectedFlaviviridaeviruses were evaluated for their anti-HCV activities. A peptide from GB virus A (GBV-A), but not other flaviviruses, demonstrated an inhibitory effect on HCV infection. Through a series of sequence optimizations and modifications of the peptide helicity and hydrophobicity, we obtained a peptide designated GBVA10-9 with highly potent anti-HCV activity. GBVA10-9 suppressed infection with both cell culture-derived and pseudotyped HCVin vitro, and the 50% cell culture inhibitory concentration ranged from 20 nM to 160 nM, depending on the genotypic origin of the envelope proteins. GBVA10-9 had no detectable effects on either HCV attachment to Huh7.5.1 cells or viral RNA replication. No virucidal activity was found with GBVA10-9, suggesting an action mechanism distinct from that of C5A. The inhibitory effect of GBVA10-9 appeared to occur at the postbinding step during viral entry. Taken together, the results with GBVA10-9 demonstrated a potent activity for blocking HCV entry that might be used in combination with other antivirals directly targeting virus-encoded enzymes. Furthermore, GBVA10-9 also provides a novel tool to dissect the detailed mechanisms of HCV entry.

scholarly journals Hepatitis C virus entry: potential receptors and their biological functions

2006 ◽  
Vol 87 (5) ◽  
pp. 1075-1084 ◽  
Author(s):  
Laurence Cocquerel ◽  
Cécile Voisset ◽  
Jean Dubuisson
Keyword(s):  
Cell Culture ◽  
Hepatitis C Virus ◽  
Hepatitis C ◽  
Low Density Lipoprotein ◽  
Heparan Sulphate ◽  
Scavenger Receptor ◽  
Density Lipoprotein ◽  
Type I ◽  
Major Step ◽  
Class B

Several cellular molecules have been identified as putative receptors for Hepatitis C virus (HCV): CD81 tetraspanin, scavenger receptor class B type I (SR-BI), mannose-binding lectins DC-SIGN and L-SIGN, low-density lipoprotein receptor, heparan sulphate proteoglycans and the asialoglycoprotein receptor. Due to difficulties in propagating HCV in cell culture, most of these molecules have been identified by analysing their interaction with a soluble, truncated form of HCV glycoprotein E2. A recent major step in investigating HCV entry was the development of pseudoparticles (HCVpp), consisting of unmodified HCV envelope glycoproteins assembled onto retroviral core particles. This system has allowed the investigation of the role of candidate receptors in the early steps of the HCV life cycle and the data obtained can now be confirmed with the help of a newly developed cell-culture system that allows efficient amplification of HCV (HCVcc). Interestingly, CD81 and SR-BI have been shown to play direct roles in HCVpp and/or HCVcc entry. However, co-expression of CD81 and SR-BI in non-hepatic cell lines does not lead to HCVpp entry, indicating that other molecule(s), expressed only in hepatic cells, are necessary for HCV entry. In this review, the molecules that have been proposed as potential HCV receptors are described and the experimental data indicating that CD81 and SR-BI are potentially involved in HCV entry are presented.

scholarly journals A Novel Small Molecule Inhibits Hepatitis C Virus Propagation in Cell Culture

2021 ◽  
Author(s):  
Ahmed K. Oraby ◽  
Cassandra L. Gardner ◽  
Robert F. Needle ◽  
Hassan M. Kofahi ◽  
Kylie R. Everard ◽  
...  
Keyword(s):  
Cell Culture ◽  
Hepatitis C Virus ◽  
Chronic Disease ◽  
Hepatitis C ◽  
Drug Discovery ◽  
Small Molecule ◽  
Antiviral Agents ◽  
Antiviral Drugs ◽  
Virus Propagation ◽  
Novel Virus

The discovery of curative antiviral drugs for a chronic disease such as HCV infection has encouraged drug discovery in the context of other viruses for which no curative drugs currently exist. Since we currently face a novel virus that has caused a pandemic, the need for new antiviral agents is more apparent than ever.

O.192 Scavenger receptor class B type I is a co-receptor for infection with cell culture-derived hepatitis C virus

2006 ◽  
Vol 36 ◽  
pp. S58
Author(s):  
M.B. Zeisel ◽  
E.K. Schnober ◽  
A. Haberstroh ◽  
D. Lavillette ◽  
F. Cosset ◽  
...  
Keyword(s):  
Cell Culture ◽  
Hepatitis C Virus ◽  
Hepatitis C ◽  
Scavenger Receptor ◽  
Type I ◽  
Scavenger Receptor Class ◽  
Class B

scholarly journals A Conserved Gly436-Trp-Leu-Ala-Gly-Leu-Phe-Tyr Motif in Hepatitis C Virus Glycoprotein E2 Is a Determinant of CD81 Binding and Viral Entry

2006 ◽  
Vol 80 (16) ◽  
pp. 7844-7853 ◽  
Author(s):  
Heidi E. Drummer ◽  
Irene Boo ◽  
Anne L. Maerz ◽  
Pantelis Poumbourios
Keyword(s):  
Hepatitis C Virus ◽  
Hepatitis C ◽  
Receptor Binding ◽  
Viral Entry ◽  
Conformational Transition ◽  
Site Directed Mutagenesis ◽  
Wild Type ◽  
Binding Ability ◽  
Group 2 ◽  
Cd81 Binding

ABSTRACT The hepatitis C virus (HCV) glycoproteins E1 and E2 form a heterodimer that mediates CD81 receptor binding and viral entry. In this study, we used site-directed mutagenesis to examine the functional role of a conserved G436WLAGLFY motif of E2. The mutants could be placed into two groups based on the ability of mature virion-incorporated E1E2 to bind the large extracellular loop (LEL) of CD81 versus the ability to mediate cellular entry of pseudotyped retroviral particles. Group 1 comprised E2 mutants where LEL binding ability largely correlated with viral entry ability, with conservative and nonconservative substitutions (W437 L/A, L438A, L441V/F, and F442A) inhibiting both functions. These data suggest that Trp-437, Leu-438, Leu-441, and Phe-442 directly interact with the LEL. Group 2 comprised E2 glycoproteins with more conservative substitutions that lacked LEL binding but retained between 20% and 60% of wild-type viral entry competence. The viral entry competence displayed by group 2 mutants was explained by residual binding by the E2 receptor binding domain to cellular full-length CD81. A subset of mutants maintained LEL binding ability in the context of intracellular E1E2 forms, but this function was largely lost in virion-incorporated glycoproteins. These data suggest that the CD81 binding site undergoes a conformational transition during glycoprotein maturation through the secretory pathway. The G436P mutant was an outlier, retaining near-wild-type levels of CD81 binding but lacking significant viral entry ability. These findings indicate that the G436WLAGLFY motif of E2 functions in CD81 binding and in pre- or post-CD81-dependent stages of viral entry.

scholarly journals Small Interfering RNA Targeted to Hepatitis C Virus 5′ NontranslatedRegion Exerts Potent Antiviral Effect

2006 ◽  
Vol 81 (2) ◽  
pp. 669-676 ◽  
Author(s):  
Tatsuo Kanda ◽  
Robert Steele ◽  
Ranjit Ray ◽  
Ratna B. Ray
Keyword(s):  
Cell Culture ◽  
Hepatitis C Virus ◽  
Hepatitis C ◽  
Virus Replication ◽  
Hcv Infection ◽  
Therapeutic Modality ◽  
Genome Replication ◽  
Entry Site ◽  
Hcv Genotype ◽  
Conserved Sequence

ABSTRACT Hepatitis C virus (HCV) is a major cause of cirrhosis and hepatocellular carcinoma. Interferon alone or together with ribavirin is the only therapy for HCV infection; however, a significant number of HCV-infected individuals do not respond to this treatment. Therefore, the development of new therapeutic options against HCV is a matter of urgency. In the present study, we have examined vectors carrying short hairpin RNA (shRNA) targeting the 5′ nontranslated conserved region of the HCV genome for inhibition of virus replication. Initially, three sequences were selected, and all three shRNAs (psh-53, psh-274, and psh-375) suppressed HCV internal ribosome entry site (IRES)-mediated translation to different degrees in Huh-7 cells. Next, we introduced siRNA into Huh-7.5 cells persistently infected with HCV genotype 2a (JFH1). The most efficient inhibition of JFH1 replication was observed with psh-274, targeted to the portion from subdomain IIId to IIIe of the IRES. Subsequently, Huh-7.5 cells stably expressing psh-274 further displayed a significant reduction in HCV JFH1 replication. The effect of psh-274 on cell-culture-grown HCV genotype 1a (H77) was also evaluated, and inhibition of virus replication and infectivity titers was observed. In the absence of a cell-culture-grown HCV genotype 1b, the effects of psh-274 on subgenomic and full-length replicons were examined, and efficient inhibition of genome replication was observed. Therefore, we have identified a conserved sequence targeted to the HCV genome that can inhibit replication of different genotypes, suggesting the potential of siRNA as an additional therapeutic modality against HCV infection.

Sign in / Sign up

Export Citation Format

Share Document