scholarly journals Evolution of a Cell Culture-Derived Genotype 1a Hepatitis C Virus (H77S.2) during Persistent Infection with Chronic Hepatitis in a Chimpanzee

2014 ◽  
Vol 88 (7) ◽  
pp. 3678-3694 ◽  
Author(s):  
M. Yi ◽  
F. Hu ◽  
M. Joyce ◽  
V. Saxena ◽  
C. Welsch ◽  
...  
Keyword(s):  
Cell Culture ◽  
Chronic Hepatitis ◽  
Hepatitis C Virus ◽  
Hepatitis C ◽  
Persistent Infection ◽  
Genotype 1A ◽  
A Cell

scholarly journals Robust production of infectious viral particles in Huh-7 cells by introducing mutations in hepatitis C virus structural proteins

2007 ◽  
Vol 88 (9) ◽  
pp. 2495-2503 ◽  
Author(s):  
David Delgrange ◽  
André Pillez ◽  
Sandrine Castelain ◽  
Laurence Cocquerel ◽  
Yves Rouillé ◽  
...  
Keyword(s):  
Cell Culture ◽  
Hepatitis C Virus ◽  
Hepatitis C ◽  
Core Protein ◽  
Potential Effect ◽  
Structural Proteins ◽  
Wild Type ◽  
Genotype 1A ◽  
Viral Particles ◽  
A Cell

Recently, the characterization of a cell culture system allowing the amplification of an authentic virus, named hepatitis C virus cell culture (HCVcc), has been reported by several groups. To obtain higher HCV particle productions, we investigated the potential effect of some amino acid changes on the infectivity of the JFH-1 isolate. As a first approach, successive infections of naïve Huh-7 cells were performed until high viral titres were obtained, and mutations that appeared during this selection were identified by sequencing. Only one major modification, N534K, located in the E2 glycoprotein sequence was found. Interestingly, this mutation prevented core glycosylation of E2 site 6. In addition, JFH-1 generated with this modification facilitated the infection of Huh-7 cells. In a second approach to identify mutations favouring HCVcc infectivity, we exploited the observation that a chimeric virus containing the genotype 1a core protein in the context of JFH-1 background was more infectious than wild-type JFH-1 isolate. Sequence alignment between JFH-1 and our chimera, led us to identify two major positions, 172 and 173, which were not occupied by similar amino acids in these two viruses. Importantly, higher viral titres were obtained by introducing these residues in the context of wild-type JFH-1. Altogether, our data indicate that a more robust production of HCVcc particles can be obtained by introducing a few specific mutations in JFH-1 structural proteins.

scholarly journals NS3 Variability in Hepatitis C Virus Genotype 1A Isolates from Liver Tissue and Serum Samples of Treatment-Naïve Patients with Chronic Hepatitis C

Intervirology ◽  
2018 ◽  
Vol 61 (1) ◽  
pp. 1-8
Author(s):  
Deborah D’Aliberti ◽  
Irene Cacciola ◽  
Cristina Musolino ◽  
Giuseppina Raffa ◽  
Roberto Filomia ◽  
...  
Keyword(s):  
Chronic Hepatitis ◽  
Hepatitis C Virus ◽  
Hepatitis C ◽  
Chronic Hepatitis C ◽  
Liver Tissue ◽  
Serum Samples ◽  
Virus Genotype ◽  
Genotype 1A ◽  
Treatment Naïve

scholarly journals Identification of Hepatitis C Virus NS5A Inhibitors

2009 ◽  
Vol 84 (1) ◽  
pp. 482-491 ◽  
Author(s):  
Julie A. Lemm ◽  
Donald O'Boyle ◽  
Mengping Liu ◽  
Peter T. Nower ◽  
Richard Colonno ◽  
...  
Keyword(s):  
Hepatitis C Virus ◽  
Hepatitis C ◽  
Amino Acid ◽  
Therapeutic Index ◽  
Dna Viruses ◽  
Genotype 1A ◽  
A Cell ◽  
Ns3 Protease ◽  
Derivatives Of ◽  
Rna And Dna

ABSTRACT Using a cell-based replicon screen, we identified a class of compounds with a thiazolidinone core structure as inhibitors of hepatitis C virus (HCV) replication. The concentration of one such compound, BMS-824, that resulted in a 50% inhibition of HCV replicon replication was ∼5 nM, with a therapeutic index of >10,000. The compound showed good specificity for HCV, as it was not active against several other RNA and DNA viruses. Replicon cells resistant to BMS-824 were isolated, and mutations were identified. A combination of amino acid substitutions of leucine to valine at residue 31 (L31V) and glutamine to leucine at residue 54 (Q54L) in NS5A conferred resistance to this chemotype, as did a single substitution of tyrosine to histidine at amino acid 93 (Y93H) in NS5A. To further explore the region(s) of NS5A involved in inhibitor sensitivity, genotype-specific NS5A inhibitors were used to evaluate a series of genotype 1a/1b hybrid replicons. Our results showed that, consistent with resistance mapping, the inhibitor sensitivity domain also mapped to the N terminus of NS5A, but it could be distinguished from the key resistance sites. In addition, we demonstrated that NS5A inhibitors, as well as an active-site inhibitor that specifically binds NS3 protease, could block the hyperphosphorylation of NS5A, which is believed to play an essential role in the viral life cycle. Clinical proof of concept has recently been achieved with derivatives of these NS5A inhibitors, indicating that small molecules targeting a nontraditional viral protein like NS5A, without any known enzymatic activity, can also have profound antiviral effects on HCV-infected subjects.

scholarly journals Mutations Identified in the Hepatitis C Virus (HCV) Polymerase of Patients with Chronic HCV Treated with Ribavirin Cause Resistance and Affect Viral Replication Fidelity

2020 ◽  
Vol 64 (12) ◽  
Author(s):  
Niels Mejer ◽  
Ulrik Fahnøe ◽  
Andrea Galli ◽  
Santseharay Ramirez ◽  
Ola Weiland ◽  
...  
Keyword(s):  
Cell Culture ◽  
Chronic Hepatitis ◽  
Hepatitis C Virus ◽  
Hepatitis C ◽  
Chronic Hepatitis C ◽  
Prolonged Exposure ◽  
Resistance Mutations ◽  
Entry Site ◽  
Chronic Hcv ◽  
Genotype 3A

ABSTRACT Ribavirin has been used for 25 years to treat patients with chronic hepatitis C virus (HCV) infection; however, its antiviral mechanism of action remains unclear. Here, we studied virus evolution in a subset of samples from a randomized 24-week trial of ribavirin monotherapy versus placebo in chronic HCV patients, as well as the viral resistance mechanisms of the observed ribavirin-associated mutations in cell culture. Thus, we performed next-generation sequencing of the full-length coding sequences of HCV recovered from patients at weeks 0, 12, 20, 32 and 40 and analyzed novel single nucleotide polymorphisms (SNPs), diversity, and mutation-linkage. At week 20, increased genetic diversity was observed in 5 ribavirin-treated compared to 4 placebo-treated HCV patients due to new synonymous SNPs, particularly G-to-A and C-to-U ribavirin-associated transitions. Moreover, emergence of 14 nonsynonymous SNPs in HCV nonstructural 5B (NS5B) occurred in treated patients, but not in placebo controls. Most substitutions located close to the NS5B polymerase nucleotide entry site. Linkage analysis showed that putative resistance mutations were found in the majority of genomes in ribavirin-treated patients. Identified NS5B mutations from genotype 3a patients were further introduced into the genotype 3a cell-culture-adapted DBN strain for studies in Huh7.5 cells. Specific NS5B substitutions, including DBN-D148N+I363V, DBN-A150V+I363V, and DBN-T227S+S183P, conferred resistance to ribavirin in long-term cell culture treatment, possibly by reducing the HCV polymerase error rate. In conclusion, prolonged exposure of HCV to ribavirin in chronic hepatitis C patients induces NS5B resistance mutations leading to increased polymerase fidelity, which could be one mechanism for ribavirin resistance.

scholarly journals Serum amyloid A has antiviral activity against hepatitis C virus by inhibiting virus entry in a cell culture system

Hepatology ◽  
2006 ◽  
Vol 44 (6) ◽  
pp. 1626-1634 ◽  
Author(s):  
Muriel Lavie ◽  
Cécile Voisset ◽  
Ngoc Vu-Dac ◽  
Virginie Zurawski ◽  
Gilles Duverlie ◽  
...  
Keyword(s):  
Cell Culture ◽  
Hepatitis C Virus ◽  
Hepatitis C ◽  
Antiviral Activity ◽  
Culture System ◽  
Serum Amyloid A ◽  
Virus Entry ◽  
Cell Culture System ◽  
Amyloid A ◽  
A Cell

scholarly journals Efficient Replication of Hepatitis C Virus Genotype 1a RNAs in Cell Culture

2003 ◽  
Vol 77 (5) ◽  
pp. 3181-3190 ◽  
Author(s):  
Keril J. Blight ◽  
Jane A. McKeating ◽  
Joseph Marcotrigiano ◽  
Charles M. Rice
Keyword(s):  
Cell Culture ◽  
Hepatitis C Virus ◽  
Hepatitis C ◽  
Rna Replication ◽  
Full Length ◽  
Hcv Rna ◽  
Metabolic Labeling ◽  
Virus Genotype ◽  
Adaptive Mutations ◽  
Genotype 1A

ABSTRACT Hepatitis C virus (HCV) genotype 1 (subtypes 1a and 1b) is responsible for the majority of treatment-resistant liver disease worldwide. Thus far, efficient HCV RNA replication has been observed only for subgenomic and full-length RNAs derived from genotype 1b isolates. Here, we report the establishment of efficient RNA replication systems for genotype 1a strain H77. Replication of subgenomic and full-length H77 1a RNAs required the highly permissive Huh-7.5 hepatoma subline and adaptive amino acid substitutions in both NS3 and NS5A. Replication could be detected by RNA quantification, fluorescence-activated cell sorting, and metabolic labeling of HCV-specific proteins. Replication efficiencies were similar for subgenomic and full-length RNAs and were most efficient for HCV RNAs lacking heterologous RNA elements. Interestingly, both subtype 1a and 1b NS3 adaptive mutations are surface exposed and present on only one face of the NS3 structure. The cell culture-adapted subtype 1a replicons should be useful for basic replication studies and for antiviral development. These results are also encouraging for the development of adapted replicons for the remaining HCV genotypes.

scholarly journals Baseline Hepatitis C Virus (HCV) NS3 Polymorphisms and Their Impact on Treatment Response in Clinical Studies of the HCV NS3 Protease Inhibitor Faldaprevir

2013 ◽  
Vol 58 (2) ◽  
pp. 698-705 ◽  
Author(s):  
Kristi L. Berger ◽  
Ibtissem Triki ◽  
Mireille Cartier ◽  
Martin Marquis ◽  
Marie-Josée Massariol ◽  
...  
Keyword(s):  
Chronic Hepatitis ◽  
Hepatitis C Virus ◽  
Hepatitis C ◽  
Amino Acid ◽  
Protease Inhibitor ◽  
Clinical Studies ◽  
Phase 2 ◽  
Genotype 1A ◽  
Treatment Naïve ◽  
Hcv Protease

ABSTRACTA challenge to the treatment of chronic hepatitis C with direct-acting antivirals is the emergence of drug-resistant hepatitis C virus (HCV) variants. HCV with preexisting polymorphisms that are associated with resistance to NS3/4A protease inhibitors have been detected in patients with chronic hepatitis C. We performed a comprehensive pooled analysis from phase 1b and phase 2 clinical studies of the HCV protease inhibitor faldaprevir to assess the population frequency of baseline protease inhibitor resistance-associated NS3 polymorphisms and their impact on response to faldaprevir treatment. A total of 980 baseline NS3 sequences were obtained (543 genotype 1b and 437 genotype 1a sequences). Substitutions associated with faldaprevir resistance (at amino acid positions 155 and 168) were rare (<1% of sequences) and did not compromise treatment response: in a phase 2 study in treatment-naive patients, six patients had faldaprevir resistance-associated polymorphisms at baseline, of whom five completed faldaprevir-based treatment and all five achieved a sustained virologic response 24 weeks after the end of treatment (SVR24). Among 13 clinically relevant amino acid positions associated with HCV protease resistance, the greatest heterogeneity was seen at NS3 codons 132 and 170 in genotype 1b, and the most common baseline substitution in genotype 1a was Q80K (99/437 [23%]). The presence of the Q80K variant did not reduce response rates to faldaprevir-based treatment. Across the three phase 2 studies, there was no significant difference in SVR24 rates between patients with genotype 1a Q80K HCV and those without Q80K HCV, whether treatment experienced (17% compared to 26%;P= 0.47) or treatment naive (62% compared to 66%;P= 0.72).

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