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Kawaguchi T, Komori A, Fujisaki K, Nishiguchi S, Kato M, Takagi H, Tanaka Y, Notsumata K, Mita E, Nomura H, Shibatoge M, Takaguchi K, Hattori T, Sata M, Koike K. Eltrombopag enables initiation and completion of pegylated interferon/ribavirin therapy in Japanese HCV-infected patients with chronic liver disease and thrombocytopenia. Exp Ther Med 2019; 18:596-604. [PMID: 31258695 PMCID: PMC6566053 DOI: 10.3892/etm.2019.7616] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/21/2018] [Accepted: 02/27/2019] [Indexed: 11/30/2022] Open
Abstract
To investigate the efficacy of eltrombopag for the treatment of thrombocytopenia in patients with chronic hepatitis C, a phase II, single-arm, open-label study with a 9-week pre-antiviral phase was conducted, followed by a 48-week antiviral phase and a 24-week follow-up phase. The proportion of patients who achieved a platelet count threshold, the proportion of patients who maintained a platelet count >50,000/µl, sustained virological response (SVR) rates and safety parameters were evaluated. Of the 45 enrolled patients (median age, 59 years; median platelet count, 63,000/µl; 98% with Child-Pugh class A), 43 (96%) achieved the platelet count threshold during the pre-antiviral phase. A total of 13 patients (29%) experienced ≥1 adverse event (AE), of which headache and vomiting were the most common, and 41 patients (mostly receiving eltrombopag 12.5 mg or 25 mg) entered the antiviral phase, of which 36 (88%) maintained the platelet count threshold; no patient platelet count decreased below 25,000/µl. Nine patients (22%) achieved an SVR at the 24-week follow-up. Grade ≥3 AEs occurred in 25 patients (61%). A total of 8 serious AEs occurred in five patients (12%). No mortality, thromboembolic events (TEEs), or cataract progression were reported. Eltrombopag increased the platelet count in chronic hepatitis C virus-infected patients with cirrhosis and thrombocytopenia and enabled them to initiate and complete interferon-based antiviral therapy (NCT01636778; first submitted: July 05, 2012).
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Affiliation(s)
- Takumi Kawaguchi
- Department of Medicine, Division of Gastroenterology, Kurume University School of Medicine, Kurume, Fukuoka 830-0011, Japan
| | - Atsumasa Komori
- Clinical Research Center, National Hospital Organization (NHO) Nagasaki Medical Center, Omura, Nagasaki 856-8562, Japan.,Department of Hepatology, Nagasaki University Graduate School of Biomedical Sciences, Nagasaki 852-8523, Japan
| | - Kunio Fujisaki
- Kirishima Medical Center, Kirishima, Kagoshima 899-5112, Japan
| | - Shuhei Nishiguchi
- Division of Hepatobiliary and Pancreatic Disease, Hyogo College of Medicine, Nishinomiya, Hyogo 663-8501, Japan
| | - Michio Kato
- National Hospital Organization Minami Wakayama Medical Center, Tanabe, Wakayama 646-8558, Japan
| | - Hitoshi Takagi
- Department of Gastroenterology, National Hospital Organization, Takasaki General Center Hospital, Takasaki, Gunma 370-0829, Japan
| | - Yasuhito Tanaka
- Liver Disease Center, Nagoya University Hospital, Nagoya, Aichi 467-8602, Japan
| | - Kazuo Notsumata
- Department of Internal Medicine, Fukuiken Saiseikai Hospital, Fukui 918-8503, Japan
| | - Eiji Mita
- Department of Gastroenterology, National Hospital Organization Osaka National Hospital, Osaka 540-0006, Japan
| | - Hideyuki Nomura
- Center for Liver Disease, Shin-kokura Hospital, Kitakyushu, Fukuoka 803-8505, Japan
| | - Mitsushige Shibatoge
- Department of Gastroenterology, Takamatsu Red-Cross Hospital, Takamatsu, Kagawa 760-0017, Japan
| | - Koichi Takaguchi
- Department of Hepatology, Kagawa Prefectural Central Hospital, Takamatsu, Kagawa 760-8557, Japan
| | | | - Michio Sata
- Research Center for Innovative Cancer Therapy, Kurume University School of Medicine, Fukuoka 830-0011, Japan
| | - Kazuhiko Koike
- Department of Gastroenterology, Graduate School of Medicine, The University of Tokyo, Tokyo 113-0033, Japan
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Strickland JC, Staton M, Leukefeld CG, Oser CB, Webster JM. Hepatitis C antibody reactivity among high-risk rural women: opportunities for services and treatment in the criminal justice system. Int J Prison Health 2019; 14:89-100. [PMID: 29869584 DOI: 10.1108/ijph-03-2017-0012] [Citation(s) in RCA: 7] [Impact Index Per Article: 1.2] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 02/06/2023]
Abstract
Purpose The purpose of this paper is to examine the drug use and criminal justice factors related to hepatitis C virus (HCV) antibody reactivity among rural women in the USA recruited from local jails. Design/methodology/approach Analyses included 277 women with a history of injection drug use from three rural jails in Kentucky. Participants completed health and drug use questionnaires and received antibody testing for HCV. Findings The majority of women tested reactive to the HCV antibody (69 percent). Reactivity was associated with risk factors, such as unsterile needle use. Criminal justice variables, including an increased likelihood of prison incarceration, an earlier age of first arrest, and a longer incarceration history, were associated with HCV reactive tests. Participants also endorsed several barriers to seeking healthcare before entering jail that were more prevalent in women testing HCV reactive regardless of HCV status awareness before entering jail. Originality/value Injection and high-risk sharing practices as well as criminal justice factors were significantly associated with HCV reactivity. Future research and practice could focus on opportunities for linkages to HCV treatment during incarceration as well as during community re-entry to help overcome real or perceived treatment barriers. The current study highlights the importance of the criminal justice system as a non-traditional, real-world setting to examine drug use and related health consequences such as HCV by describing the association of high-risk drug use and criminal justice consequences with HCV among rural women recruited from local jails.
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Affiliation(s)
- Justin C Strickland
- Department of Psychology, College of Arts and Sciences, University of Kentucky , Lexington, Kentucky, USA
| | - Michele Staton
- Department of Behavioral Science, College of Medicine, University of Kentucky , Lexington, Kentucky, USA.,Center on Drug and Alcohol Research, University of Kentucky , Lexington, Kentucky, USA
| | - Carl G Leukefeld
- Department of Behavioral Science, College of Medicine, University of Kentucky , Lexington, Kentucky, USA.,Center on Drug and Alcohol Research, University of Kentucky , Lexington, Kentucky, USA
| | - Carrie B Oser
- Center on Drug and Alcohol Research, University of Kentucky , Lexington, Kentucky, USA.,Department of Sociology, College of Arts and Sciences, University of Kentucky , Lexington, Kentucky, USA
| | - J Matthew Webster
- Department of Behavioral Science, College of Medicine, University of Kentucky , Lexington, Kentucky, USA.,Center on Drug and Alcohol Research, University of Kentucky , Lexington, Kentucky, USA
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Kuwashiro T, Iwane S, Jinghe X, Matsuhashi S, Eguchi Y, Anzai K, Fujimoto K, Mizuta T, Sakamoto N, Ikeda M, Kato N, Ozaki I. Regulation of interferon signaling and HCV‑RNA replication by extracellular matrix. Int J Mol Med 2018; 42:957-965. [PMID: 29786754 PMCID: PMC6034922 DOI: 10.3892/ijmm.2018.3693] [Citation(s) in RCA: 7] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/12/2017] [Accepted: 05/09/2018] [Indexed: 01/08/2023] Open
Abstract
Although interferon (IFN)‑based treatment of patients with chronic hepatitis C virus (HCV) infection is widely applied, treatment resistance is often observed in patients with advanced liver fibrosis. Given that the molecular mechanisms of IFN resistance in liver fibrosis remain elusive, the present study investigated the effects of extracellular matrix (ECM) on IFN signaling in hepatic cells. The native HuH‑7 human hepatoma cell line and HuH‑7 cells were stably transfected with full‑length HCV‑RNA fused with Renilla luciferase (OR6 cells) were cultured on ECM‑coated dishes or non‑coated plastic dishes (NDs), and treated with human IFN‑α. In Huh‑7 cells cultured on coated dishes, the IFN‑stimulated response element (ISRE) luciferase activity was measured following ISRE plasmid transfection and the expression of IFN‑stimulated genes (ISG) were significantly lower than those in cells cultured on NDs. In addition, after IFN‑α treatment, the amount of HCV‑RNA and viral protein produced by OR6 cells cultured on coated dishes was higher than that produced by cells cultured on NDs. When cells were treated with β1‑integrin‑blocking antibody to disrupt the cell‑matrix interaction, the ISRE luciferase activity was restored, and the protein expression of ISG was increased, while that of HCV proteins was suppressed. Treatment of cells with integrin‑linked kinase (ILK) inhibitor or focal adhesion kinase (FAK) inhibitor restored the ISRE luciferase activity and expression of ISG proteins. These results suggested that β1‑integrin‑mediated signals affected the IFN signaling and promoted HCV replication. Therefore, the accumulation of ECM in liver fibrosis may impair IFN signaling through β1‑integrin‑mediated signaling involving ILK and FAK.
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Affiliation(s)
- Takuya Kuwashiro
- Department of Internal Medicine, Saga Medical School, Saga University, Saga 849‑8501, Japan
| | - Shinji Iwane
- Department of Internal Medicine, Saga Medical School, Saga University, Saga 849‑8501, Japan
| | - Xia Jinghe
- Department of Internal Medicine, Saga Medical School, Saga University, Saga 849‑8501, Japan
| | - Sachiko Matsuhashi
- Department of Internal Medicine, Saga Medical School, Saga University, Saga 849‑8501, Japan
| | - Yuichiro Eguchi
- Department of Internal Medicine, Saga Medical School, Saga University, Saga 849‑8501, Japan
| | - Keizo Anzai
- Department of Internal Medicine, Saga Medical School, Saga University, Saga 849‑8501, Japan
| | - Kazuma Fujimoto
- Department of Internal Medicine, Saga Medical School, Saga University, Saga 849‑8501, Japan
| | - Toshihiko Mizuta
- Department of Internal Medicine, Saga Medical School, Saga University, Saga 849‑8501, Japan
| | - Naoya Sakamoto
- Department of Gastroenterology and Hepatology, Hokkaido University Graduate School of Medicine, Department of Gastroenterology and Hepatology, Graduate School of Medicine, Hokkaido University, Sapporo, Hokkaido 060‑8638, Japan
| | - Masanori Ikeda
- Department of Tumor Virology, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical Sciences, Okayama 700‑8558, Japan
| | - Nobuyuki Kato
- Department of Tumor Virology, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical Sciences, Okayama 700‑8558, Japan
| | - Iwata Ozaki
- Department of Internal Medicine, Saga Medical School, Saga University, Saga 849‑8501, Japan
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Favorable Outcomes of Chinese HCV-Related Cirrhotic Patients with Sustained Virological Response after Pegylated Interferon Plus Ribavirin Treatment. BIOMED RESEARCH INTERNATIONAL 2017; 2017:8061091. [PMID: 28232944 PMCID: PMC5292367 DOI: 10.1155/2017/8061091] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.1] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Subscribe] [Scholar Register] [Received: 10/20/2016] [Revised: 12/08/2016] [Accepted: 01/04/2017] [Indexed: 02/07/2023]
Abstract
Few studies have conducted follow-up investigations of the clinical course in HCV-related cirrhotic patients who achieved a sustained virological response (SVR) with pegylated interferon plus ribavirin treatment (PegIFN + RBV). We investigated the clinical course and laboratory data in a prospective cohort study enrolling HCV-related cirrhotic patients who received PegIFN + RBV between August 2008 and July 2013 in China. Complete blood counts, liver function tests, and HCV-RNA were serially examined. Liver-related complications were recorded. To detect hepatocellular carcinoma (HCC), alpha-fetoprotein assays, and ultrasound scans were repeated at 6-month intervals. Twenty-five patients were enrolled, including 8 patients with decompensation events before treatment. Eighteen patients achieved SVR with a mean follow-up period of 25.78 months. During the follow-up period, only one patient exhibited HCV-RNA positivity and no decompensation events were detected, but 4 patients developed HCC after SVR. APRI decreased more in patients with SVR than in patients with non-SVR (median, -1.33 versus 0.86, P < 0.001). The albumin levels and platelet counts significantly increased during the follow-up period after SVR (44.27 ± 4.09 versus 42.63 ± 4.37, P = 0.037 and 173.89 ± 87.36 versus 160.11 ± 77.97, P = 0.047). These data indicated that HCV-related cirrhotic patients with SVR after PegIFN + RBV may have a favorable clinical course and improvements in laboratory data. Moreover, HCC should be monitored.
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5
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Budzko L, Marcinkowska-Swojak M, Jackowiak P, Kozlowski P, Figlerowicz M. Copy number variation of genes involved in the hepatitis C virus-human interactome. Sci Rep 2016; 6:31340. [PMID: 27510840 PMCID: PMC4980658 DOI: 10.1038/srep31340] [Citation(s) in RCA: 5] [Impact Index Per Article: 0.6] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/07/2016] [Accepted: 07/18/2016] [Indexed: 02/06/2023] Open
Abstract
Copy number variation (CNV) is a newly discovered form of intra-species genetic polymorphism that is defined as deletions or duplications of genome segments ranging from 1 kbp to several Mbp. CNV accounts for the majority of the genetic variation observed in humans (CNV regions cover more than 10% of the human genome); therefore, it may significantly influence both the phenotype and susceptibility to various diseases. Unfortunately, the impact of CNV on a number of diseases, including hepatitis C virus (HCV) infection, remains largely unexplored. Here, we analyzed 421 human genes encoding proteins that have been shown to interact with HCV proteins or genomic RNA (proteins from the HCV-human interactome). We found that 19 of the 421 candidate genes are located in putative CNV regions. For all of these genes, copy numbers were determined for European, Asiatic and African populations using the multiplex ligation-dependent amplification (MLPA) method. As a result, we identified 4 genes, IGLL1, MLLT4, PDPK1, PPP1R13L, for which the CN-genotype ranged from 1 to 6. All of these genes are involved in host-virus interaction; thus, their polymorphism has a potential impact on the development of HCV infection and/or therapy outcome.
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Affiliation(s)
- Lucyna Budzko
- Institute of Bioorganic Chemistry, Polish Academy of Sciences, Poznan, Poland
| | | | - Paulina Jackowiak
- Institute of Bioorganic Chemistry, Polish Academy of Sciences, Poznan, Poland
- Institute of Chemical Technology and Engineering, Poznan University of Technology, Poznan, Poland
| | - Piotr Kozlowski
- Institute of Bioorganic Chemistry, Polish Academy of Sciences, Poznan, Poland
- Institute of Chemical Technology and Engineering, Poznan University of Technology, Poznan, Poland
| | - Marek Figlerowicz
- Institute of Bioorganic Chemistry, Polish Academy of Sciences, Poznan, Poland
- Institute of Computing Science, Poznan University of Technology, Poznan, Poland
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Frimpong JA, D'Aunno T, Perlman DC, Strauss SM, Mallow A, Hernandez D, Schackman BR, Feaster DJ, Metsch LR. On-site bundled rapid HIV/HCV testing in substance use disorder treatment programs: study protocol for a hybrid design randomized controlled trial. Trials 2016; 17:117. [PMID: 26936623 PMCID: PMC4776446 DOI: 10.1186/s13063-016-1225-4] [Citation(s) in RCA: 12] [Impact Index Per Article: 1.3] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/04/2015] [Accepted: 02/10/2016] [Indexed: 12/11/2022] Open
Abstract
BACKGROUND More than 1.2 million people in the United States are living with human immunodeficiency virus (HIV), and 3.2 million are living with hepatitis C virus (HCV). An estimated 25 % of persons living with HIV also have HCV. It is therefore of great public health importance to ensure the prompt diagnosis of both HIV and HCV in populations that have the highest prevalence of both infections, including individuals with substance use disorders (SUD). METHODS/DESIGN In this theory-driven, efficacy-effectiveness-implementation hybrid study, we will develop and test an on-site bundled rapid HIV/HCV testing intervention for SUD treatment programs. Its aim is to increase the receipt of HIV and HCV test results among SUD treatment patients. Using a rigorous process involving patients, providers, and program managers, we will incorporate rapid HCV testing into evidence-based HIV testing and linkage to care interventions. We will then test, in a randomized controlled trial, the extent to which this bundled rapid HIV/HCV testing approach increases receipt of HIV and HCV test results. Lastly, we will conduct formative research to understand the barriers to, and facilitators of, the adoption, implementation, and sustainability of the bundled rapid testing strategy in SUD treatment programs. DISCUSSION Novel approaches that effectively integrate on-site rapid HIV and rapid HCV testing are needed to address both the HIV and HCV epidemics. If feasible and efficacious, bundled rapid HIV/HCV testing may offer a scalable, potentially cost-effective approach to testing high-risk populations, such as patients of SUD treatment programs. It may ultimately lead to improved linkage to care and progress through the HIV and HCV care and treatment cascades. TRIAL REGISTRATION ClinicalTrials.gov: NCT02355080 . (30 January 2015).
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Affiliation(s)
- Jemima A Frimpong
- Department of Health Policy and Management, Mailman School of Public Health, Columbia University, New York, USA.
| | - Thomas D'Aunno
- Robert F. Wagner Graduate School of Public Service, New York University, New York, USA.
| | - David C Perlman
- Mount Sinai Beth Israel; Ichan School of Medicine at Mount Sinai, New York, USA.
| | | | - Alissa Mallow
- Montefiore Health System, New York, USA, New York, USA.
| | - Diana Hernandez
- Department of Sociomedical Sciences, Mailman School of Public Health, Columbia University, New York, USA.
| | - Bruce R Schackman
- Department of Healthcare Policy and Research, Weill Cornell Medical College, New York, USA.
| | - Daniel J Feaster
- Division of Biostatistics, Department of Public Health Sciences, Miller School of Medicine, University of Miami, Miami, USA.
| | - Lisa R Metsch
- Department of Sociomedical Sciences, Mailman School of Public Health, Columbia University, New York, USA.
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7
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Wang Y, Li J, Wang X, Peña JC, Li K, Zhang T, Ho W. (-)-Epigallocatechin-3-Gallate Enhances Hepatitis C Virus Double-Stranded RNA Intermediates-Triggered Innate Immune Responses in Hepatocytes. Sci Rep 2016; 6:21595. [PMID: 26879672 PMCID: PMC4754899 DOI: 10.1038/srep21595] [Citation(s) in RCA: 20] [Impact Index Per Article: 2.2] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/13/2015] [Accepted: 01/27/2016] [Indexed: 12/14/2022] Open
Abstract
(-)-Epigallocatechin-3-gallate (EGCG), a major polyphenol component of green tea, has recently been identified as an inhibitor of hepatitis C virus (HCV) entry. Here, we examined whether EGCG can enhance hepatocyte-mediated intracellular innate immunity against HCV. HCV dsRNAs (Core, E1-P7, NS-3'NTR and NS5A) induced interferon-λ1 (IFN-λ1) expression in human hepatocytes. These HCV dsRNAs also induced the expression of Toll-like receptor 3 (TLR3), retinoic acid-inducible gene I (RIG-I) and several antiviral IFN-stimulated genes (ISGs) expression. Although EGCG treatment of hepatocytes alone had little effect on TLR3 and RIG-I signaling pathways, EGCG significantly enhanced HCV dsRNAs-induced the expression of IFN-λ1, TLR3, RIG-I and antiviral ISGs in hepatocytes. Furthermore, treatment of HCV-infected hepatocytes with EGCG and HCV dsRNAs inhibited viral replication. Given that EGCG has the ability to enhance HCV dsRNAs-induced intracellular antiviral innate immunity against HCV, suggesting the potential application of EGCG as a new anti-HCV agent for HCV therapy.
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Affiliation(s)
- Yizhong Wang
- Department of Infectious Diseases, Shanghai Children’s Hospital, Shanghai Jiao Tong University, Shanghai 200040, P.R. China
| | - Jieliang Li
- Department of Pathology and Laboratory Medicine, Temple University School of Medicine, Philadelphia, PA 19140, USA
| | - Xu Wang
- Department of Pathology and Laboratory Medicine, Temple University School of Medicine, Philadelphia, PA 19140, USA
| | - Juliet C. Peña
- Department of Pathology and Laboratory Medicine, Temple University School of Medicine, Philadelphia, PA 19140, USA
| | - Kui Li
- Deparment of Microbiology, Immunology, and Biochemistry, University of Tennessee Health Science Center, Memphis, TN 38163, USA
| | - Ting Zhang
- Department of Infectious Diseases, Shanghai Children’s Hospital, Shanghai Jiao Tong University, Shanghai 200040, P.R. China
| | - Wenzhe Ho
- Department of Pathology and Laboratory Medicine, Temple University School of Medicine, Philadelphia, PA 19140, USA
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Perlman DC, Jordan AE, McKnight C, Young C, Delucchi KL, Sorensen JL, Des Jarlais DC, Masson CL. Viral hepatitis among drug users in methadone maintenance: associated factors, vaccination outcomes, and interventions. J Addict Dis 2015; 33:322-31. [PMID: 25299236 DOI: 10.1080/10550887.2014.969623] [Citation(s) in RCA: 8] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/05/2023]
Abstract
Drug users are at high risk of viral Hepatitis A, B, and C. The prevalence of Hepatitis A, Hepatitis B, and Hepatitis C, associated factors, and vaccine seroconversion among drug treatment program participants in a randomized controlled trial of hepatitis care coordination were examined. Of 489 participants, 44 and 47% required Hepatitis A/Hepatitis B vaccinations, respectively; 59% were Hepatitis C positive requiring linkage to care. Factors associated with serologic statuses, and vaccine seroconversion are reported; implications for strategies in drug treatment settings are discussed. Results suggest generalizable strategies for drug treatment programs to expand viral hepatitis screening, prevention, vaccination, and linkage to care.
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Affiliation(s)
- David C Perlman
- a Chemical Dependency Institute, Mount Sinai Beth Israel , New York , New York , USA
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9
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Teimourpour R, Meshkat Z, Gholoubi A, Nomani H, Rostami S. Viral Load Analysis of Hepatitis C Virus in Huh7.5 Cell Culture System. Jundishapur J Microbiol 2015; 8:e19279. [PMID: 26290686 PMCID: PMC4537521 DOI: 10.5812/jjm.8(5)2015.19279] [Citation(s) in RCA: 8] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/05/2014] [Revised: 06/26/2014] [Accepted: 07/23/2014] [Indexed: 01/26/2023] Open
Abstract
BACKGROUND Previous studies using cell culture systems for the replication of hepatitis C virus have opened new research dimensions, and paved the ways for further and detailed studies of the virus in vitro. OBJECTIVES The purpose of the present study was to cultivate hepatitis C virus in a cell culture system and evaluate viral amplification. MATERIALS AND METHODS In order to propagate hepatitis C virus, cloned whole genome of virus, JFH-1, was used. JFH-1 cDNA was introduced into strain JM109 of Escherichia coli and plasmid, containing the viral genome was purified from transformed bacteria. After XbaI digestion, RNA synthesis was induced using T7 RNA polymerase enzyme. Next, eukaryotic cell line Huh 7.5 was transfected by the purified RNA. Finally, Huh-7.5 cell line was infected with replicated virus and viral load was determined using real-time PCR (Polymerase Chain Reaction). RESULTS The amount of viral load, which was measured using real-time PCR was 17592 IU/mL. CONCLUSIONS In the present study, using cell culture, a high titer (in acceptable range) of infectious hepatitis C virus was produced. This method could be used in future studies.
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Affiliation(s)
- Roghayeh Teimourpour
- Antimicrobial Resistance Research Center, Mashhad University of Medical Sciences, Mashhad, IR Iran
| | - Zahra Meshkat
- Antimicrobial Resistance Research Center, Mashhad University of Medical Sciences, Mashhad, IR Iran
- Corresponding author: Zahra Meshkat, Antimicrobial Resistance Research Center, Mashhad University of Medical Sciences, P. O. Box: 9196773117, Mashhad, IR Iran. Tel: +98-5138012453, Fax: +98-5138002287, E-mail:
| | - Aida Gholoubi
- Department of Modern Sciences and Technologies, School of Medicine, Mashhad University of Medical Sciences, Mashhad, IR Iran
| | - Hosein Nomani
- Antimicrobial Resistance Research Center, Mashhad University of Medical Sciences, Mashhad, IR Iran
| | - Sina Rostami
- Department of Biology, Faculty of Sciences, Ferdowsi University of Mashhad, Mashhad, IR Iran
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Pfeil AM, Reich O, Guerra IM, Cure S, Negro F, Müllhaupt B, Lavanchy D, Schwenkglenks M. Cost-effectiveness analysis of sofosbuvir compared to current standard treatment in Swiss patients with chronic hepatitis C. PLoS One 2015; 10:e0126984. [PMID: 25974722 PMCID: PMC4431849 DOI: 10.1371/journal.pone.0126984] [Citation(s) in RCA: 35] [Impact Index Per Article: 3.5] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/13/2015] [Accepted: 04/09/2015] [Indexed: 12/17/2022] Open
Abstract
In clinical trials, sofosbuvir showed high antiviral activity in patients infected with hepatitis C virus (HCV) across all genotypes. We aimed to determine the cost-effectiveness of sofosbuvir-based treatment compared to current standard treatment in mono-infected patients with chronic hepatitis C (CHC) genotypes 1-4 in Switzerland. Cost-effectiveness was modelled from the perspective of the Swiss health care system using a lifetime Markov model. Incremental cost-effectiveness ratios (ICERs) used an endpoint of cost per quality-adjusted life year (QALY) gained. Treatment characteristics, quality of life, and transition probabilities were obtained from published literature. Country-specific model inputs such as patient characteristics, mortality and costs were obtained from Swiss sources. We performed extensive sensitivity analyses. Costs and effects were discounted at 3% (range: 0-5%) per year. Sofosbuvir-containing treatment in mixed cohorts of cirrhotic and non-cirrhotic patients with CHC genotypes 1-4 showed ICERs between CHF 10,337 and CHF 91,570 per QALY gained. In subgroup analyses, sofosbuvir dominated telaprevir- and boceprevir-containing treatment in treatment-naïve genotype 1 cirrhotic patients. ICERs of sofosbuvir were above CHF 100,000 per QALY in treatment-naïve, interferon eligible, non-cirrhotic patients infected with genotypes 2 or 3. In deterministic and probabilistic sensitivity analyses, results were generally robust. From a Swiss health care system perspective, treatment of mixed cohorts of cirrhotic and non-cirrhotic patients with CHC genotypes 1-4 with sofosbuvir-containing treatment versus standard treatment would be cost-effective if a threshold of CHF 100,000 per QALY was assumed.
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Affiliation(s)
- Alena M. Pfeil
- Institute of Pharmaceutical Medicine (ECPM), University of Basel, Basel, Switzerland
| | - Oliver Reich
- Department of Health Sciences, Helsana Group, Zurich, Switzerland
| | | | | | - Francesco Negro
- Divisions of Gastroenterology and Hepatology and of Clinical Pathology, University Hospital Geneva, Geneva, Switzerland
| | - Beat Müllhaupt
- Swiss HPB (Hepato-Pancreato-Biliary) Center and Department of Gastroenterology and Hepatology, University Hospital Zürich, Zurich, Switzerland
| | - Daniel Lavanchy
- World Health Organization (WHO), African Union, Governments, Denges VD, Switzerland
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11
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Zhu YP, Peng ZG, Wu ZY, Li JR, Huang MH, Si SY, Jiang JD. Host APOBEC3G protein inhibits HCV replication through direct binding at NS3. PLoS One 2015; 10:e0121608. [PMID: 25811715 PMCID: PMC4374698 DOI: 10.1371/journal.pone.0121608] [Citation(s) in RCA: 26] [Impact Index Per Article: 2.6] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/04/2014] [Accepted: 02/02/2015] [Indexed: 02/07/2023] Open
Abstract
Human APOBEC3G (hA3G) is a cytidine deaminase that restricts replication of certain viruses. We have previously reported that hA3G was a host restriction factor against hepatitis C virus (HCV) replication, and hA3G stabilizers showed a significant inhibitory activity against HCV. However, the molecular mechanism of hA3G against HCV remains unknown. We show in this study that hA3G’s C-terminal directly binds HCV non-structural protein NS3 at its C-terminus, which is responsible for NS3’s helicase and NTPase activity. Binding of hA3G to the C-terminus of NS3 reduced helicase activity, and therefore inhibited HCV replication. The anti-HCV mechanism of hA3G appeared to be independent of its deamination activity. Although early stage HCV infection resulted in an increase in host hA3G as an intracellular response against HCV replication, hA3G was gradually diminished after a long-term incubation, suggesting an unknown mechanism(s) that protects HCV NS3 from inactivation by hA3G. The process represents, at least partially, a cellular defensive mechanism against HCV and the action is mediated through a direct interaction between host hA3G and HCV NS3. We believe that understanding of the antiviral mechanism of hA3G against HCV might open an interesting avenue to explore hA3G stabilizers as a new class of anti-HCV agents.
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Affiliation(s)
- Yan-Ping Zhu
- Laboratory of Antiviral Research, Institute of Medicinal Biotechnology, Chinese Academy of Medical Sciences/Peking Union Medical College, Beijing, 100050, China
| | - Zong-Gen Peng
- Laboratory of Antiviral Research, Institute of Medicinal Biotechnology, Chinese Academy of Medical Sciences/Peking Union Medical College, Beijing, 100050, China
- * E-mail: (ZGP); (JDJ)
| | - Zhou-Yi Wu
- Laboratory of Antiviral Research, Institute of Medicinal Biotechnology, Chinese Academy of Medical Sciences/Peking Union Medical College, Beijing, 100050, China
| | - Jian-Rui Li
- Laboratory of Antiviral Research, Institute of Medicinal Biotechnology, Chinese Academy of Medical Sciences/Peking Union Medical College, Beijing, 100050, China
| | - Meng-Hao Huang
- Laboratory of Antiviral Research, Institute of Medicinal Biotechnology, Chinese Academy of Medical Sciences/Peking Union Medical College, Beijing, 100050, China
| | - Shu-Yi Si
- Laboratory of Antiviral Research, Institute of Medicinal Biotechnology, Chinese Academy of Medical Sciences/Peking Union Medical College, Beijing, 100050, China
| | - Jian-Dong Jiang
- Laboratory of Antiviral Research, Institute of Medicinal Biotechnology, Chinese Academy of Medical Sciences/Peking Union Medical College, Beijing, 100050, China
- State Key Laboratory of Bioactive Substance and Function of Natural Medicines, Institute of Materia Medica, Chinese Academy of Medical Sciences/Peking Union Medical College, Beijing, 100050, China
- * E-mail: (ZGP); (JDJ)
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12
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Lim SG. Chronic hepatitis C genotype 1 treatment roadmap for resource constrained settings. World J Gastroenterol 2015; 21:1972-1981. [PMID: 25684966 PMCID: PMC4323477 DOI: 10.3748/wjg.v21.i6.1972] [Citation(s) in RCA: 9] [Impact Index Per Article: 0.9] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 06/18/2014] [Revised: 07/21/2014] [Accepted: 09/19/2014] [Indexed: 02/06/2023] Open
Abstract
AIM: To use existing hepatitis C virus (HCV) antiviral therapies as access to new treatments is limited.
METHODS: A PubMed search for randomised control trials or meta-analysis related to response-guided therapy of HCV genotype 1 patients was undertaken using pegylated interferon and ribavirin (PR), boceprevir (B) and telaprevir (T) and lead-in where response-guided therapy at TW4(TW4), 8(TW8), 10(TW10), or 12(TW12) based on HCVRNA(+) or HCVRNA(-). Studies presented at major conferences were also used. Where necessary, a post-hoc analysis was performed. A response-guided management roadmap was created based on sustained virological response (SVR).
RESULTS: Starting with PR, those with HCVRNA(-) at TW4 have > 86% SVR, while those are HCVRNA(+) have 34%-41.7% SVR. HCVRNA(-) TW4 patients can have 24 wk PR if HCVRNA < 400000 IU/mL. Alternatively, 28 wk BPR has similar SVR. If HCVRNA(+) at TW4, 72 wk PR leads to 53% SVR, hence BPR is a better option, and if HCVRNA(-) by TW8, 28 wk therapy is sufficient. If HCVRNA(+) at TW8, then HCVRNA should be checked at TW10 and TW12. By TW12, HCVRNA ≥ 100 IU/mL activates the stopping rule. This roadmap is applicable for treatment-naïve, treatment failures and cirrhotic patients. Validation from an Asia Pacific early access boceprevir program confirmed the findings that HCVRNA(-) at TW4, or TW8 conferred > 80% SVR, leading to the “80-80” rule.
CONCLUSION: Using a roadmap based on HCVRNA(-) at TW4 or TW8 (the “80-80” rule), high SVR can be achieved, and guide the best choices for treatment, and also reduces drug exposure in poor responders.
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13
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Han T, Wan Y, Wang J, Zhao P, Yuan Y, Wang L, She Y, Broering R, Lu M, Ye L, Zhu Y. Set7 facilitates hepatitis C virus replication via enzymatic activity-dependent attenuation of the IFN-related pathway. THE JOURNAL OF IMMUNOLOGY 2015; 194:2757-68. [PMID: 25681344 DOI: 10.4049/jimmunol.1400583] [Citation(s) in RCA: 14] [Impact Index Per Article: 1.4] [Reference Citation Analysis] [Abstract] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 12/25/2022]
Abstract
Hepatitis C virus (HCV) infection is a major cause of chronic liver disease, usually resulting in persistent infection involving hepatic steatosis, cirrhosis, and hepatocellular carcinoma via escape of the host's immune response. Set7 is a lysine-specific methyltransferase that is involved in gene regulation and virus replication. However, the mechanism underlying the immune evasion between HCV and Set7 is not well understood. In this study, we observed that the expression of Set7 in Huh7.5.1 cells was upregulated by HCV infection, and high levels of Set7 expression were also found in the sera, PBMCs, and liver tissue of HCV patients relative to healthy individuals. Further investigation showed that Set7 enhanced HCV replication in an enzymatic activity-dependent manner. Moreover, our data showed that Set7 decreased the expression of virus-induced IFN and IFN-related effectors, such as dsRNA-activated protein kinase and 2',5'-oligoadenylate synthetase. Further investigation suggested that Set7 suppressed the endogenous IFN expression by reducing the nuclear translocation of IFN regulatory factor 3/7 and the p65 subunit of NF-κB and reduced IFN-induced dsRNA-activated protein kinase and 2',5'-oligoadenylate synthetase via attenuation of the phosphorylation of STAT1 and STAT2. Additionally, IFN receptors, including IFNAR1 and IFNAR2, which are located upstream of the JAK/STAT pathway, were reduced by Set7. Taken together, our results reveal that Set7 facilitates HCV replication through the attenuation of IFN signaling pathways and IFN-related effectors.
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Affiliation(s)
- Tao Han
- State Key Laboratory of Virology, College of Life Sciences, Wuhan University, Wuhan, Hubei 430072, China
| | - Yushun Wan
- State Key Laboratory of Virology, College of Life Sciences, Wuhan University, Wuhan, Hubei 430072, China
| | - Jun Wang
- State Key Laboratory of Virology, College of Life Sciences, Wuhan University, Wuhan, Hubei 430072, China
| | - Peng Zhao
- State Key Laboratory of Virology, College of Life Sciences, Wuhan University, Wuhan, Hubei 430072, China
| | - Yue Yuan
- State Key Laboratory of Virology, College of Life Sciences, Wuhan University, Wuhan, Hubei 430072, China
| | - Li Wang
- State Key Laboratory of Virology, College of Life Sciences, Wuhan University, Wuhan, Hubei 430072, China
| | - Yinglong She
- State Key Laboratory of Virology, College of Life Sciences, Wuhan University, Wuhan, Hubei 430072, China
| | - Ruth Broering
- Medical Faculty, Department of Gastroenterology and Hepatology, University of Duisburg-Essen, 45127 Essen, Germany; and
| | - Mengji Lu
- Institute of Virology, University of Duisburg-Essen, 45127 Essen, Germany
| | - Linbai Ye
- State Key Laboratory of Virology, College of Life Sciences, Wuhan University, Wuhan, Hubei 430072, China
| | - Ying Zhu
- State Key Laboratory of Virology, College of Life Sciences, Wuhan University, Wuhan, Hubei 430072, China;
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14
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Sedano CD, Sarnow P. Interaction of host cell microRNAs with the HCV RNA genome during infection of liver cells. Semin Liver Dis 2015; 35:75-80. [PMID: 25632937 PMCID: PMC4832929 DOI: 10.1055/s-0034-1397351] [Citation(s) in RCA: 14] [Impact Index Per Article: 1.4] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 02/01/2023]
Abstract
It has remained an enigma how hepatitis C viral (HCV) RNA can persist in the liver of infected patients for many decades. With the recent discovery of roles for microRNAs in gene expression, it was reported that the HCV RNA genome subverts liver-specific microRNA miR-122 to protect its 5' end from degradation by host cell exoribonucleases. Sequestration of miR-122 in cultured liver cells and in the liver of chimpanzees by small, modified antisense RNAs resulted in dramatic loss of HCV RNA and viral yield. This finding led to the first successful human trial in which subcutaneous administration of antisense molecules against miR-122 lowered viral yield in HCV patients, without the emergence of resistant virus. In this review, the authors summarize the molecular mechanism by which miR-122 protects the HCV RNA genome from degradation by exoribonucleases Xrn1 and Xrn2 and discuss the application of miR-122 antisense molecules in the clinic.
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Affiliation(s)
- Cecilia D. Sedano
- Department of Microbiology and Immunology, Stanford University, School of Medicine, Stanford, California
| | - Peter Sarnow
- Department of Microbiology and Immunology, Stanford University, School of Medicine, Stanford, California
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15
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Sagan SM, Chahal J, Sarnow P. cis-Acting RNA elements in the hepatitis C virus RNA genome. Virus Res 2015; 206:90-8. [PMID: 25576644 DOI: 10.1016/j.virusres.2014.12.029] [Citation(s) in RCA: 30] [Impact Index Per Article: 3.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/01/2014] [Revised: 12/12/2014] [Accepted: 12/24/2014] [Indexed: 12/22/2022]
Abstract
Hepatitis C virus (HCV) infection is a rapidly increasing global health problem with an estimated 170 million people infected worldwide. HCV is a hepatotropic, positive-sense RNA virus of the family Flaviviridae. As a positive-sense RNA virus, the HCV genome itself must serve as a template for translation, replication and packaging. The viral RNA must therefore be a dynamic structure that is able to readily accommodate structural changes to expose different regions of the genome to viral and cellular proteins to carry out the HCV life cycle. The ∼ 9600 nucleotide viral genome contains a single long open reading frame flanked by 5' and 3' non-coding regions that contain cis-acting RNA elements important for viral translation, replication and stability. Additional cis-acting RNA elements have also been identified in the coding sequences as well as in the 3' end of the negative-strand replicative intermediate. Herein, we provide an overview of the importance of these cis-acting RNA elements in the HCV life cycle.
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Affiliation(s)
- Selena M Sagan
- Department of Microbiology & Immunology, McGill University, Montreal, QC, Canada
| | - Jasmin Chahal
- Department of Microbiology & Immunology, McGill University, Montreal, QC, Canada
| | - Peter Sarnow
- Department of Microbiology and Immunology, Stanford University, Stanford, CA, United States.
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16
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Zou X, Chi X, Pan Y, Du D, Sun H, Matsuda A, Li W, Kuno A, Zhang X, Narimatsu H, Niu J, Zhang Y. LecT-Hepa facilitates estimating treatment outcome during interferon therapy in chronic hepatitis C patients. Clin Proteomics 2014; 11:44. [PMID: 25593566 PMCID: PMC4276098 DOI: 10.1186/1559-0275-11-44] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.1] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/25/2014] [Accepted: 11/25/2014] [Indexed: 02/07/2023] Open
Abstract
Background A combination treatment of interferon and ribavirin is the standard and the commonly used treatment for chronic hepatitis C (CHC). Developing noninvasive tests like serum indicators that can predict treatment outcome at an early stage of therapy is beneficial for individualized treatment and management of CHC. A glyco-indicator based on the glyco-alteration of serum α1-acid glycoprotein, LecT-Hepa, was discovered by glycomics technologies as a robust indicator of liver fibrosis. Here, we investigated the clinical utility of LecT-Hepa for evaluation of treatment outcome. Results Firstly, ninety-seven patients with CHC were used for comparison of LecT-Hepa in serum and plasma. We found no significant difference in the concentrations of LecT-Hepa in serum and plasma. And then, 213 serum specimens from 45 patients who received 48 weeks of treatment with interferon and ribavirin were followed up for 96 weeks, and were used for evaluation of the role of LecT-Hepa. We found that LecT-Hepa might reflect the change in fibrosis regression during the treatment process. Moreover, the change of LecT-Hepa at the first 12 weeks of treatment could already predict the antiviral treatment response, which was more superior to FIB-4 index and aspartate aminotransferase-to-platelet ratio index (APRI) in this study. Conclusions These results provide a new perspective that serum glycoprotein could be used as a joint diagnosis indicator for estimation treatment outcome of viral hepatitis at earlier stage of therapy. Electronic supplementary material The online version of this article (doi:10.1186/1559-0275-11-44) contains supplementary material, which is available to authorized users.
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Affiliation(s)
- Xia Zou
- Ministry of Education Key Laboratory of Systems Biomedicine, Shanghai Center for Systems Biomedicine, Shanghai Jiao Tong University, 800 Dong Chuan Road, Minhang Shanghai, 200240 China.,SCSB (China) - AIST (Japan) Joint Medical Glycomics Laboratory, 800 Dong Chuan Road, Minhang Shanghai, 200240 China.,Department of Surgery, Ruijin Hospital, Shanghai Jiao Tong University School of Medicine, 197, Ruijin Er Road, Shanghai, 200025 China
| | - Xiumei Chi
- Department of Hepatology, First Hospital, Jilin University, Changchun, 130021 China
| | - Yu Pan
- Department of Hepatology, First Hospital, Jilin University, Changchun, 130021 China
| | - Dongning Du
- Research Center for Medical Glycoscience (RCMG), National Institute of Advanced Industrial Science and Technology (AIST), 1-1-1 Umezono, Tsukuba, Ibaraki, 305-8568 Japan.,SCSB (China) - AIST (Japan) Joint Medical Glycomics Laboratory, 800 Dong Chuan Road, Minhang Shanghai, 200240 China
| | - Haibo Sun
- Department of Hepatology, First Hospital, Jilin University, Changchun, 130021 China
| | - Atsushi Matsuda
- Research Center for Medical Glycoscience (RCMG), National Institute of Advanced Industrial Science and Technology (AIST), 1-1-1 Umezono, Tsukuba, Ibaraki, 305-8568 Japan.,SCSB (China) - AIST (Japan) Joint Medical Glycomics Laboratory, 800 Dong Chuan Road, Minhang Shanghai, 200240 China
| | - Wei Li
- Ministry of Education Key Laboratory of Systems Biomedicine, Shanghai Center for Systems Biomedicine, Shanghai Jiao Tong University, 800 Dong Chuan Road, Minhang Shanghai, 200240 China.,SCSB (China) - AIST (Japan) Joint Medical Glycomics Laboratory, 800 Dong Chuan Road, Minhang Shanghai, 200240 China
| | - Atsushi Kuno
- Research Center for Medical Glycoscience (RCMG), National Institute of Advanced Industrial Science and Technology (AIST), 1-1-1 Umezono, Tsukuba, Ibaraki, 305-8568 Japan.,SCSB (China) - AIST (Japan) Joint Medical Glycomics Laboratory, 800 Dong Chuan Road, Minhang Shanghai, 200240 China
| | - Xinxin Zhang
- Department of Infectious Diseases, Ruijin Hospital, Shanghai Jiao Tong University School of Medicine, 197, Ruijin Er Road, Shanghai, 200025 China
| | - Hisashi Narimatsu
- Research Center for Medical Glycoscience (RCMG), National Institute of Advanced Industrial Science and Technology (AIST), 1-1-1 Umezono, Tsukuba, Ibaraki, 305-8568 Japan.,SCSB (China) - AIST (Japan) Joint Medical Glycomics Laboratory, 800 Dong Chuan Road, Minhang Shanghai, 200240 China
| | - Junqi Niu
- Department of Hepatology, First Hospital, Jilin University, Changchun, 130021 China
| | - Yan Zhang
- Ministry of Education Key Laboratory of Systems Biomedicine, Shanghai Center for Systems Biomedicine, Shanghai Jiao Tong University, 800 Dong Chuan Road, Minhang Shanghai, 200240 China.,SCSB (China) - AIST (Japan) Joint Medical Glycomics Laboratory, 800 Dong Chuan Road, Minhang Shanghai, 200240 China
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17
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Lendvai G, Jármay K, Karácsony G, Halász T, Kovalszky I, Baghy K, Wittmann T, Schaff Z, Kiss A. Elevated miR-33a and miR-224 in steatotic chronic hepatitis C liver biopsies. World J Gastroenterol 2014; 20:15343-15350. [PMID: 25386083 PMCID: PMC4223268 DOI: 10.3748/wjg.v20.i41.15343] [Citation(s) in RCA: 26] [Impact Index Per Article: 2.4] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 02/11/2014] [Revised: 05/07/2014] [Accepted: 06/13/2014] [Indexed: 02/07/2023] Open
Abstract
AIM To assess the expression of selected microRNAs (miRNA) in hepatitis C, steatotic hepatitis C, noninfected steatotic and normal liver tissues. METHODS The relative expression levels of miR-21, miR-33a, miR-96, miR-122, miR-125b, miR-221 and miR-224 were determined in 76 RNA samples isolated from 18 non-steatotic and 28 steatotic chronic hepatitis C (CHC and CHC-Steatosis, respectively) cases, 18 non-infected, steatotic liver biopsies of metabolic origin (Steatosis) and 12 normal formalin-fixed paraffin-embedded liver tissues using TaqMan MicroRNA Assays. All CHC biopsy samples were obtained prior to initiating therapy. Patients' serum biochemical values, which included glucose, triglyceride, cholesterol, alanine aminotransferase (ALT), aspartate aminotransferase (AST), gamma-glutamyl-transferase (GGT), alkaline phosphatase (AP), were obtained and correlated with relative miRNA expression. RESULTS When compared with control non-infected liver samples, miR-122 and miR-221 levels were reduced in CHC-Steatosis (P < 0.03) and in CHC, CHC-Steatosis and Steatosis (P < 0.01). Alternatively, the expression of miR-33a and miR-224 were elevated in CHC-Steatosis and Steatosis in comparison to control tissue (P < 0.01). The levels of miR-33a and miR-224 in CHC-Steatosis (P < 0.02) and miR-224 in Steatosis (P < 0.001) were increased in comparison to CHC samples. By contrast, the expression of miR-21 did not differ statistically between diseased and normal liver samples. Levels of miR-33a correlated negatively with serum AST and AP levels in Steatosis as well as with necroinflammatory grade in CHC, whereas miR-21 correlated positively with AST in Steatosis and displayed negative correlation with triglyceride level in CHC-Steatosis. In contrast, miRNA levels were not correlated with ALT, GGT, cholesterol levels or fibrosis stage. CONCLUSION Differences in miRNA expression were observed between CHC and steatotic CHC, CHC and steatotic liver, but not between steatotic CHC and steatotic liver of metabolic origin.
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18
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Persistent hepatitis C virus infection impairs ribavirin antiviral activity through clathrin-mediated trafficking of equilibrative nucleoside transporter 1. J Virol 2014; 89:626-42. [PMID: 25339775 DOI: 10.1128/jvi.02492-14] [Citation(s) in RCA: 20] [Impact Index Per Article: 1.8] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/14/2022] Open
Abstract
UNLABELLED Ribavirin (RBV) continues to be an important component of interferon-free hepatitis C treatment regimens, as RBV alone does not inhibit hepatitis C virus (HCV) replication effectively; the reason for this ineffectiveness has not been established. In this study, we investigated the RBV resistance mechanism using a persistently HCV-infected cell culture system. The antiviral activity of RBV against HCV was progressively impaired in the persistently infected culture, whereas interferon lambda 1 (IFN-λ1), a type III IFN, showed a strong antiviral response and induced viral clearance. We found that HCV replication in persistently infected cultures induces an autophagy response that impairs RBV uptake by preventing the expression of equilibrative nucleoside transporter 1 (ENT1). The Huh-7.5 cell line treated with an autophagy inducer, Torin 1, downregulated membrane expression of ENT1 and terminated RBV uptake. In contrast, the autophagy inhibitors hydroxychloroquine (HCQ), 3-methyladenine (3-MA), and bafilomycin A1 (BafA1) prevented ENT1 degradation and enhanced RBV antiviral activity. The HCV-induced autophagy response, as well as treatment with Torin 1, degrades clathrin heavy chain expression in a hepatoma cell line. Reduced expression of the clathrin heavy chain by HCV prevents ENT1 recycling to the plasma membrane and forces ENT1 to the lysosome for degradation. This study provides a potential mechanism for the impairment of RBV antiviral activity in persistently HCV-infected cell cultures and suggests that inhibition of the HCV-induced autophagy response could be used as a strategy for improving RBV antiviral activity against HCV infection. IMPORTANCE The results from this work will allow a review of the competing theories of antiviral therapy development in the field of HCV virology. Ribavirin (RBV) remains an important component of interferon-free hepatitis C treatment regimens. The reason why RBV alone does not inhibit HCV replication effectively has not been established. This study provides a potential mechanism for why RBV antiviral activity is impaired in persistently HCV-infected cell cultures and suggests that inhibition of the HCV-induced autophagy response could be used as a strategy to increase RBV antiviral activity against HCV infection. Therefore, it is anticipated that this work would generate a great deal of interest, not only among virologists but also among the general public.
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19
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Increased prevalence of hepatitis C virus subtype 6a in China: a comparison between 2004-2007 and 2008-2011. Arch Virol 2014; 159:3231-7. [PMID: 25085624 PMCID: PMC4221604 DOI: 10.1007/s00705-014-2185-1] [Citation(s) in RCA: 27] [Impact Index Per Article: 2.5] [Reference Citation Analysis] [Abstract] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/11/2014] [Accepted: 07/16/2014] [Indexed: 02/07/2023]
Abstract
Different hepatitis C virus (HCV) genotypes exhibit differences in disease pathogenesis and progression, as well as disease outcomes and response to therapy. Tracking the change of HCV genotypes in various epidemiological settings is critical for both disease surveillance and the development of improved antiviral treatment. Here, we tracked the changes in the prevalence of the HCV genotypes in China between 2004-2007 and 2008-2011. HCV-RNA-positive sera were collected from volunteer blood donors during the period 2008-2011. The genotypes were determined by phylogenic analysis using the NS5B and E1 sequences. Geographical and demographic distribution patterns related to the HCV genotypes obtained in 2008-2011 were compared with our previous study, which recorded data in the period 2004-2007. Pearson chi-square test and t-test were used to statistically analyze the results. In 2008-2011, HCV subtypes 1b and 6a were detected in 43.8 % (184/420) and 34.3 % (144/420), respectively. The male/female ratio was found to be higher for HCV genotype 6 than for genotypes 1 and 2. When compared with the period of 2004-2007, although no significant difference was found in gender or age for genotypes 1, 2, 3 and 6, the subtype 6a frequency was significantly increased from 11 % to 26.5 % in the blood donors from outside of Guangdong Province in 2008-2011. A pattern of increase in HCV subtype 6a was found in blood donors outside of Guangdong Province, indicating that HCV subtype 6a has rapidly spread from Guangdong to other regions of China over the past 10 years.
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20
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Meissner EG, Wu D, Osinusi A, Bon D, Virtaneva K, Sturdevant D, Porcella S, Wang H, Herrmann E, McHutchison J, Suffredini AF, Polis M, Hewitt S, Prokunina-Olsson L, Masur H, Fauci AS, Kottilil S. Endogenous intrahepatic IFNs and association with IFN-free HCV treatment outcome. J Clin Invest 2014; 124:3352-63. [PMID: 24983321 PMCID: PMC4109554 DOI: 10.1172/jci75938] [Citation(s) in RCA: 175] [Impact Index Per Article: 15.9] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/04/2014] [Accepted: 05/19/2014] [Indexed: 12/14/2022] Open
Abstract
BACKGROUND. Hepatitis C virus (HCV) infects approximately 170 million people worldwide and may lead to cirrhosis and hepatocellular carcinoma in chronically infected individuals. Treatment is rapidly evolving from IFN-α-based therapies to IFN-α-free regimens that consist of directly acting antiviral agents (DAAs), which demonstrate improved efficacy and tolerability in clinical trials. Virologic relapse after DAA therapy is a common cause of treatment failure; however, it is not clear why relapse occurs or whether certain individuals are more prone to recurrent viremia. METHODS. We conducted a clinical trial using the DAA sofosbuvir plus ribavirin (SOF/RBV) and performed detailed mRNA expression analysis in liver and peripheral blood from patients who achieved either a sustained virologic response (SVR) or relapsed. RESULTS. On-treatment viral clearance was accompanied by rapid downregulation of IFN-stimulated genes (ISGs) in liver and blood, regardless of treatment outcome. Analysis of paired pretreatment and end of treatment (EOT) liver biopsies from SVR patients showed that viral clearance was accompanied by decreased expression of type II and III IFNs, but unexpectedly increased expression of the type I IFN IFNA2. mRNA expression of ISGs was higher in EOT liver biopsies of patients who achieved SVR than in patients who later relapsed. CONCLUSION. These results suggest that restoration of type I intrahepatic IFN signaling by EOT may facilitate HCV eradication and prevention of relapse upon withdrawal of SOF/RBV. TRIAL REGISTRATION. ClinicalTrials.gov NCT01441180.
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Affiliation(s)
- Eric G. Meissner
- Laboratory of Immunoregulation, NIAID, NIH, Bethesda, Maryland, USA. Division of Infectious Diseases, Institute of Human Virology, University of Maryland Medical School, Baltimore, Maryland, USA. Institute of Biostatistics and Mathematical Modeling, Johann Wolfgang Goethe University, Frankfurt, Germany. Genomics Unit, Research Technologies Section, Rocky Mountain Laboratories, NIAID, NIH, Hamilton, Montana, USA. Critical Care Medicine Department, Clinical Center, NIH, Bethesda, Maryland, USA. Gilead Sciences, Foster City, California, USA. Department of Pathology, NCI, NIH, Bethesda, Maryland, USA. Laboratory of Translational Genomics, Division of Cancer Epidemiology and Genetics, NCI, NIH, Bethesda, Maryland, USA
| | - David Wu
- Laboratory of Immunoregulation, NIAID, NIH, Bethesda, Maryland, USA. Division of Infectious Diseases, Institute of Human Virology, University of Maryland Medical School, Baltimore, Maryland, USA. Institute of Biostatistics and Mathematical Modeling, Johann Wolfgang Goethe University, Frankfurt, Germany. Genomics Unit, Research Technologies Section, Rocky Mountain Laboratories, NIAID, NIH, Hamilton, Montana, USA. Critical Care Medicine Department, Clinical Center, NIH, Bethesda, Maryland, USA. Gilead Sciences, Foster City, California, USA. Department of Pathology, NCI, NIH, Bethesda, Maryland, USA. Laboratory of Translational Genomics, Division of Cancer Epidemiology and Genetics, NCI, NIH, Bethesda, Maryland, USA
| | - Anu Osinusi
- Laboratory of Immunoregulation, NIAID, NIH, Bethesda, Maryland, USA. Division of Infectious Diseases, Institute of Human Virology, University of Maryland Medical School, Baltimore, Maryland, USA. Institute of Biostatistics and Mathematical Modeling, Johann Wolfgang Goethe University, Frankfurt, Germany. Genomics Unit, Research Technologies Section, Rocky Mountain Laboratories, NIAID, NIH, Hamilton, Montana, USA. Critical Care Medicine Department, Clinical Center, NIH, Bethesda, Maryland, USA. Gilead Sciences, Foster City, California, USA. Department of Pathology, NCI, NIH, Bethesda, Maryland, USA. Laboratory of Translational Genomics, Division of Cancer Epidemiology and Genetics, NCI, NIH, Bethesda, Maryland, USA
| | - Dimitra Bon
- Laboratory of Immunoregulation, NIAID, NIH, Bethesda, Maryland, USA. Division of Infectious Diseases, Institute of Human Virology, University of Maryland Medical School, Baltimore, Maryland, USA. Institute of Biostatistics and Mathematical Modeling, Johann Wolfgang Goethe University, Frankfurt, Germany. Genomics Unit, Research Technologies Section, Rocky Mountain Laboratories, NIAID, NIH, Hamilton, Montana, USA. Critical Care Medicine Department, Clinical Center, NIH, Bethesda, Maryland, USA. Gilead Sciences, Foster City, California, USA. Department of Pathology, NCI, NIH, Bethesda, Maryland, USA. Laboratory of Translational Genomics, Division of Cancer Epidemiology and Genetics, NCI, NIH, Bethesda, Maryland, USA
| | - Kimmo Virtaneva
- Laboratory of Immunoregulation, NIAID, NIH, Bethesda, Maryland, USA. Division of Infectious Diseases, Institute of Human Virology, University of Maryland Medical School, Baltimore, Maryland, USA. Institute of Biostatistics and Mathematical Modeling, Johann Wolfgang Goethe University, Frankfurt, Germany. Genomics Unit, Research Technologies Section, Rocky Mountain Laboratories, NIAID, NIH, Hamilton, Montana, USA. Critical Care Medicine Department, Clinical Center, NIH, Bethesda, Maryland, USA. Gilead Sciences, Foster City, California, USA. Department of Pathology, NCI, NIH, Bethesda, Maryland, USA. Laboratory of Translational Genomics, Division of Cancer Epidemiology and Genetics, NCI, NIH, Bethesda, Maryland, USA
| | - Dan Sturdevant
- Laboratory of Immunoregulation, NIAID, NIH, Bethesda, Maryland, USA. Division of Infectious Diseases, Institute of Human Virology, University of Maryland Medical School, Baltimore, Maryland, USA. Institute of Biostatistics and Mathematical Modeling, Johann Wolfgang Goethe University, Frankfurt, Germany. Genomics Unit, Research Technologies Section, Rocky Mountain Laboratories, NIAID, NIH, Hamilton, Montana, USA. Critical Care Medicine Department, Clinical Center, NIH, Bethesda, Maryland, USA. Gilead Sciences, Foster City, California, USA. Department of Pathology, NCI, NIH, Bethesda, Maryland, USA. Laboratory of Translational Genomics, Division of Cancer Epidemiology and Genetics, NCI, NIH, Bethesda, Maryland, USA
| | - Steve Porcella
- Laboratory of Immunoregulation, NIAID, NIH, Bethesda, Maryland, USA. Division of Infectious Diseases, Institute of Human Virology, University of Maryland Medical School, Baltimore, Maryland, USA. Institute of Biostatistics and Mathematical Modeling, Johann Wolfgang Goethe University, Frankfurt, Germany. Genomics Unit, Research Technologies Section, Rocky Mountain Laboratories, NIAID, NIH, Hamilton, Montana, USA. Critical Care Medicine Department, Clinical Center, NIH, Bethesda, Maryland, USA. Gilead Sciences, Foster City, California, USA. Department of Pathology, NCI, NIH, Bethesda, Maryland, USA. Laboratory of Translational Genomics, Division of Cancer Epidemiology and Genetics, NCI, NIH, Bethesda, Maryland, USA
| | - Honghui Wang
- Laboratory of Immunoregulation, NIAID, NIH, Bethesda, Maryland, USA. Division of Infectious Diseases, Institute of Human Virology, University of Maryland Medical School, Baltimore, Maryland, USA. Institute of Biostatistics and Mathematical Modeling, Johann Wolfgang Goethe University, Frankfurt, Germany. Genomics Unit, Research Technologies Section, Rocky Mountain Laboratories, NIAID, NIH, Hamilton, Montana, USA. Critical Care Medicine Department, Clinical Center, NIH, Bethesda, Maryland, USA. Gilead Sciences, Foster City, California, USA. Department of Pathology, NCI, NIH, Bethesda, Maryland, USA. Laboratory of Translational Genomics, Division of Cancer Epidemiology and Genetics, NCI, NIH, Bethesda, Maryland, USA
| | - Eva Herrmann
- Laboratory of Immunoregulation, NIAID, NIH, Bethesda, Maryland, USA. Division of Infectious Diseases, Institute of Human Virology, University of Maryland Medical School, Baltimore, Maryland, USA. Institute of Biostatistics and Mathematical Modeling, Johann Wolfgang Goethe University, Frankfurt, Germany. Genomics Unit, Research Technologies Section, Rocky Mountain Laboratories, NIAID, NIH, Hamilton, Montana, USA. Critical Care Medicine Department, Clinical Center, NIH, Bethesda, Maryland, USA. Gilead Sciences, Foster City, California, USA. Department of Pathology, NCI, NIH, Bethesda, Maryland, USA. Laboratory of Translational Genomics, Division of Cancer Epidemiology and Genetics, NCI, NIH, Bethesda, Maryland, USA
| | - John McHutchison
- Laboratory of Immunoregulation, NIAID, NIH, Bethesda, Maryland, USA. Division of Infectious Diseases, Institute of Human Virology, University of Maryland Medical School, Baltimore, Maryland, USA. Institute of Biostatistics and Mathematical Modeling, Johann Wolfgang Goethe University, Frankfurt, Germany. Genomics Unit, Research Technologies Section, Rocky Mountain Laboratories, NIAID, NIH, Hamilton, Montana, USA. Critical Care Medicine Department, Clinical Center, NIH, Bethesda, Maryland, USA. Gilead Sciences, Foster City, California, USA. Department of Pathology, NCI, NIH, Bethesda, Maryland, USA. Laboratory of Translational Genomics, Division of Cancer Epidemiology and Genetics, NCI, NIH, Bethesda, Maryland, USA
| | - Anthony F. Suffredini
- Laboratory of Immunoregulation, NIAID, NIH, Bethesda, Maryland, USA. Division of Infectious Diseases, Institute of Human Virology, University of Maryland Medical School, Baltimore, Maryland, USA. Institute of Biostatistics and Mathematical Modeling, Johann Wolfgang Goethe University, Frankfurt, Germany. Genomics Unit, Research Technologies Section, Rocky Mountain Laboratories, NIAID, NIH, Hamilton, Montana, USA. Critical Care Medicine Department, Clinical Center, NIH, Bethesda, Maryland, USA. Gilead Sciences, Foster City, California, USA. Department of Pathology, NCI, NIH, Bethesda, Maryland, USA. Laboratory of Translational Genomics, Division of Cancer Epidemiology and Genetics, NCI, NIH, Bethesda, Maryland, USA
| | - Michael Polis
- Laboratory of Immunoregulation, NIAID, NIH, Bethesda, Maryland, USA. Division of Infectious Diseases, Institute of Human Virology, University of Maryland Medical School, Baltimore, Maryland, USA. Institute of Biostatistics and Mathematical Modeling, Johann Wolfgang Goethe University, Frankfurt, Germany. Genomics Unit, Research Technologies Section, Rocky Mountain Laboratories, NIAID, NIH, Hamilton, Montana, USA. Critical Care Medicine Department, Clinical Center, NIH, Bethesda, Maryland, USA. Gilead Sciences, Foster City, California, USA. Department of Pathology, NCI, NIH, Bethesda, Maryland, USA. Laboratory of Translational Genomics, Division of Cancer Epidemiology and Genetics, NCI, NIH, Bethesda, Maryland, USA
| | - Stephen Hewitt
- Laboratory of Immunoregulation, NIAID, NIH, Bethesda, Maryland, USA. Division of Infectious Diseases, Institute of Human Virology, University of Maryland Medical School, Baltimore, Maryland, USA. Institute of Biostatistics and Mathematical Modeling, Johann Wolfgang Goethe University, Frankfurt, Germany. Genomics Unit, Research Technologies Section, Rocky Mountain Laboratories, NIAID, NIH, Hamilton, Montana, USA. Critical Care Medicine Department, Clinical Center, NIH, Bethesda, Maryland, USA. Gilead Sciences, Foster City, California, USA. Department of Pathology, NCI, NIH, Bethesda, Maryland, USA. Laboratory of Translational Genomics, Division of Cancer Epidemiology and Genetics, NCI, NIH, Bethesda, Maryland, USA
| | - Ludmila Prokunina-Olsson
- Laboratory of Immunoregulation, NIAID, NIH, Bethesda, Maryland, USA. Division of Infectious Diseases, Institute of Human Virology, University of Maryland Medical School, Baltimore, Maryland, USA. Institute of Biostatistics and Mathematical Modeling, Johann Wolfgang Goethe University, Frankfurt, Germany. Genomics Unit, Research Technologies Section, Rocky Mountain Laboratories, NIAID, NIH, Hamilton, Montana, USA. Critical Care Medicine Department, Clinical Center, NIH, Bethesda, Maryland, USA. Gilead Sciences, Foster City, California, USA. Department of Pathology, NCI, NIH, Bethesda, Maryland, USA. Laboratory of Translational Genomics, Division of Cancer Epidemiology and Genetics, NCI, NIH, Bethesda, Maryland, USA
| | - Henry Masur
- Laboratory of Immunoregulation, NIAID, NIH, Bethesda, Maryland, USA. Division of Infectious Diseases, Institute of Human Virology, University of Maryland Medical School, Baltimore, Maryland, USA. Institute of Biostatistics and Mathematical Modeling, Johann Wolfgang Goethe University, Frankfurt, Germany. Genomics Unit, Research Technologies Section, Rocky Mountain Laboratories, NIAID, NIH, Hamilton, Montana, USA. Critical Care Medicine Department, Clinical Center, NIH, Bethesda, Maryland, USA. Gilead Sciences, Foster City, California, USA. Department of Pathology, NCI, NIH, Bethesda, Maryland, USA. Laboratory of Translational Genomics, Division of Cancer Epidemiology and Genetics, NCI, NIH, Bethesda, Maryland, USA
| | - Anthony S. Fauci
- Laboratory of Immunoregulation, NIAID, NIH, Bethesda, Maryland, USA. Division of Infectious Diseases, Institute of Human Virology, University of Maryland Medical School, Baltimore, Maryland, USA. Institute of Biostatistics and Mathematical Modeling, Johann Wolfgang Goethe University, Frankfurt, Germany. Genomics Unit, Research Technologies Section, Rocky Mountain Laboratories, NIAID, NIH, Hamilton, Montana, USA. Critical Care Medicine Department, Clinical Center, NIH, Bethesda, Maryland, USA. Gilead Sciences, Foster City, California, USA. Department of Pathology, NCI, NIH, Bethesda, Maryland, USA. Laboratory of Translational Genomics, Division of Cancer Epidemiology and Genetics, NCI, NIH, Bethesda, Maryland, USA
| | - Shyamasundaran Kottilil
- Laboratory of Immunoregulation, NIAID, NIH, Bethesda, Maryland, USA. Division of Infectious Diseases, Institute of Human Virology, University of Maryland Medical School, Baltimore, Maryland, USA. Institute of Biostatistics and Mathematical Modeling, Johann Wolfgang Goethe University, Frankfurt, Germany. Genomics Unit, Research Technologies Section, Rocky Mountain Laboratories, NIAID, NIH, Hamilton, Montana, USA. Critical Care Medicine Department, Clinical Center, NIH, Bethesda, Maryland, USA. Gilead Sciences, Foster City, California, USA. Department of Pathology, NCI, NIH, Bethesda, Maryland, USA. Laboratory of Translational Genomics, Division of Cancer Epidemiology and Genetics, NCI, NIH, Bethesda, Maryland, USA
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21
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Genotypic and phenotypic analyses of hepatitis C virus variants observed in clinical studies of VX-222, a nonnucleoside NS5B polymerase inhibitor. Antimicrob Agents Chemother 2014; 58:5456-65. [PMID: 24982088 DOI: 10.1128/aac.03052-14] [Citation(s) in RCA: 17] [Impact Index Per Article: 1.5] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/11/2022] Open
Abstract
VX-222, a thiophene-2-carboxylic acid derivative, is a selective nonnucleoside inhibitor of the hepatitis C virus (HCV) NS5B RNA-dependent RNA polymerase. In phase 1 and 2 clinical studies, VX-222 demonstrated effective antiviral efficacy, with substantial reductions in plasma HCV RNA in patients chronically infected with genotype 1 HCV. To characterize the potential for selection of VX-222-resistant variants in HCV-infected patients, the HCV NS5B gene was sequenced at baseline and during and after 3 days of VX-222 dosing (monotherapy) in a phase 1 study. Variants with the substitutions L419C/I/M/P/S/V, R422K, M423I/T/V, I482L/N/T, A486S/T/V, and V494A were selected during VX-222 dosing, and their levels declined over time after the end of dosing. Phenotypic analysis of these variants was conducted using HCV replicons carrying site-directed mutations. Of the 17 variants, 14 showed reduced susceptibility to VX-222 compared with the wild type, with the L419C/S and R422K variants having higher levels of resistance (>200-fold) than the rest of the variants (6.8- to 76-fold). The M423I and A486S variants remained susceptible to VX-222. The 50% effective concentration (EC50) for the L419P variant could not be obtained due to the poor replication of this replicon. The majority of the variants (15/17) were less fit than the wild type. A subset of the variants, predominately the L419S and R422K variants, were observed when the efficacy and safety of VX-222- and telaprevir-based regimens given for 12 weeks were investigated in genotype 1 HCV-infected patients in a phase 2 study. The NS3 and NS5B variants selected during the dual combination therapy showed reduced susceptibility to both telaprevir and VX-222 and had a lower replication capacity than the wild type. The phase 1b study has the ClinicalTrials.gov identifier NCT00911963, and the phase 2a study has ClinicalTrials.gov identifier NCT01080222.
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22
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Meissner EG, Nelson A, Marti M, Masur H, Osinusi A, Kottilil S. Sustained Virologic Response for Chronic Hepatitis C Infection after 27 Days of Treatment with Sofosbuvir and Ribavirin. Open Forum Infect Dis 2014; 1:013. [PMID: 25411655 PMCID: PMC4233673 DOI: 10.1093/ofid/ofu013] [Citation(s) in RCA: 11] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/30/2022] Open
Abstract
Successful treatment of chronic hepatitis C virus infection can now be achieved using direct-acting antiviral agents without interferon. In this report, we present a patient who achieved a sustained virologic response after 27 days of treatment with sofosbuvir and ribavirin. It is imperative to identify factors that allow for shorter treatment times in some individuals.
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Affiliation(s)
- Eric G Meissner
- Laboratory of Immunoregulation/NIAID/NIH, Bethesda, MD, 20892 USA
| | - Amy Nelson
- Laboratory of Immunoregulation/NIAID/NIH, Bethesda, MD, 20892 USA
| | - Miriam Marti
- Laboratory of Immunoregulation/NIAID/NIH, Bethesda, MD, 20892 USA
| | - Henry Masur
- Critical Care Medicine Department/Clinical Center/NIH, Bethesda, MD, 20892 USA
| | - Anu Osinusi
- Laboratory of Immunoregulation/NIAID/NIH, Bethesda, MD, 20892 USA ; Division of Infectious Diseases, Institute of Human Virology, University of Maryland Medical School, Baltimore, MD, USA
| | - Shyam Kottilil
- Laboratory of Immunoregulation/NIAID/NIH, Bethesda, MD, 20892 USA
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23
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Pfaender S, Brown RJ, Pietschmann T, Steinmann E. Natural reservoirs for homologs of hepatitis C virus. Emerg Microbes Infect 2014; 3:e21. [PMID: 26038514 PMCID: PMC3974340 DOI: 10.1038/emi.2014.19] [Citation(s) in RCA: 68] [Impact Index Per Article: 6.2] [Reference Citation Analysis] [Abstract] [Key Words] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/18/2013] [Revised: 01/07/2014] [Accepted: 01/22/2014] [Indexed: 12/31/2022]
Abstract
Hepatitis C virus is considered a major public health problem, infecting 2%–3% of the human population. Hepatitis C virus infection causes acute and chronic liver disease, including chronic hepatitis, cirrhosis and hepatocellular carcinoma. In fact, hepatitis C virus infection is the most frequent indication for liver transplantation and a vaccine is not available. Hepatitis C virus displays a narrow host species tropism, naturally infecting only humans, although chimpanzees are also susceptible to experimental infection. To date, there is no evidence for an animal reservoir of viruses closely related to hepatitis C virus which may have crossed the species barrier to cause disease in humans and resulted in the current pandemic. In fact, due to this restricted host range, a robust immunocompetent small animal model is still lacking, hampering mechanistic analysis of virus pathogenesis, immune control and prophylactic vaccine development. Recently, several studies discovered new viruses related to hepatitis C virus, belonging to the hepaci- and pegivirus genera, in small wild mammals (rodents and bats) and domesticated animals which live in close contact with humans (dogs and horses). Genetic and biological characterization of these newly discovered hepatitis C virus-like viruses infecting different mammals will contribute to our understanding of the origins of hepatitis C virus in humans and enhance our ability to study pathogenesis and immune responses using tractable animal models. In this review article, we start with an introduction on the genetic diversity of hepatitis C virus and then focus on the newly discovered viruses closely related to hepatitis C virus. Finally, we discuss possible theories about the origin of this important viral human pathogen.
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Affiliation(s)
- Stephanie Pfaender
- Institute for Experimental Virology, Twincore Centre of Experimental and Clinical Infection Research; a joint venture between the Hannover Medical School and the Helmholtz Centre for Infection Research , Hannover 30625, Germany
| | - Richard Jp Brown
- Institute for Experimental Virology, Twincore Centre of Experimental and Clinical Infection Research; a joint venture between the Hannover Medical School and the Helmholtz Centre for Infection Research , Hannover 30625, Germany
| | - Thomas Pietschmann
- Institute for Experimental Virology, Twincore Centre of Experimental and Clinical Infection Research; a joint venture between the Hannover Medical School and the Helmholtz Centre for Infection Research , Hannover 30625, Germany
| | - Eike Steinmann
- Institute for Experimental Virology, Twincore Centre of Experimental and Clinical Infection Research; a joint venture between the Hannover Medical School and the Helmholtz Centre for Infection Research , Hannover 30625, Germany
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24
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Blass B. HCV polymerase inhibitors. ACS Med Chem Lett 2014; 5:232-3. [PMID: 24900809 DOI: 10.1021/ml400428b] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/26/2013] [Indexed: 11/28/2022] Open
Affiliation(s)
- Benjamin Blass
- Temple University School of Pharmacy, 3307 North Broad Street, Philadelphia, Pennsylvania 19140, United States
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25
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Blass B. Antiviral compounds for the treatment of HCV. ACS Med Chem Lett 2014; 5:230-1. [PMID: 24900808 DOI: 10.1021/ml400426t] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/26/2013] [Indexed: 11/29/2022] Open
Affiliation(s)
- Benjamin Blass
- Temple University School of Pharmacy, 3307 North Broad Street, Philadelphia, Pennsylvania 19140, United States
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26
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Hepatitis C virus genetic variability and the presence of NS5B resistance-associated mutations as natural polymorphisms in selected genotypes could affect the response to NS5B inhibitors. Antimicrob Agents Chemother 2014; 58:2781-97. [PMID: 24590484 DOI: 10.1128/aac.02386-13] [Citation(s) in RCA: 50] [Impact Index Per Article: 4.5] [Reference Citation Analysis] [Abstract] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 02/06/2023] Open
Abstract
Because of the extreme genetic variability of hepatitis C virus (HCV), we analyzed the NS5B polymerase genetic variability in circulating HCV genotypes/subtypes and its impact on the genetic barrier for the development of resistance to clinically relevant nucleoside inhibitors (NIs)/nonnucleoside inhibitors (NNIs). The study included 1,145 NS5B polymerase sequences retrieved from the Los Alamos HCV database and GenBank. The genetic barrier was calculated for drug resistance emergence. Prevalence and genetic barrier were calculated for 1 major NI and 32 NNI resistance variants (13 major and 19 minor) at 21 total NS5B positions. Docking calculations were used to analyze sofosbuvir affinity toward the diverse HCV genotypes. Overall, NS5B polymerase was moderately conserved among all HCV genotypes, with 313/591 amino acid residues (53.0%) showing ≤1% variability and 83/591 residues (14.0%) showing high variability (≥25.1%). Nine NNI resistance variants (2 major variants, 414L and 423I; 7 minor variants, 316N, 421V, 445F, 482L, 494A, 499A, and 556G) were found as natural polymorphisms in selected genotypes. In particular, 414L and 423I were found in HCV genotype 4 (HCV-4) (n = 14/38, 36.8%) and in all HCV-5 sequences (n = 17, 100%), respectively. Regardless of HCV genotype, the 282T major NI resistance variant and 10 major NNI resistance variants (316Y, 414L, 423I/T/V, 448H, 486V, 495L, 554D, and 559G) always required a single nucleotide substitution to be generated. Conversely, the other 3 major NNI resistance variants (414T, 419S, and 422K) were associated with a different genetic barrier score development among the six HCV genotypes. Sofosbuvir docking analysis highlighted a better ligand affinity toward HCV-2 than toward HCV-3, in agreement with the experimental observations. The genetic variability among HCV genotypes, particularly with the presence of polymorphisms at NNI resistance positions, could affect their responsiveness to NS5B inhibitors. A pretherapy HCV NS5B sequencing could help to provide patients with the full efficacy of NNI-containing regimens.
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Lee CM, Chen CY, Chien RN, Tseng KC, Peng CY, Tung SY, Fang YJ, Huang YH, Lu SN, Hung CH, Tsai TJ, Fang CC, Hsu CW, Yeh CT. A double-blind randomized controlled study to evaluate the efficacy of low-dose oral interferon-alpha in preventing hepatitis C relapse. J Interferon Cytokine Res 2014; 34:187-94. [PMID: 24237300 PMCID: PMC3942686 DOI: 10.1089/jir.2013.0074] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/18/2013] [Accepted: 09/03/2013] [Indexed: 11/12/2022] Open
Abstract
Low-dose oral interferon could exert immune-modulating effects in human. We conducted a clinical trial to investigate the efficacy of oral interferon-alpha in preventing hepatitis C relapse. Totally 169 genotype 1b chronic hepatitis C patients having achieved end-of-therapy virological clearance were randomized to receive interferon-alpha lozenge 500 IU/day (n=59), 1,500 IU/day (n=53), or placebo (n=57) for 24 weeks. Overall, no significant differences were found for the relapse rates in the 3 groups (P>0.05). However, in patients with fibroindex 1.4-1.7, relapse occurred in 1/12 (8.3%) 500 IU-group patients versus 9/21 (42.9%) patients of the other groups (P=0.05). In 158 patients receiving at least 4 weeks of oral interferon, significantly higher platelet count was found at the end of trial in the 500 IU group (P=0.003). In thrombocytopenic patients, a significantly expedited recovery of platelet count was found in the 500 IU group (P=0.002). No drug-related severe adverse events were reported. In conclusion, at 500 IU/day, oral interferon exerted a borderline suppression effect of virological relapse in chronic hepatitis C patients with mild liver fibrosis. Additionally, it significantly expedited platelet count recovery after the end of peginterferon therapy.
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Affiliation(s)
- Chuan-Mo Lee
- Department of Hepato-Gastroenterology, Kaohsiung Chang Gung Memorial Hospital and Chang Gung University College of Medicine, Kaohsiung, Taiwan
| | - Chi-Yi Chen
- Department of Hepato-Gastroenterology, Chiayi Christian Hospital, Chiayi, Taiwan
| | - Rong-Nan Chien
- Liver Research Unit, Keelung Chang Gung Memorial Hospital, Keelung, Taiwan
| | - Kuo-Chih Tseng
- Department of Hepato-Gastroenterology, Buddhist Dalin Tzu Chi General Hospital, Chiayi and School of Medicine, Tzuchi University, Hualien, Taiwan
| | - Cheng-Yuan Peng
- Department of Hepato-Gastroenterology, China Medical University Hospital, Taichung, Taiwan
| | - Shui-Yi Tung
- Department of Hepato-Gastroenterology, Chiayi Chang Gung Memorial Hospital, Chiayi, Taiwan
| | - Yi-Jen Fang
- Department of Hepato-Gastroenterology, Show-Chwan Memorial Hospital, Changhua, Taiwan
| | - Yi-Hsiang Huang
- Division of Gastreoenterology, Taipei Veterans General Hospital and Institute of Clinical Medicine, National Yang-Ming University, Taipei, Taiwan
| | - Sheng-Nan Lu
- Department of Hepato-Gastroenterology, Kaohsiung Chang Gung Memorial Hospital and Chang Gung University College of Medicine, Kaohsiung, Taiwan
| | - Chao-Hung Hung
- Department of Hepato-Gastroenterology, Kaohsiung Chang Gung Memorial Hospital and Chang Gung University College of Medicine, Kaohsiung, Taiwan
| | - Tsung-Jang Tsai
- Department of Hepato-Gastroenterology, Chiayi Christian Hospital, Chiayi, Taiwan
| | - Chien-Chung Fang
- Department of Hepato-Gastroenterology, Chiayi Christian Hospital, Chiayi, Taiwan
| | - Chao-Wei Hsu
- Liver Research Center, Linko Chang Gung Memorial Hospital and School of Medicine, Chang Gung University, Taoyuan, Taiwan
| | - Chau-Ting Yeh
- Liver Research Center, Linko Chang Gung Memorial Hospital and School of Medicine, Chang Gung University, Taoyuan, Taiwan
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McWilliam Leitch EC, McLauchlan J. Determining the cellular diversity of hepatitis C virus quasispecies by single-cell viral sequencing. J Virol 2013; 87:12648-55. [PMID: 24049174 PMCID: PMC3838117 DOI: 10.1128/jvi.01602-13] [Citation(s) in RCA: 18] [Impact Index Per Article: 1.5] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/12/2013] [Accepted: 09/07/2013] [Indexed: 12/15/2022] Open
Abstract
Single-cell genomics is emerging as an important tool in cellular biology. We describe for the first time a system to investigate RNA virus quasispecies diversity at the cellular level utilizing hepatitis C virus (HCV) replicons. A high-fidelity nested reverse transcription (RT)-PCR assay was developed, and validation using control transcripts of known copy number indicated a detection limit of 3 copies of viral RNA/reaction. This system was used to determine the cellular diversity of subgenomic JFH-1 HCV replicons constitutively expressed in Huh7 cells. Each cell contained a unique quasispecies that was much less diverse than the quasispecies of the bulk cell population from which the single cells were derived, suggesting the occurrence of independent evolution at the cellular level. An assessment of the replicative fitness of the predominant single-cell quasispecies variants indicated a modest reduction in fitness compared to the wild type. Real-time RT-PCR methods capable of determining single-cell viral loads were developed and indicated an average of 113 copies of replicon RNA per cell, correlating with calculated RNA copy numbers in the bulk cell population. This study introduces a single-cell RNA viral-sequencing method with numerous potential applications to explore host-virus interactions during infection. HCV quasispecies diversity varied greatly between cells in vitro, suggesting different within-cell evolutionary pathways. Such divergent trajectories in vivo could have implications for the evolution and establishment of antiviral-resistant variants and host immune escape mutants.
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29
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Arciello M, Gori M, Balsano C. Mitochondrial dysfunctions and altered metals homeostasis: new weapons to counteract HCV-related oxidative stress. OXIDATIVE MEDICINE AND CELLULAR LONGEVITY 2013; 2013:971024. [PMID: 24371505 PMCID: PMC3859171 DOI: 10.1155/2013/971024] [Citation(s) in RCA: 21] [Impact Index Per Article: 1.8] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Subscribe] [Scholar Register] [Received: 06/10/2013] [Revised: 10/18/2013] [Accepted: 10/28/2013] [Indexed: 02/06/2023]
Abstract
The hepatitis C virus (HCV) infection produces several pathological effects in host organism through a wide number of molecular/metabolic pathways. Today it is worldwide accepted that oxidative stress actively participates in HCV pathology, even if the antioxidant therapies adopted until now were scarcely effective. HCV causes oxidative stress by a variety of processes, such as activation of prooxidant enzymes, weakening of antioxidant defenses, organelle damage, and metals unbalance. A focal point, in HCV-related oxidative stress onset, is the mitochondrial failure. These organelles, known to be the "power plants" of cells, have a central role in energy production, metabolism, and metals homeostasis, mainly copper and iron. Furthermore, mitochondria are direct viral targets, because many HCV proteins associate with them. They are the main intracellular free radicals producers and targets. Mitochondrial dysfunctions play a key role in the metal imbalance. This event, today overlooked, is involved in oxidative stress exacerbation and may play a role in HCV life cycle. In this review, we summarize the role of mitochondria and metals in HCV-related oxidative stress, highlighting the need to consider their deregulation in the HCV-related liver damage and in the antiviral management of patients.
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Affiliation(s)
- Mario Arciello
- Department of Internal Medicine and Medical Specialties, “Sapienza” University of Rome, Via del Policlinico 155, 00161 Rome, Italy
- Francesco Balsano Foundation, Via G.B. Martini 6, 00198 Rome, Italy
| | - Manuele Gori
- Francesco Balsano Foundation, Via G.B. Martini 6, 00198 Rome, Italy
| | - Clara Balsano
- Francesco Balsano Foundation, Via G.B. Martini 6, 00198 Rome, Italy
- Institute of Molecular Biology and Pathology (IBPM); CNR, Piazzale Aldo Moro 7, 00185 Rome, Italy
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30
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A novel dengue virus inhibitor, BP13944, discovered by high-throughput screening with dengue virus replicon cells selects for resistance in the viral NS2B/NS3 protease. Antimicrob Agents Chemother 2013; 58:110-9. [PMID: 24145533 DOI: 10.1128/aac.01281-13] [Citation(s) in RCA: 65] [Impact Index Per Article: 5.4] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/18/2022] Open
Abstract
Dengue virus (DENV) causes disease globally, resulting in an estimated 25 to 100 million new infections per year. No effective DENV vaccine is available, and the current treatment is only supportive. Thus, there is an urgent need to develop therapeutic agents to cure this epidemic disease. In the present study, we identified a potential small-molecule inhibitor, BP13944, via high-throughput screening (HTS) of 60,000 compounds using a stable cell line harboring an efficient luciferase replicon of DENV serotype 2 (DENV-2). BP13944 reduced the expression of the DENV replicon reporter in cells, showing a 50% effective concentration (EC50) of 1.03 ± 0.09 μM. Without detectable cytotoxicity, the compound inhibited replication or viral RNA synthesis in all four serotypes of DENV but not in Japanese encephalitis virus (JEV). Sequencing analyses of several individual clones derived from BP13944-resistant RNAs purified from cells harboring the DENV-2 replicon revealed a consensus amino acid substitution (E66G) in the region of the NS3 protease domain. Introduction of E66G into the DENV replicon, an infectious DENV cDNA clone, and recombinant NS2B/NS3 protease constructs conferred 15.2-, 17.2-, and 3.1-fold resistance to BP13944, respectively. Our results identify an effective small-molecule inhibitor, BP13944, which likely targets the DENV NS3 protease. BP13944 could be considered part of a more effective treatment regime for inhibiting DENV in the future.
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