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Liu T, Wang H, Zhao Y, Wang YX, Xing X, Gao P. Drug development for chronic hepatitis B functional cure: Recent progress. World J Hepatol 2025; 17:105797. [PMID: 40308829 PMCID: PMC12038417 DOI: 10.4254/wjh.v17.i4.105797] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 02/08/2025] [Revised: 03/21/2025] [Accepted: 04/09/2025] [Indexed: 04/25/2025] Open
Abstract
Chronic hepatitis B virus (HBV) infection affects approximately 254 million individuals globally, contributing to significant morbidity and mortality due to HBV-related liver failure and cirrhosis, which result in millions of fatalities each year. Although approved antiviral nucleos(t)ide analogues can effectively suppress HBV replication, their ability to reduce hepatitis B surface antigen (HBsAg) levels in plasma remains limited. The clinical application of the immunomodulator interferon-alpha is restricted by concerns regarding its safety and the severity of associated adverse reactions, rendering long-term administration challenging. Therefore, current drug development efforts for chronic hepatitis B aim to achieve a functional cure, which is defined as HBsAg serological clearance and sustained suppression of HBV DNA. This review discusses recent advancements in novel direct-acting therapeutic strategies for the treatment of chronic hepatitis B by focusing on the progresses in HBV entry inhibitors, monoclonal antibodies, RNA interferences, and other agents that directly target the virus. Furthermore, we discuss the development of immunomodulatory therapies, including TLR-7/8 agonists, immune checkpoint inhibitors, and therapeutic vaccines. In the end, we conclude by highlighting the importance of the rational combination-strategy design to improve the functional cure rate of HBV.
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Affiliation(s)
- Ting Liu
- Department of Clinical Laboratory, The Second Affiliated Hospital of Dalian Medical University, Dalian 116021, Liaoning Province, China
| | - He Wang
- Department of Clinical Laboratory, The Second Affiliated Hospital of Dalian Medical University, Dalian 116021, Liaoning Province, China
| | - Yue Zhao
- Graduate School Base Office, Dalian Medical University, Dalian 116000, Liaoning Province, China
| | - Ying-Xin Wang
- Department of Clinical Laboratory, The Second Affiliated Hospital of Dalian Medical University, Dalian 116021, Liaoning Province, China
| | - Xue Xing
- Department of Clinical Laboratory, The Second Affiliated Hospital of Dalian Medical University, Dalian 116021, Liaoning Province, China
| | - Peng Gao
- Department of Clinical Laboratory, The Second Affiliated Hospital of Dalian Medical University, Dalian 116021, Liaoning Province, China.
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Shekhtman L, Ishida Y, Tsuge M, Reinharz V, Yamao M, Takahashi M, Tateno C, Uprichard SL, Dahari H, Chayama K. Modeling of hepatitis B virus kinetics and accumulation of cccDNA in primary human hepatocytes. JHEP Rep 2025; 7:101311. [PMID: 40236628 PMCID: PMC11997607 DOI: 10.1016/j.jhepr.2024.101311] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 11/05/2024] [Revised: 12/11/2024] [Accepted: 12/16/2024] [Indexed: 04/17/2025] Open
Abstract
Background & Aims Knowledge about early HBV covalently closed circular DNA (cccDNA) accumulation post infection is lacking. We characterized and mathematically modeled HBV infection kinetics during early infection and treatment in primary human hepatocytes (PHHs). Methods PHHs were inoculated with HBV, and infection was monitored with and without treatment with the nucleoside analog entecavir (ETV), the HBV-entry inhibitor Myr-preS1, or both ETV + Myr-preS1. Extracellular HBV DNA (exHBV), total intracellular HBV DNA (inHBV), and cccDNA were frequently measured during the 12 days post inoculation. A multicompartmental mathematical model was developed to explain HBV infection dynamics. Results Multiphasic exHBV and inHBV kinetics were overall similar in untreated and Myr-preS1-treated PHHs. In ETV-treated PHHs (either alone or with Myr-preS1), exHBV and inHBV initially declined and did not resurge. ETV-treated cultures had significantly (p=0.002) lower mean cccDNA levels at Day 2 post inoculation (4.3 ± 0.1 vs. 4.7 ± 0.1) and reached plateau slower (5 vs. 2 days) than untreated and Myr-preS1-treated cultures, respectively. Modeling predicts that the recycling of inHBV into cccDNA stops when cccDNA reaches a maximum and HBV secretion changes depending on the concentration of inHBV. Even when initiated at the time of inoculation, ETV did not prevent or eradicate infection but rather blocked inHBV accumulation gradually, reaching 97% efficacy by the end of the 12-day experiment and resulting in an average 44% slower cccDNA accumulation. Conclusions The study provides insight into the interrelationships and dynamics of cccDNA accumulation, inHBV accumulation, and secretion of exHBV containing particles. Although kinetics and modeling support the conclusion that the level of cccDNA in the cell is regulated, the mechanisms that determine HBV capsid secretion vs. recycling to the nucleus for cccDNA accumulation require further investigation. Impact and implications Using primary human hepatocytes (PHHs), we characterize early HBV kinetics post infection. HBV-infected PHH were treated with an entry inhibitor to characterize the accumulation of intracellular and extracellular HBV DNA and the nuclear episomal viral genome called covalently closed circular DNA (cccDNA) in the absence of HBV spread. Kinetic and mathematical modeling in PHHs confirms that the replication inhibitor, entecavir, strongly blocks intracellular HBV DNA accumulation, which also slowed the accumulation of cccDNA. However, modeling indicates that effects on cccDNA accumulation are not directly determined by intracellular HBV DNA levels, supporting the conclusion that cccDNA levels within the cell are regulated in some yet-to-be-elucidated manner.
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Affiliation(s)
- Louis Shekhtman
- The Program for Experimental and Theoretical Modeling, Division of Hepatology, Department of Medicine, Loyola University Medical Center, Maywood, IL, USA
- Department of Information Science, Bar-Ilan University, Ramat Gan, Israel
| | - Yuji Ishida
- PhoenixBio Co., Ltd., Higashi-Hiroshima, Japan
| | - Masataka Tsuge
- Liver Center, Hiroshima University Hospital, Hiroshima, Japan
- Department of Gastroenterology, Graduate School of Biomedical and Health Sciences, Hiroshima University, Hiroshima, Japan
| | - Vladimir Reinharz
- Department of Computer Science, Université du Québec à Montréal, Montréal, QC, Canada
| | | | | | | | - Susan L. Uprichard
- The Program for Experimental and Theoretical Modeling, Division of Hepatology, Department of Medicine, Loyola University Medical Center, Maywood, IL, USA
| | - Harel Dahari
- The Program for Experimental and Theoretical Modeling, Division of Hepatology, Department of Medicine, Loyola University Medical Center, Maywood, IL, USA
| | - Kazuaki Chayama
- Hiroshima Institute of Life Sciences, Hiroshima, Japan
- RIKEN Center for Integrative Medical Sciences, Yokohama, Japan
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Jin Y, Wang S, Tang K, Zhan P, Liu X. Recent advances in screening methods enabling the discovery of novel anti-hepatitis B virus drug candidates. Eur J Med Chem 2025; 282:117093. [PMID: 39612566 DOI: 10.1016/j.ejmech.2024.117093] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/11/2024] [Revised: 11/15/2024] [Accepted: 11/21/2024] [Indexed: 12/01/2024]
Abstract
The global population affected by Hepatitis B virus (HBV) is approximately 296 million, but few drugs have been able to completely eradicate HBV and the range of effective treatments remains limited. Recent advancements in molecular biology and artificial intelligence, as well as a comprehensive understanding of the molecular structure of HBV, have greatly aided the rational development of anti-HBV agents. Such advancements have facilitated an increasing array of candidate drugs transitioning into clinical trials, however, no novel target-based compounds have been approved for clinical application. To expedite the progression of anti-HBV drug development, establishing a reliable and robust in vitro HBV infection system is of great importance. However, owing to the host and tissue specificity of HBV, identifying a stable and dependable cell culture system for screening all anti-HBV agents poses significant challenges. In this review, we summarize recent advances in screening methods for small-molecule inhibitors that target key stages of the HBV replication cycle from a medicinal chemistry perspective.
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Affiliation(s)
- Yu Jin
- Department of Medicinal Chemistry, Key Laboratory of Chemical Biology (Ministry of Education), School of Pharmaceutical Sciences, Shandong University, 44 West Culture Road, 250012, Jinan, Shandong, PR China
| | - Shuo Wang
- Department of Medicinal Chemistry, Key Laboratory of Chemical Biology (Ministry of Education), School of Pharmaceutical Sciences, Shandong University, 44 West Culture Road, 250012, Jinan, Shandong, PR China
| | - Kai Tang
- Department of Medicinal Chemistry, Key Laboratory of Chemical Biology (Ministry of Education), School of Pharmaceutical Sciences, Shandong University, 44 West Culture Road, 250012, Jinan, Shandong, PR China
| | - Peng Zhan
- Department of Medicinal Chemistry, Key Laboratory of Chemical Biology (Ministry of Education), School of Pharmaceutical Sciences, Shandong University, 44 West Culture Road, 250012, Jinan, Shandong, PR China.
| | - Xinyong Liu
- Department of Medicinal Chemistry, Key Laboratory of Chemical Biology (Ministry of Education), School of Pharmaceutical Sciences, Shandong University, 44 West Culture Road, 250012, Jinan, Shandong, PR China.
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Li S, Hao L, Deng J, Zhang J, Yu F, Ye F, Li N, Hu X. The Culprit Behind HBV-Infected Hepatocytes: NTCP. Drug Des Devel Ther 2024; 18:4839-4858. [PMID: 39494152 PMCID: PMC11529284 DOI: 10.2147/dddt.s480151] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/27/2024] [Accepted: 10/16/2024] [Indexed: 11/05/2024] Open
Abstract
Hepatitis B virus (HBV) is a globally prevalent human DNA virus responsible for over 250 million cases of chronic liver infections, leading to conditions such as liver inflammation, cirrhosis and hepatocellular carcinoma (HCC). Sodium taurocholate co-transporting polypeptide (NTCP) is a transmembrane protein highly expressed in human hepatocytes and functions as a bile acid (BA) transporter. NTCP has been identified as the receptor that HBV and its satellite virus, hepatitis delta virus (HDV), use to enter hepatocytes. HBV entry into hepatocytes is tightly regulated by various signaling pathways, and NTCP plays an important role as the initial stage of HBV infection. NTCP acts as an initiation signal, causing metabolic changes in hepatocytes and facilitating the entry of HBV into hepatocytes. Thus, a comprehensive understanding of NTCP's role is crucial. In this review, we will examine the regulatory mechanisms governing HBV pre-S1 binding to liver membrane NTCP, the role of NTCP in HBV internalization, and the transcriptional and translational regulation of NTCP expression. Additionally, we will discuss clinical drugs targeting NTCP, including combination therapies involving NTCP inhibitors, and consider the safety of NTCP as a therapeutic target.
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Affiliation(s)
- Shenghao Li
- School of Clinical Medicine, Chengdu University of Traditional Chinese Medicine, Chengdu, Sichuan Province, People’s Republic of China
- Department of Infectious Diseases, Hospital of Chengdu University of Traditional Chinese Medicine, Chengdu, Sichuan Province, People’s Republic of China
| | - Liyuan Hao
- School of Clinical Medicine, Chengdu University of Traditional Chinese Medicine, Chengdu, Sichuan Province, People’s Republic of China
- Department of Infectious Diseases, Hospital of Chengdu University of Traditional Chinese Medicine, Chengdu, Sichuan Province, People’s Republic of China
| | - Jiali Deng
- School of Clinical Medicine, Chengdu University of Traditional Chinese Medicine, Chengdu, Sichuan Province, People’s Republic of China
- Department of Infectious Diseases, Hospital of Chengdu University of Traditional Chinese Medicine, Chengdu, Sichuan Province, People’s Republic of China
| | - Junli Zhang
- Department of Infectious Diseases, Jiangsu Province Hospital of Chinese Medicine, Nanjing, Jiangsu Province, People’s Republic of China
| | - Fei Yu
- School of Clinical Medicine, Chengdu University of Traditional Chinese Medicine, Chengdu, Sichuan Province, People’s Republic of China
- Department of Infectious Diseases, Hospital of Chengdu University of Traditional Chinese Medicine, Chengdu, Sichuan Province, People’s Republic of China
| | - Fanghang Ye
- School of Clinical Medicine, Chengdu University of Traditional Chinese Medicine, Chengdu, Sichuan Province, People’s Republic of China
- Department of Infectious Diseases, Hospital of Chengdu University of Traditional Chinese Medicine, Chengdu, Sichuan Province, People’s Republic of China
| | - Na Li
- School of Clinical Medicine, Chengdu University of Traditional Chinese Medicine, Chengdu, Sichuan Province, People’s Republic of China
- Department of Infectious Diseases, Hospital of Chengdu University of Traditional Chinese Medicine, Chengdu, Sichuan Province, People’s Republic of China
| | - Xiaoyu Hu
- Department of Infectious Diseases, Hospital of Chengdu University of Traditional Chinese Medicine, Chengdu, Sichuan Province, People’s Republic of China
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Xu ZM, Gnouamozi GE, Rüeger S, Shea PR, Buti M, Chan HL, Marcellin P, Lawless D, Naret O, Zeller M, Schneuing A, Scheck A, Junier T, Moradpour D, Podlaha O, Suri V, Gaggar A, Subramanian M, Correia B, Gfeller D, Urban S, Fellay J. Joint host-pathogen genomic analysis identifies hepatitis B virus mutations associated with human NTCP and HLA class I variation. Am J Hum Genet 2024; 111:1018-1034. [PMID: 38749427 PMCID: PMC11179264 DOI: 10.1016/j.ajhg.2024.04.013] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/08/2024] [Revised: 04/17/2024] [Accepted: 04/18/2024] [Indexed: 06/09/2024] Open
Abstract
Evolutionary changes in the hepatitis B virus (HBV) genome could reflect its adaptation to host-induced selective pressure. Leveraging paired human exome and ultra-deep HBV genome-sequencing data from 567 affected individuals with chronic hepatitis B, we comprehensively searched for the signatures of this evolutionary process by conducting "genome-to-genome" association tests between all human genetic variants and viral mutations. We identified significant associations between an East Asian-specific missense variant in the gene encoding the HBV entry receptor NTCP (rs2296651, NTCP S267F) and mutations within the receptor-binding region of HBV preS1. Through in silico modeling and in vitro preS1-NTCP binding assays, we observed that the associated HBV mutations are in proximity to the NTCP variant when bound and together partially increase binding affinity to NTCP S267F. Furthermore, we identified significant associations between HLA-A variation and viral mutations in HLA-A-restricted T cell epitopes. We used in silico binding prediction tools to evaluate the impact of the associated HBV mutations on HLA presentation and observed that mutations that result in weaker binding affinities to their cognate HLA alleles were enriched. Overall, our results suggest the emergence of HBV escape mutations that might alter the interaction between HBV PreS1 and its cellular receptor NTCP during viral entry into hepatocytes and confirm the role of HLA class I restriction in inducing HBV epitope variations.
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Affiliation(s)
- Zhi Ming Xu
- School of Life Sciences, École Polytechnique Fédérale de Lausanne, Lausanne, Switzerland; Swiss Institute of Bioinformatics, Lausanne, Switzerland
| | - Gnimah Eva Gnouamozi
- Department of Infectious Diseases, Molecular Virology, University Hospital Heidelberg, Heidelberg, Germany
| | - Sina Rüeger
- School of Life Sciences, École Polytechnique Fédérale de Lausanne, Lausanne, Switzerland; Swiss Institute of Bioinformatics, Lausanne, Switzerland
| | - Patrick R Shea
- Institute for Genomic Medicine, Columbia University, New York, NY, USA
| | - Maria Buti
- Liver Unit, Hospital Universitario Vall d'Hebron and CIBEREHD del Instituto Carlos III, Barcelona, Spain
| | - Henry Ly Chan
- The Chinese University of Hong Kong, Hong Kong, China
| | | | - Dylan Lawless
- School of Life Sciences, École Polytechnique Fédérale de Lausanne, Lausanne, Switzerland; Swiss Institute of Bioinformatics, Lausanne, Switzerland
| | - Olivier Naret
- School of Life Sciences, École Polytechnique Fédérale de Lausanne, Lausanne, Switzerland; Swiss Institute of Bioinformatics, Lausanne, Switzerland
| | - Matthias Zeller
- School of Life Sciences, École Polytechnique Fédérale de Lausanne, Lausanne, Switzerland
| | - Arne Schneuing
- School of Life Sciences, École Polytechnique Fédérale de Lausanne, Lausanne, Switzerland
| | - Andreas Scheck
- School of Life Sciences, École Polytechnique Fédérale de Lausanne, Lausanne, Switzerland
| | - Thomas Junier
- School of Life Sciences, École Polytechnique Fédérale de Lausanne, Lausanne, Switzerland; Swiss Institute of Bioinformatics, Lausanne, Switzerland
| | - Darius Moradpour
- Division of Gastroenterology and Hepatology, Lausanne University Hospital and University of Lausanne, Lausanne, Switzerland
| | | | | | | | | | - Bruno Correia
- School of Life Sciences, École Polytechnique Fédérale de Lausanne, Lausanne, Switzerland
| | - David Gfeller
- Department of Oncology UNIL-CHUV, Lausanne University Hospital, Ludwig Institute for Cancer Research, University of Lausanne, Lausanne, Switzerland; Swiss Institute of Bioinformatics, Lausanne, Switzerland
| | - Stephan Urban
- Department of Infectious Diseases, Molecular Virology, University Hospital Heidelberg, Heidelberg, Germany; German Center for Infection Research (DZIF), Partner Site Heidelberg, Heidelberg, Germany
| | - Jacques Fellay
- School of Life Sciences, École Polytechnique Fédérale de Lausanne, Lausanne, Switzerland; Swiss Institute of Bioinformatics, Lausanne, Switzerland; Precision Medicine Unit, Biomedical Data Science Center, Lausanne University Hospital and University of Lausanne, Lausanne, Switzerland.
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Yu X, Gao Y, Zhang X, Ji L, Fang M, Li M, Gao Y. Hepatitis B: Model Systems and Therapeutic Approaches. J Immunol Res 2024; 2024:4722047. [PMID: 38745751 PMCID: PMC11093688 DOI: 10.1155/2024/4722047] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/06/2023] [Revised: 03/29/2024] [Accepted: 04/05/2024] [Indexed: 05/16/2024] Open
Abstract
Hepatitis B virus (HBV) infection is a major global health issue and ranks among the top causes of liver cirrhosis and hepatocellular carcinoma. Although current antiviral medications, including nucleot(s)ide analogs and interferons, could inhibit the replication of HBV and alleviate the disease, HBV cannot be fully eradicated. The development of cellular and animal models for HBV infection plays an important role in exploring effective anti-HBV medicine. During the past decades, advancements in several cell culture systems, such as HepG2.2.15, HepAD38, HepaRG, hepatocyte-like cells, and primary human hepatocytes, have propelled the research in inhibiting HBV replication and expression and thus enriched our comprehension of the viral life cycle and enhancing antiviral drug evaluation efficacy. Mouse models, in particular, have emerged as the most extensively studied HBV animal models. Additionally, the present landscape of HBV therapeutics research now encompasses a comprehensive assessment of the virus's life cycle, targeting numerous facets and employing a variety of immunomodulatory approaches, including entry inhibitors, strategies aimed at cccDNA, RNA interference technologies, toll-like receptor agonists, and, notably, traditional Chinese medicine (TCM). This review describes the attributes and limitations of existing HBV model systems and surveys novel advancements in HBV treatment modalities, which will offer deeper insights toward discovering potentially efficacious pharmaceutical interventions.
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Affiliation(s)
- Xiaoxiao Yu
- Laboratory of Cellular Immunity, Shuguang Hospital, Affiliated to Shanghai University of Traditional Chinese Medicine, Shanghai, China
| | - Yating Gao
- Laboratory of Cellular Immunity, Shuguang Hospital, Affiliated to Shanghai University of Traditional Chinese Medicine, Shanghai, China
| | - Xin Zhang
- Laboratory of Cellular Immunity, Shuguang Hospital, Affiliated to Shanghai University of Traditional Chinese Medicine, Shanghai, China
| | - Longshan Ji
- Laboratory of Cellular Immunity, Shuguang Hospital, Affiliated to Shanghai University of Traditional Chinese Medicine, Shanghai, China
| | - Miao Fang
- Laboratory of Cellular Immunity, Shuguang Hospital, Affiliated to Shanghai University of Traditional Chinese Medicine, Shanghai, China
| | - Man Li
- Laboratory of Cellular Immunity, Shuguang Hospital, Affiliated to Shanghai University of Traditional Chinese Medicine, Shanghai, China
| | - Yueqiu Gao
- Laboratory of Cellular Immunity, Shuguang Hospital, Affiliated to Shanghai University of Traditional Chinese Medicine, Shanghai, China
- Department of Hepatopathy, Shuguang Hospital, Affiliated to Shanghai University of Traditional Chinese Medicine, Shanghai, China
- Institute of Infectious Diseases of Integrated Traditional Chinese and Western Medicine, Shanghai, China
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Roll V, Diesendorf V, Roewer N, Abdelgawad A, Roewer J, Trimpert J, Bodem J. A systematic analysis of anthocyanins inhibiting human, murine, and equine herpesviruses. PHYTOMEDICINE : INTERNATIONAL JOURNAL OF PHYTOTHERAPY AND PHYTOPHARMACOLOGY 2024; 124:155314. [PMID: 38190783 DOI: 10.1016/j.phymed.2023.155314] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Subscribe] [Scholar Register] [Received: 08/14/2023] [Revised: 11/22/2023] [Accepted: 12/25/2023] [Indexed: 01/10/2024]
Abstract
BACKGROUND Herpesviruses are common animal and human pathogens that cause severe health problems in children, immunocompromised patients, and infected animals with a host range from fish to mammals. Anthocyanin-containing plant extracts have been described as potent antivirals, which might cause fewer harmful side effects than direct-acting antivirals. Here, we report that an extract of Aristotelia chilensis (Molina) Stuntz (Elaeocarpaceae) (MBE) with a high content of the anthocyanin delphinidin suppresses lytic replication of equine, murine and human herpesviruses of replication in vitro. METHODS We treated cultured cells with MBE and purified individual anthocyanins present in the extract to determine the most active compound at different concentrations. We subsequently infected the cultures with human herpesviruses 1 (HSV-1) or 8 (HHV-8), murine cytomegalovirus (CMV), or equine herpesviruses 1 (EHV-1) and determined the number of infected cells and viral infectivity. RESULTS MBE inhibited the HSV-1, murine CMV, and EHV-1 by up to 2 orders of magnitude. In the presence of the stabilizing randomly methylated-beta-cyclodextrin, the inhibitory concentration could be lowered significantly. We identified delphinidin as an active antiviral compound and showed that the non-glycosylated delphinidin solved and stabilized with sulfobutylether-beta-cyclodextrin allowed usage of approximately 50 times lower concentrations. CONCLUSION Glycosylated delphinidin derivatives were identified as active antiviral compounds of MBE. This suggests that plant extracts rich in delphinidin-anthocyanins have potent antiviral properties that could be used in treatment and prevention.
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Affiliation(s)
- Valeria Roll
- Julius-Maximilians-University of Würzburg, Institute for Virology and Immunobiology, Versbacher Strasse 7, 97078 Würzburg, Germany
| | - Viktoria Diesendorf
- Julius-Maximilians-University of Würzburg, Institute for Virology and Immunobiology, Versbacher Strasse 7, 97078 Würzburg, Germany
| | - Norbert Roewer
- Universitätsklinikum Würzburg, Josef-Schneider-Str. 2, 97080 Würzburg, Germany
| | - Azza Abdelgawad
- Freie Universität Berlin, Institut für Virologie, Robert von Ostertag-Str. 7, 14163 Berlin, Germany
| | - Joachim Roewer
- Dermatologikum Hamburg, Stephansplatz 5, 20354 Hamburg, Germany
| | - Jakob Trimpert
- Freie Universität Berlin, Institut für Virologie, Robert von Ostertag-Str. 7, 14163 Berlin, Germany
| | - Jochen Bodem
- Julius-Maximilians-University of Würzburg, Institute for Virology and Immunobiology, Versbacher Strasse 7, 97078 Würzburg, Germany.
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Hu JL, Huang AL. Classifying hepatitis B therapies with insights from covalently closed circular DNA dynamics. Virol Sin 2024; 39:9-23. [PMID: 38110037 PMCID: PMC10877440 DOI: 10.1016/j.virs.2023.12.005] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/29/2023] [Accepted: 12/13/2023] [Indexed: 12/20/2023] Open
Abstract
The achievement of a functional cure for chronic hepatitis B (CHB) remains limited to a minority of patients treated with currently approved drugs. The primary objective in developing new anti-HBV drugs is to enhance the functional cure rates for CHB. A critical prerequisite for the functional cure of CHB is a substantial reduction, or even eradication of covalently closed circular DNA (cccDNA). Within this context, the changes in cccDNA levels during treatment become as a pivotal concern. We have previously analyzed the factors influencing cccDNA dynamics and introduced a preliminary classification of hepatitis B treatment strategies based on these dynamics. In this review, we employ a systems thinking perspective to elucidate the fundamental aspects of the HBV replication cycle and to rationalize the classification of treatment strategies according to their impact on the dynamic equilibrium of cccDNA. Building upon this foundation, we categorize current anti-HBV strategies into two distinct groups and advocate for their combined use to significantly reduce cccDNA levels within a well-defined timeframe.
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Affiliation(s)
- Jie-Li Hu
- Key Laboratory of Molecular Biology on Infectious Diseases, Ministry of Education, Chongqing Medical University, Chongqing, 400016, China.
| | - Ai-Long Huang
- Key Laboratory of Molecular Biology on Infectious Diseases, Ministry of Education, Chongqing Medical University, Chongqing, 400016, China.
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9
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Zhang Z, Zhang Q, Zhang Y, Lou Y, Ge L, Zhang W, Zhang W, Song F, Huang P. Role of sodium taurocholate cotransporting polypeptide (NTCP) in HBV-induced hepatitis: Opportunities for developing novel therapeutics. Biochem Pharmacol 2024; 219:115956. [PMID: 38049009 DOI: 10.1016/j.bcp.2023.115956] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/23/2023] [Revised: 11/29/2023] [Accepted: 11/29/2023] [Indexed: 12/06/2023]
Abstract
Hepatitis B is an infectious disease caused by the HBV virus. It presents a significant challenge for treatment due to its chronic nature and the potential for developing severe complications, including hepatocirrhosis and hepatocellular carcinoma. These complications not only cause physical and psychological distress to patients but also impose substantial economic and social burdens on both individuals and society as a whole. The internalization of HBV relies on endocytosis and necessitates the involvement of various proteins, including heparin sulfate proteoglycans, epidermal growth factor receptors, and NTCP. Among these proteins, NTCP is pivotal in HBV internalization and is primarily located in the liver's basement membrane. As a transporter of bile acids, NTCP also serves as a receptor facilitating HBV entry into cells. Numerous molecules have been identified to thwart HBV infection by stifling NTCP activity, although only a handful exhibit low IC50 values. In this systematic review, our primary focus dwells on the structure and regulation of NTCP, as well as the mechanism involved in HBV internalization. We underscore recent drug breakthroughs that specifically target NTCP to combat HBV infection. By shedding light on these advances, this review contributes novel insights into developing effective anti-HBV medications.
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Affiliation(s)
- Zhentao Zhang
- Department of Pharmacology, College of Pharmaceutical Sciences, Zhejiang University of Technology, Hangzhou, China
| | - Qi Zhang
- Department of Pharmacology, College of Pharmaceutical Sciences, Zhejiang University of Technology, Hangzhou, China
| | - Yiwen Zhang
- Center for Clinical Pharmacy, Cancer Center, Department of Pharmacy, Zhejiang Provincial People's Hospital (Affiliated People's Hospital), Hangzhou Medical College, Hangzhou, Zhejiang, China; Key Laboratory of Endocrine Gland Diseases of Zhejiang Province, Hangzhou, China
| | - Yutao Lou
- Department of Pharmacology, College of Pharmaceutical Sciences, Zhejiang University of Technology, Hangzhou, China
| | - Luqi Ge
- Department of Pharmacology, College of Pharmaceutical Sciences, Zhejiang University of Technology, Hangzhou, China
| | - Wanli Zhang
- Department of Pharmacology, College of Pharmaceutical Sciences, Zhejiang University of Technology, Hangzhou, China
| | - Wen Zhang
- Department of Pharmacology, College of Pharmaceutical Sciences, Zhejiang University of Technology, Hangzhou, China
| | - Feifeng Song
- Center for Clinical Pharmacy, Cancer Center, Department of Pharmacy, Zhejiang Provincial People's Hospital (Affiliated People's Hospital), Hangzhou Medical College, Hangzhou, Zhejiang, China; Key Laboratory of Endocrine Gland Diseases of Zhejiang Province, Hangzhou, China.
| | - Ping Huang
- Center for Clinical Pharmacy, Cancer Center, Department of Pharmacy, Zhejiang Provincial People's Hospital (Affiliated People's Hospital), Hangzhou Medical College, Hangzhou, Zhejiang, China; Key Laboratory of Endocrine Gland Diseases of Zhejiang Province, Hangzhou, China.
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10
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Abdelwahed AH, Heineman BD, Wu GY. Novel Approaches to Inhibition of HBsAg Expression from cccDNA and Chromosomal Integrants: A Review. J Clin Transl Hepatol 2023; 11:1485-1497. [PMID: 38161502 PMCID: PMC10752814 DOI: 10.14218/jcth.2023.00067] [Citation(s) in RCA: 2] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 02/15/2023] [Revised: 07/19/2023] [Accepted: 08/16/2023] [Indexed: 01/03/2024] Open
Abstract
Hepatitis B virus (HBV) is a widely prevalent liver infection that can cause acute or chronic hepatitis. Although current treatment modalities are highly effective in the suppression of viral levels, they cannot eliminate the virus or achieve definitive cure. This is a consequence of the complex nature of HBV-host interactions. Major challenges to achieving sustained viral suppression include the presence of a high viral burden from the HBV DNA and hepatitis B surface antigen (HBsAg), the presence of reservoirs for HBV replication and antigen production, and the HBV-impaired innate and adaptive immune response of the host. Those therapeutic methods include cell entry inhibitors, HBsAg inhibitors, gene editing approaches, immune-targeting therapies and direct inhibitors of covalently closed circular DNA (cccDNA). Novel approaches that target these key mechanisms are now being studied in preclinical and clinical phases. In this review article, we provide a comprehensive review on mechanisms by which HBV escapes elimination from current treatments, and highlight new agents to achieve a definitive HBV cure.
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Affiliation(s)
- Ahmed H. Abdelwahed
- Department of Medicine, Division of Gastroenterology-Hepatology, University of Connecticut Health Center, Farmington, CT, USA
| | - Brent D. Heineman
- Department of Medicine, Division of Gastroenterology-Hepatology, University of Connecticut Health Center, Farmington, CT, USA
| | - George Y. Wu
- Department of Medicine, Division of Gastroenterology-Hepatology, University of Connecticut Health Center, Farmington, CT, USA
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11
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Sandmann L, Berg T, Deterding K, Fischer N, Hinrichsen H, Petersen J, Tacke F, Cornberg M. Addendum „Antivirale Therapie der chronischen Hepatitis-D-Virusinfektion“ zur S3-Leitlinie „Prophylaxe, Diagnostik und Therapie der Hepatitis-B-Virusinfektion“ der Deutschen Gesellschaft für Gastroenterologie, Verdauungs- und Stoffwechselkrankheiten (DGVS). ZEITSCHRIFT FUR GASTROENTEROLOGIE 2023; 61:1635-1653. [PMID: 38081179 DOI: 10.1055/a-2181-3046] [Citation(s) in RCA: 2] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [MESH Headings] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 12/18/2023]
Affiliation(s)
- Lisa Sandmann
- Klinik für Gastroenterologie, Hepatologie, Infektiologie und Endokrinologie, Medizinische Hochschule Hannover, Hannover, Deutschland
- D-SOLVE Consortium, Horizon Europe Project, partner-site Medizinische Hochschule Hannover, Hannover, Deutschland
| | - Thomas Berg
- Bereich Hepatologie, Medizinische Klinik II, Universitätsklinikum Leipzig, Leipzig, Deutschland
| | - Katja Deterding
- Klinik für Gastroenterologie, Hepatologie, Infektiologie und Endokrinologie, Medizinische Hochschule Hannover, Hannover, Deutschland
| | - Nadine Fischer
- Deutsche Gesellschaft für Gastroenterologie, Verdauungs- und Stoffwechselkrankheiten (DGVS), Berlin, Deutschland
| | | | - Jörg Petersen
- IFI Institut für Interdisziplinäre Medizin an der Asklepios Klinik St Georg, Hamburg, Deutschland
| | - Frank Tacke
- Medizinische Klinik m. S. Hepatologie und Gastroenterologie, Charité Universitätsmedizin Berlin, Berlin, Deutschland
| | - Markus Cornberg
- Klinik für Gastroenterologie, Hepatologie, Infektiologie und Endokrinologie, Medizinische Hochschule Hannover, Hannover, Deutschland
- Centre for individualised infection Medicine (CiiM), Hannover, Deutschland
- Deutsches Zentrum für Infektionsforschung (DZIF), partner-site Hannover-Braunschweig, Deutschland
- D-SOLVE Consortium, Horizon Europe Project, partner-site Medizinische Hochschule Hannover, Hannover, Deutschland
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12
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Sandmann L, Berg T, Deterding K, Fischer N, Hinrichsen H, Petersen J, Tacke F, Cornberg M. Antiviral Therapy of Chronic Hepatitis D Virus Infection - Addendum to the S3 Guideline "Prophylaxis, Diagnosis and Therapy of Hepatitis B Virus Infection" of the German Society for Gastroenterology, Digestive and Metabolic Diseases (DGVS). ZEITSCHRIFT FUR GASTROENTEROLOGIE 2023; 61:e715-e732. [PMID: 38081178 DOI: 10.1055/a-2181-3345] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [MESH Headings] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 12/18/2023]
Affiliation(s)
- Lisa Sandmann
- Department of Gastroenterology, Hepatology, Infectious Diseases and Endocrinology, Hannover Medical School, Hannover, Germany
- D-SOLVE Consortium, Horizon Europe Project, partner-site Hannover Medical School, Germany
| | - Thomas Berg
- Division of Hepatology, Department of Medicine II, Leipzig University Medical Center, Leipzig, Germany
| | - Katja Deterding
- Department of Gastroenterology, Hepatology, Infectious Diseases and Endocrinology, Hannover Medical School, Hannover, Germany
| | - Nadine Fischer
- German Society for Gastroenterology, Digestive and Metabolic Diseases (DGVS), Berlin, Germany
| | | | - Jörg Petersen
- IFI Institute for Interdisciplinary Medicine at Asklepios Klinik St Georg, Hamburg, Germany
| | - Frank Tacke
- Department of Hepatology and Gastroenterology, Charité - Universitätsmedizin Berlin, Berlin, Germany
| | - Markus Cornberg
- Department of Gastroenterology, Hepatology, Infectious Diseases and Endocrinology, Hannover Medical School, Hannover, Germany
- Centre for individualised infection Medicine (CiiM), Hannover, Germany
- German Center for Infection Research (DZIF), partner-site Hannover-Braunschweig, Germany
- D-SOLVE Consortium, Horizon Europe Project, partner-site Hannover Medical School, Germany
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13
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Zhuang AQ, Chen Y, Chen SM, Liu WC, Li Y, Zhang WJ, Wu YH. Current Status and Challenges in Anti-Hepatitis B Virus Agents Based on Inactivation/Inhibition or Elimination of Hepatitis B Virus Covalently Closed Circular DNA. Viruses 2023; 15:2315. [PMID: 38140556 PMCID: PMC10747957 DOI: 10.3390/v15122315] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/15/2023] [Revised: 11/21/2023] [Accepted: 11/22/2023] [Indexed: 12/24/2023] Open
Abstract
There has been over half a century since the discovery of hepatitis B virus (HBV) to now, but approximately 300 million patients with chronic hepatitis B (CHB) still live in the world, resulting in about one million deaths every year. Although currently approved antivirals (e.g., nucleoside analogues) are effective at reducing HBV replication, they have almost no impact on the existing HBV covalently closed circular DNA (cccDNA) reservoir. HBV cccDNA is a critical obstacle to the complete elimination of the virus via antiviral therapy. The true cure of HBV infection requires the eradication of viral cccDNA from HBV-infected cells; thus, the development of new agents directly or indirectly targeting HBV cccDNA is urgently needed due to the limitations of current available drugs against HBV infection. In this regard, it is the major focus of current anti-HBV research worldwide via different mechanisms to either inactivate/inhibit (functional cure) or eliminate (complete cure) HBV cccDNA. Therefore, this review discussed and summarized recent advances and challenges in efforts to inactivate/silence or eliminate viral cccDNA using anti-HBV agents from different sources, such as small molecules (including epigenetic drugs) and polypeptides/proteins, and siRNA or gene-editing approaches targeting/attenuating HBV cccDNA via different mechanisms, as well as future directions that may be considered in efforts to truly cure chronic HBV infection. In conclusion, no breakthrough has been made yet in attenuating HBV cccDNA, although a number of candidates have advanced into the phase of clinical trials. Furthermore, the overwhelming majority of the candidates function to indirectly target HBV cccDNA. No outstanding candidate directly targets HBV cccDNA. Relatively speaking, CCC_R08 and nitazoxanide may be some of the most promising agents to clear HBV infection in small molecule compounds. Additionally, CRISPR-Cas9 systems can directly target HBV cccDNA for decay and demonstrate significant anti-HBV activity. Consequently, gene-editing approaches targeting HBV cccDNA may be one of the most promising means to achieve the core goal of anti-HBV therapeutic strategies. In short, more basic studies on HBV infection need to be carried out to overcome these challenges.
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Affiliation(s)
| | | | | | | | | | | | - Yi-Hang Wu
- Zhejiang Provincial Key Laboratory of Biometrology and Inspection & Quarantine, Department of Pharmacy, College of Life Sciences, China Jiliang University, Hangzhou 310018, China
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14
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Mouzannar K, Liang TJ. Development of a highly potent anti-HBs monoclonal antibody for HBV and HDV therapy: An improvement with unsettled questions. J Hepatol 2023; 79:1079-1081. [PMID: 37586647 PMCID: PMC11653528 DOI: 10.1016/j.jhep.2023.08.002] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Grants] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 08/03/2023] [Accepted: 08/05/2023] [Indexed: 08/18/2023]
Affiliation(s)
- Karim Mouzannar
- Liver Diseases Branch, National Institute of Diabetics and Digestive and Kidney Diseases, NIH, Bethesda, Maryland, USA
| | - T Jake Liang
- Liver Diseases Branch, National Institute of Diabetics and Digestive and Kidney Diseases, NIH, Bethesda, Maryland, USA.
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15
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Olenginski LT, Attionu SK, Henninger EN, LeBlanc RM, Longhini AP, Dayie TK. Hepatitis B Virus Epsilon (ε) RNA Element: Dynamic Regulator of Viral Replication and Attractive Therapeutic Target. Viruses 2023; 15:1913. [PMID: 37766319 PMCID: PMC10534774 DOI: 10.3390/v15091913] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/01/2023] [Revised: 09/01/2023] [Accepted: 09/05/2023] [Indexed: 09/29/2023] Open
Abstract
Hepatitis B virus (HBV) chronically infects millions of people worldwide, which underscores the importance of discovering and designing novel anti-HBV therapeutics to complement current treatment strategies. An underexploited but attractive therapeutic target is ε, a cis-acting regulatory stem-loop RNA situated within the HBV pregenomic RNA (pgRNA). The binding of ε to the viral polymerase protein (P) is pivotal, as it triggers the packaging of pgRNA and P, as well as the reverse transcription of the viral genome. Consequently, small molecules capable of disrupting this interaction hold the potential to inhibit the early stages of HBV replication. The rational design of such ligands necessitates high-resolution structural information for the ε-P complex or its individual components. While these data are currently unavailable for P, our recent structural elucidation of ε through solution nuclear magnetic resonance spectroscopy marks a significant advancement in this area. In this review, we provide a brief overview of HBV replication and some of the therapeutic strategies to combat chronic HBV infection. These descriptions are intended to contextualize our recent experimental efforts to characterize ε and identify ε-targeting ligands, with the ultimate goal of developing novel anti-HBV therapeutics.
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Affiliation(s)
- Lukasz T. Olenginski
- Center for Biomolecular Structure and Organization, Department of Chemistry and Biochemistry, University of Maryland, College Park, MD 20742, USA (R.M.L.)
- Department of Biochemistry, University of Colorado, Boulder, CO 80309, USA
| | - Solomon K. Attionu
- Center for Biomolecular Structure and Organization, Department of Chemistry and Biochemistry, University of Maryland, College Park, MD 20742, USA (R.M.L.)
| | - Erica N. Henninger
- Center for Biomolecular Structure and Organization, Department of Chemistry and Biochemistry, University of Maryland, College Park, MD 20742, USA (R.M.L.)
| | - Regan M. LeBlanc
- Center for Biomolecular Structure and Organization, Department of Chemistry and Biochemistry, University of Maryland, College Park, MD 20742, USA (R.M.L.)
| | - Andrew P. Longhini
- Center for Biomolecular Structure and Organization, Department of Chemistry and Biochemistry, University of Maryland, College Park, MD 20742, USA (R.M.L.)
- Neuroscience Research Institute, University of California, Santa Barbara, Santa Barbara, CA 93106, USA
- Department of Molecular, Cellular and Developmental Biology, University of California, Santa Barbara, Santa Barbara, CA 93106, USA
| | - Theodore K. Dayie
- Center for Biomolecular Structure and Organization, Department of Chemistry and Biochemistry, University of Maryland, College Park, MD 20742, USA (R.M.L.)
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16
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Brunetto MR, Ricco G, Negro F, Wedemeyer H, Yurdaydin C, Asselah T, Papatheodoridis G, Gheorghe L, Agarwal K, Farci P, Buti M. EASL Clinical Practice Guidelines on hepatitis delta virus. J Hepatol 2023; 79:433-460. [PMID: 37364791 DOI: 10.1016/j.jhep.2023.05.001] [Citation(s) in RCA: 109] [Impact Index Per Article: 54.5] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 05/01/2023] [Accepted: 05/01/2023] [Indexed: 06/28/2023]
Abstract
Hepatitis D virus (HDV) is a defective virus that requires the hepatitis B virus to complete its life cycle and cause liver damage in humans. HDV is responsible for rare acute and chronic liver diseases and is considered the most aggressive hepatitis virus. Acute infection can cause acute liver failure, while persistent infection typically causes a severe form of chronic hepatitis which is associated with rapid and frequent progression to cirrhosis and its end-stage complications, hepatic decompensation and hepatocellular carcinoma. Major diagnostic and therapeutic innovations prompted the EASL Governing Board to commission specific Clinical Practice Guidelines on the identification, virologic and clinical characterisation, prognostic assessment, and appropriate clinical and therapeutic management of HDV-infected individuals.
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17
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Liu Y, Cafiero TR, Park D, Biswas A, Winer BY, Cho CH, Bram Y, Chandar V, Connell AKO, Gertje HP, Crossland N, Schwartz RE, Ploss A. Targeted viral adaptation generates a simian-tropic hepatitis B virus that infects marmoset cells. Nat Commun 2023; 14:3582. [PMID: 37328459 PMCID: PMC10276007 DOI: 10.1038/s41467-023-39148-3] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/19/2022] [Accepted: 05/26/2023] [Indexed: 06/18/2023] Open
Abstract
Hepatitis B virus (HBV) only infects humans and chimpanzees, posing major challenges for modeling HBV infection and chronic viral hepatitis. The major barrier in establishing HBV infection in non-human primates lies at incompatibilities between HBV and simian orthologues of the HBV receptor, sodium taurocholate co-transporting polypeptide (NTCP). Through mutagenesis analysis and screening among NTCP orthologues from Old World monkeys, New World monkeys and prosimians, we determined key residues responsible for viral binding and internalization, respectively and identified marmosets as a suitable candidate for HBV infection. Primary marmoset hepatocytes and induced pluripotent stem cell-derived hepatocyte-like cells support HBV and more efficient woolly monkey HBV (WMHBV) infection. Adapted chimeric HBV genome harboring residues 1-48 of WMHBV preS1 generated here led to a more efficient infection than wild-type HBV in primary and stem cell derived marmoset hepatocytes. Collectively, our data demonstrate that minimal targeted simianization of HBV can break the species barrier in small NHPs, paving the path for an HBV primate model.
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Affiliation(s)
- Yongzhen Liu
- Department of Molecular Biology, Princeton University, Princeton, NJ, 08544, USA
| | - Thomas R Cafiero
- Department of Molecular Biology, Princeton University, Princeton, NJ, 08544, USA
| | - Debby Park
- Department of Molecular Biology, Princeton University, Princeton, NJ, 08544, USA
| | - Abhishek Biswas
- Department of Molecular Biology, Princeton University, Princeton, NJ, 08544, USA
- Research Computing, Office of Information Technology, Princeton University, Princeton, NJ, 08544, USA
| | - Benjamin Y Winer
- Department of Molecular Biology, Princeton University, Princeton, NJ, 08544, USA
- Memorial Sloan-Kettering Cancer Center, 1275 York Avenue, New York, NY, 10065, USA
| | | | - Yaron Bram
- Division of Gastroenterology and Hepatology, Department of Medicine, Weill Cornell Medicine, New York, NY, 10065, USA
| | - Vasuretha Chandar
- Division of Gastroenterology and Hepatology, Department of Medicine, Weill Cornell Medicine, New York, NY, 10065, USA
| | - Aoife K O' Connell
- National Emerging Infectious Diseases Laboratories, Boston University, Boston, MA, 02118, USA
| | - Hans P Gertje
- National Emerging Infectious Diseases Laboratories, Boston University, Boston, MA, 02118, USA
| | - Nicholas Crossland
- National Emerging Infectious Diseases Laboratories, Boston University, Boston, MA, 02118, USA
- Department of Pathology and Laboratory Medicine, Boston University Chobanian & Avedisian School of Medicine, Boston, MA, 02118, USA
| | - Robert E Schwartz
- Division of Gastroenterology and Hepatology, Department of Medicine, Weill Cornell Medicine, New York, NY, 10065, USA
| | - Alexander Ploss
- Department of Molecular Biology, Princeton University, Princeton, NJ, 08544, USA.
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18
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Kostyushev D, Brezgin S, Kostyusheva A, Ponomareva N, Bayurova E, Zakirova N, Kondrashova A, Goptar I, Nikiforova A, Sudina A, Babin Y, Gordeychuk I, Lukashev A, Zamyatnin AA, Ivanov A, Chulanov V. Transient and tunable CRISPRa regulation of APOBEC/AID genes for targeting hepatitis B virus. MOLECULAR THERAPY. NUCLEIC ACIDS 2023; 32:478-493. [PMID: 37187708 PMCID: PMC10176074 DOI: 10.1016/j.omtn.2023.04.016] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Figures] [Subscribe] [Scholar Register] [Received: 10/25/2022] [Accepted: 04/17/2023] [Indexed: 05/17/2023]
Abstract
APOBEC/AID cytidine deaminases play an important role in innate immunity and antiviral defenses and were shown to suppress hepatitis B virus (HBV) replication by deaminating and destroying the major form of HBV genome, covalently closed circular DNA (cccDNA), without toxicity to the infected cells. However, developing anti-HBV therapeutics based on APOBEC/AID is complicated by the lack of tools for activating and controlling their expression. Here, we developed a CRISPR-activation-based approach (CRISPRa) to induce APOBEC/AID transient overexpression (>4-800,000-fold increase in mRNA levels). Using this new strategy, we were able to control APOBEC/AID expression and monitor their effects on HBV replication, mutation, and cellular toxicity. CRISPRa prominently reduced HBV replication (∼90%-99% decline of viral intermediates), deaminated and destroyed cccDNA, but induced mutagenesis in cancer-related genes. By coupling CRISPRa with attenuated sgRNA technology, we demonstrate that APOBEC/AID activation can be precisely controlled, eliminating off-site mutagenesis in virus-containing cells while preserving prominent antiviral activity. This study untangles the differences in the effects of physiologically expressed APOBEC/AID on HBV replication and cellular genome, provides insights into the molecular mechanisms of HBV cccDNA mutagenesis, repair, and degradation, and, finally, presents a strategy for a tunable control of APOBEC/AID expression and for suppressing HBV replication without toxicity.
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Affiliation(s)
- Dmitry Kostyushev
- Martsinovsky Institute of Medical Parasitology, Tropical and Vector-Borne Diseases, Sechenov University, 119991 Moscow, Russia
- Scientific Center for Genetics and Life Sciences, Division of Biotechnology, Sirius University of Science and Technology, 354340 Sochi, Russia
| | - Sergey Brezgin
- Martsinovsky Institute of Medical Parasitology, Tropical and Vector-Borne Diseases, Sechenov University, 119991 Moscow, Russia
- Scientific Center for Genetics and Life Sciences, Division of Biotechnology, Sirius University of Science and Technology, 354340 Sochi, Russia
| | - Anastasiya Kostyusheva
- Martsinovsky Institute of Medical Parasitology, Tropical and Vector-Borne Diseases, Sechenov University, 119991 Moscow, Russia
| | - Natalia Ponomareva
- Martsinovsky Institute of Medical Parasitology, Tropical and Vector-Borne Diseases, Sechenov University, 119991 Moscow, Russia
- Scientific Center for Genetics and Life Sciences, Division of Biotechnology, Sirius University of Science and Technology, 354340 Sochi, Russia
- Department of Pharmaceutical and Toxicological Chemistry, Sechenov First Moscow State Medical University, 119146 Moscow, Russia
| | - Ekaterina Bayurova
- Chumakov Federal Scientific Center for Research and Development of Immune and Biological Products of Russian Academy of Sciences, 108819 Moscow, Russia
| | - Natalia Zakirova
- Center for Precision Genome Editing and Genetic Technologies for Biomedicine, Engelhardt Institute of Molecular Biology, Russian Academy of Science, 119991 Moscow, Russia
| | - Alla Kondrashova
- Chumakov Federal Scientific Center for Research and Development of Immune and Biological Products of Russian Academy of Sciences, 108819 Moscow, Russia
| | - Irina Goptar
- Izmerov Research Institute of Occupational Health, 105275 Moscow, Russia
| | | | - Anna Sudina
- Federal State Budgetary Institution Centre for Strategic Planning and Management of Biomedical Health Risks of the Federal Medical Biological Agency, 119435 Moscow, Russia
| | - Yurii Babin
- Martsinovsky Institute of Medical Parasitology, Tropical and Vector-Borne Diseases, Sechenov University, 119991 Moscow, Russia
| | - Ilya Gordeychuk
- Chumakov Federal Scientific Center for Research and Development of Immune and Biological Products of Russian Academy of Sciences, 108819 Moscow, Russia
- Institute for Translational Medicine and Biotechnology, Sechenov First Moscow State Medical University, 127994 Moscow, Russia
- Department of Infectious Diseases, Sechenov First Moscow State Medical University, 119146 Moscow, Russia
| | - Alexander Lukashev
- Martsinovsky Institute of Medical Parasitology, Tropical and Vector-Borne Diseases, Sechenov University, 119991 Moscow, Russia
| | - Andrey A. Zamyatnin
- Scientific Center for Genetics and Life Sciences, Division of Biotechnology, Sirius University of Science and Technology, 354340 Sochi, Russia
- Institute of Molecular Medicine, Sechenov First Moscow State Medical University, 119991 Moscow, Russia
- Belozersky Institute of Physico-Chemical Biology, Lomonosov Moscow State University, 119992 Moscow, Russia
- Faculty of Health and Medical Sciences, University of Surrey, Guildford GU2 7X, UK
| | - Alexander Ivanov
- Center for Precision Genome Editing and Genetic Technologies for Biomedicine, Engelhardt Institute of Molecular Biology, Russian Academy of Science, 119991 Moscow, Russia
| | - Vladimir Chulanov
- Scientific Center for Genetics and Life Sciences, Division of Biotechnology, Sirius University of Science and Technology, 354340 Sochi, Russia
- Institute for Translational Medicine and Biotechnology, Sechenov First Moscow State Medical University, 127994 Moscow, Russia
- Department of Infectious Diseases, Sechenov First Moscow State Medical University, 119146 Moscow, Russia
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19
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Allweiss L, Testoni B, Yu M, Lucifora J, Ko C, Qu B, Lütgehetmann M, Guo H, Urban S, Fletcher SP, Protzer U, Levrero M, Zoulim F, Dandri M. Quantification of the hepatitis B virus cccDNA: evidence-based guidelines for monitoring the key obstacle of HBV cure. Gut 2023; 72:972-983. [PMID: 36707234 PMCID: PMC10086470 DOI: 10.1136/gutjnl-2022-328380] [Citation(s) in RCA: 13] [Impact Index Per Article: 6.5] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 07/29/2022] [Accepted: 01/15/2023] [Indexed: 01/29/2023]
Abstract
OBJECTIVES A major goal of curative hepatitis B virus (HBV) treatments is the reduction or inactivation of intrahepatic viral covalently closed circular DNA (cccDNA). Hence, precise cccDNA quantification is essential in preclinical and clinical studies. Southern blot (SB) permits cccDNA visualisation but lacks sensitivity and is very laborious. Quantitative PCR (qPCR) has no such limitations but inaccurate quantification due to codetection of viral replicative intermediates (RI) can occur. The use of different samples, preservation conditions, DNA extraction, nuclease digestion methods and qPCR strategies has hindered standardisation. Within the ICE-HBV consortium, available and novel protocols for cccDNA isolation and qPCR quantification in liver tissues and cell cultures were compared in six laboratories to develop evidence-based guidance for best practices. DESIGN Reference material (HBV-infected humanised mouse livers and HepG2-NTCP cells) was exchanged for cross-validation. Each group compared different DNA extraction methods (Hirt extraction, total DNA extraction with or without proteinase K treatment (+PK/-PK)) and nuclease digestion protocols (plasmid-safe ATP-dependent DNase (PSD), T5 exonuclease, exonucleases I/III). Samples were analysed by qPCR and SB. RESULTS Hirt and -PK extraction reduced coexisting RI forms. However, both cccDNA and the protein-free relaxed circular HBV DNA (pf-rcDNA) form were detected by qPCR. T5 and Exo I/III nucleases efficiently removed all RI forms. In contrast, PSD did not digest pf-rcDNA, but was less prone to induce cccDNA overdigestion. In stabilised tissues (eg, Allprotect), nucleases had detrimental effects on cccDNA. CONCLUSIONS We present here a comprehensive evidence-based guidance for optimising, controlling and validating cccDNA measurements using available qPCR assays.
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Affiliation(s)
- Lena Allweiss
- I. Medical Clinic and Polyclinic, University Medical Center Hamburg-Eppendorf, Hamburg, Germany
- German Center for Infection Research, Hamburg-Lübeck-Borstel-Riems, Munich and Heidelberg sites, Germany
| | - Barbara Testoni
- Cancer Research Center of Lyon, INSERM U1052, Lyon University, Hospices de Lyon, Lyon, France
- ANRS HBV Cure Task Force, Lyon, France
| | - Mei Yu
- Gilead Sciences, Foster City, California, USA
| | - Julie Lucifora
- Cancer Research Center of Lyon, INSERM U1052, Lyon University, Hospices de Lyon, Lyon, France
- ANRS HBV Cure Task Force, Lyon, France
- CIRI, Centre International de Recherche en Infectiologie, INSERM U1111, Université Claude Bernard Lyon 1, Lyon, France
| | - Chunkyu Ko
- Institute of Virology, Technical University of Munich, Munchen, Germany
- Infectious Diseases Therapeutic Research Center, Therapeutics & Biotechnology Division, Korea Research Institute of Chemical Technology, Daejeon, Korea (the Republic of)
| | - Bingqian Qu
- Department of Infectious Diseases, Molecular Virology, University Hospital Heidelberg, Heidelberg, Germany
- Division of Veterinary Medicine, Paul-Ehrlich-Institut, Langen, Germany
| | - Marc Lütgehetmann
- German Center for Infection Research, Hamburg-Lübeck-Borstel-Riems, Munich and Heidelberg sites, Germany
- Institute of Medical Microbiology, Virology and Hygiene, University Medical Center Hamburg-Eppendorf Hamburg, Hamburg, Germany
| | - Haitao Guo
- Indiana University School of Medicine, Indianapolis, Indiana, USA
- Department of Microbiology and Molecular Genetics, Cancer Virology Program, UPMC Hillman Cancer Center, University of Pittsburgh School of Medicine, Pittsburgh, Pennsylvania, USA
| | - Stephan Urban
- German Center for Infection Research, Hamburg-Lübeck-Borstel-Riems, Munich and Heidelberg sites, Germany
- Department of Infectious Diseases, Molecular Virology, University Hospital Heidelberg, Heidelberg, Germany
| | | | - Ulrike Protzer
- German Center for Infection Research, Hamburg-Lübeck-Borstel-Riems, Munich and Heidelberg sites, Germany
- Institute of Virology, Technical University of Munich, Munchen, Germany
| | - Massimo Levrero
- Cancer Research Center of Lyon, INSERM U1052, Lyon University, Hospices de Lyon, Lyon, France
- ANRS HBV Cure Task Force, Lyon, France
| | - Fabien Zoulim
- Cancer Research Center of Lyon, INSERM U1052, Lyon University, Hospices de Lyon, Lyon, France
- ANRS HBV Cure Task Force, Lyon, France
| | - Maura Dandri
- I. Medical Clinic and Polyclinic, University Medical Center Hamburg-Eppendorf, Hamburg, Germany
- German Center for Infection Research, Hamburg-Lübeck-Borstel-Riems, Munich and Heidelberg sites, Germany
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Burm R, Van Houtte F, Verhoye L, Mesalam AA, Ciesek S, Roingeard P, Wedemeyer H, Leroux-Roels G, Meuleman P. A human monoclonal antibody against HBsAg for the prevention and treatment of chronic HBV and HDV infection. JHEP Rep 2023; 5:100646. [PMID: 36748051 PMCID: PMC9898450 DOI: 10.1016/j.jhepr.2022.100646] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 10/07/2022] [Revised: 11/17/2022] [Accepted: 11/21/2022] [Indexed: 12/12/2022] Open
Abstract
Background & Aims Elimination of chronic HBV/HDV infection remains a major global health challenge. Targeting excessive hepatitis B surface antigen (HBsAg) release may provide an interesting window of opportunity to break immune tolerance and to achieve a functional cure using additional antivirals. Methods We evaluated a HBsAg-specific human monoclonal antibody, as part of either a prophylactic or therapeutic strategy, against HBV/HDV infection in cell culture models and in human-liver chimeric mice. To assess prophylactic efficacy, mice were passively immunized prior to infection with HBV or HBV/HDV (coinfection and superinfection setting). Therapeutic efficacy was assessed in HBV and HBV/HDV-coinfected mice receiving 4 weeks of treatment. Viral parameters (HBV DNA, HDV RNA and HBsAg) were assessed in mouse plasma. Results The antibody could effectively prevent HBV/HDV infection in a dose-dependent manner with IC50 values of ∼3.5 ng/ml. Passive immunization showed complete protection of mice from both HBV and HBV/HDV coinfection. Moreover, HDV superinfection was either completely prevented or at least attenuated in HBV-infected mice. Finally, antibody treatment in mice with established HBV/HDV infection resulted in a significant decline in viremia and a concomitant drop in on-treatment HBsAg, with a moderate viral rebound following treatment cessation. Conclusion We present data on a valuable antibody candidate that could complement other antivirals in strategies aimed at achieving functional cure of chronic HBV and HDV infection. Impact and implications Patients chronically infected with HBV may eventually develop liver cancer and are at great risk of being superinfected with HDV, which worsens and accelerates disease progression. Unfortunately, current treatments can rarely eliminate both viruses from chronically infected patients. In this study, we present data on a novel antibody that is able to prevent chronic HBV/HDV infection in a mouse model with a humanized liver. Moreover, antibody treatment of HBV/HDV-infected mice strongly diminishes viral loads during therapy. This antibody is a valuable candidate for further clinical development.
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Affiliation(s)
- Rani Burm
- Laboratory of Liver Infectious Diseases (LLID), Department of Diagnostic Sciences, Faculty of Medicine and Health Sciences, Ghent University, Ghent, Belgium
| | - Freya Van Houtte
- Laboratory of Liver Infectious Diseases (LLID), Department of Diagnostic Sciences, Faculty of Medicine and Health Sciences, Ghent University, Ghent, Belgium
| | - Lieven Verhoye
- Laboratory of Liver Infectious Diseases (LLID), Department of Diagnostic Sciences, Faculty of Medicine and Health Sciences, Ghent University, Ghent, Belgium
| | - Ahmed Atef Mesalam
- Laboratory of Liver Infectious Diseases (LLID), Department of Diagnostic Sciences, Faculty of Medicine and Health Sciences, Ghent University, Ghent, Belgium
- Department of Therapeutic Chemistry, Pharmaceutical and Drug Industries Research Institute, National Research Centre (NRC), Dokki, Cairo 12622, Egypt
| | - Sandra Ciesek
- Institute for Medical Virology, University Hospital, Goethe University, Frankfurt am Main, Germany
- German Center for Infection Research, DZIF, External Partner Site, Frankfurt am Main, Germany
- Fraunhofer Institute for Translational Medicine and Pharmacology (ITMP), Theodor Stern Kai 7, Frankfurt am Main, Germany
| | - Philippe Roingeard
- INSERM U966, Université François Rabelais and CHRU de Tours, Tours, France
| | - Heiner Wedemeyer
- Department of Gastroenterology, Hepatology and Endocrinology, Hannover Medical School, Hannover, Germany
| | - Geert Leroux-Roels
- Center for Vaccinology, Faculty of Medicine and Health Sciences, Ghent University and University Hospital, Ghent, Belgium
| | - Philip Meuleman
- Laboratory of Liver Infectious Diseases (LLID), Department of Diagnostic Sciences, Faculty of Medicine and Health Sciences, Ghent University, Ghent, Belgium
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Heat Shock Protein Family A Member 1 Promotes Intracellular Amplification of Hepatitis B Virus Covalently Closed Circular DNA. J Virol 2023; 97:e0126122. [PMID: 36519896 PMCID: PMC9888207 DOI: 10.1128/jvi.01261-22] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/23/2022] Open
Abstract
Hepatitis B virus (HBV) contains a partially double-stranded relaxed circular DNA (rcDNA) genome that is converted into a covalently closed circular DNA (cccDNA) in the nucleus of the infected hepatocyte by cellular DNA repair machinery. cccDNA associates with nucleosomes to form a minichromosome that transcribes RNA to support the expression of viral proteins and reverse transcriptional replication of viral DNA. In addition to the de novo synthesis from incoming virion rcDNA, cccDNA can also be synthesized from rcDNA in the progeny nucleocapsids within the cytoplasm of infected hepatocytes via the intracellular amplification pathway. In our efforts to identify cellular DNA repair proteins required for cccDNA synthesis using a chemogenetic screen, we found that B02, a small-molecule inhibitor of DNA homologous recombination repair protein RAD51, significantly enhanced the synthesis of cccDNA via the intracellular amplification pathway in human hepatoma cells. Ironically, neither small interfering RNA (siRNA) knockdown of RAD51 expression nor treatment with another structurally distinct RAD51 inhibitor or activator altered cccDNA amplification. Instead, it was found that B02 treatment significantly elevated the levels of multiple heat shock protein mRNA, and siRNA knockdown of HSPA1 expression or treatment with HSPA1 inhibitors significantly attenuated B02 enhancement of cccDNA amplification. Moreover, B02-enhanced cccDNA amplification was efficiently inhibited by compounds that selectively inhibit DNA polymerase α or topoisomerase II, the enzymes required for cccDNA intracellular amplification. Our results thus indicate that B02 treatment induces a heat shock protein-mediated cellular response that positively regulates the conversion of rcDNA into cccDNA via the authentic intracellular amplification pathway. IMPORTANCE Elimination or functional inactivation of cccDNA minichromosomes in HBV-infected hepatocytes is essential for the cure of chronic hepatitis B virus (HBV) infection. However, lack of knowledge of the molecular mechanisms of cccDNA metabolism and regulation hampers the development of antiviral drugs to achieve this therapeutic goal. Our findings reported here imply that enhanced cccDNA amplification may occur under selected pathobiological conditions, such as cellular stress, to subvert the dilution or elimination of cccDNA and maintain the persistence of HBV infection. Therapeutic inhibition of HSPA1-enhanced cccDNA amplification under these pathobiological conditions should facilitate the elimination of cccDNA and cure of chronic hepatitis B.
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Khalfi P, Kennedy PT, Majzoub K, Asselah T. Hepatitis D virus: Improving virological knowledge to develop new treatments. Antiviral Res 2023; 209:105461. [PMID: 36396025 DOI: 10.1016/j.antiviral.2022.105461] [Citation(s) in RCA: 13] [Impact Index Per Article: 6.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/04/2022] [Revised: 10/21/2022] [Accepted: 11/05/2022] [Indexed: 11/16/2022]
Abstract
Hepatitis delta virus (HDV), a satellite of hepatitis B virus (HBV), possesses the smallest viral genome known to infect animals. HDV needs HBV surface protein for secretion and entry into target liver cells. However, HBV is dispensable for HDV genome amplification, as it relies almost exclusively on cellular host factors for replication. HBV/HDV co-infections affect over 12 million people worldwide and constitute the most severe form of viral hepatitis. Co-infected individuals are at higher risk of developing liver cirrhosis and hepatocellular carcinoma compared to HBV mono-infected patients. Bulevirtide, an entry inhibitor, was conditionally approved in July 2020 in the European Union for adult patients with chronic hepatitis delta (CHD) and compensated liver disease. There are several drugs in development, including lonafarnib and interferon lambda, with different modes of action. In this review, we detail our current fundamental knowledge of HDV lifecycle and review antiviral treatments under development against this virus, outlining their respective mechanisms-of-action. Finally, we describe the antiviral effect these compounds are showing in ongoing clinical trials, discussing their promise and potential pitfalls for managing HDV infected patients.
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Affiliation(s)
- Pierre Khalfi
- Institut de Génétique Moléculaire de Montpellier, University of Montpellier, CNRS-UMR 5535, Montpellier 34293 cedex 5, France
| | - Patrick T Kennedy
- The Blizard Institute, Queen Mary University of London, The Royal London Hospital, Barts Health NHS Trust, London, UK
| | - Karim Majzoub
- Institut de Génétique Moléculaire de Montpellier, University of Montpellier, CNRS-UMR 5535, Montpellier 34293 cedex 5, France.
| | - Tarik Asselah
- Université de Paris, Cité CRI, INSERM UMR 1149, Department of Hepatology, AP-HP Hôpital Beaujon, Clichy, France.
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23
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Sivarajan R, Oberwinkler H, Roll V, König EM, Steinke M, Bodem J. A defined anthocyanin mixture sourced from bilberry and black currant inhibits Measles virus and various herpesviruses. BMC Complement Med Ther 2022; 22:181. [PMID: 35804339 PMCID: PMC9264518 DOI: 10.1186/s12906-022-03661-7] [Citation(s) in RCA: 5] [Impact Index Per Article: 1.7] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/18/2021] [Accepted: 06/29/2022] [Indexed: 11/10/2022] Open
Abstract
Abstract
Background
Anthocyanin-containing plant extracts and carotenoids, such as astaxanthin, have been well-known for their antiviral and anti-inflammatory activity, respectively. We hypothesised that a mixture of Ribes nigrum L. (Grossulariaceae) (common name black currant (BC)) and Vaccinium myrtillus L. (Ericaceae) (common name bilberry (BL)) extracts (BC/BL) with standardised anthocyanin content as well as single plant extracts interfered with the replication of Measles virus and Herpesviruses in vitro.
Methods
We treated cell cultures with BC/BL or defined single plant extracts, purified anthocyanins and astaxanthin in different concentrations and subsequently infected the cultures with the Measles virus (wild-type or vaccine strain Edmonston), Herpesvirus 1 or 8, or murine Cytomegalovirus. Then, we analysed the number of infected cells and viral infectivity and compared the data to non-treated controls.
Results
The BC/BL extract inhibited wild-type Measles virus replication, syncytia formation and cell-to-cell spread. This suppression was dependent on the wild-type virus-receptor-interaction since the Measles vaccine strain was unaffected by BC/BL treatment. Furthermore, the evidence was provided that the delphinidin-3-rutinoside chloride, a component of BC/BL, and purified astaxanthin, were effective anti-Measles virus compounds. Human Herpesvirus 1 and murine Cytomegalovirus replication was inhibited by BC/BL, single bilberry or black currant extracts, and the BC/BL component delphinidin-3-glucoside chloride. Additionally, we observed that BC/BL seemed to act synergistically with aciclovir. Moreover, BC/BL, the single bilberry and black currant extracts, and the BC/BL components delphinidin-3-glucoside chloride, cyanidin-3-glucoside, delphinidin-3-rutinoside chloride, and petunidin-3-galactoside inhibited human Herpesvirus 8 replication.
Conclusions
Our data indicate that Measles viruses and Herpesviruses are differentially susceptible to a specific BC/BL mixture, single plant extracts, purified anthocyanins and astaxanthin. These compounds might be used in the prevention of viral diseases and in addition to direct-acting antivirals, such as aciclovir.
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24
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Chen S, Zhang L, Chen Y, Fu L. Inhibiting Sodium Taurocholate Cotransporting Polypeptide in HBV-Related Diseases: From Biological Function to Therapeutic Potential. J Med Chem 2022; 65:12546-12561. [DOI: 10.1021/acs.jmedchem.2c01097] [Citation(s) in RCA: 10] [Impact Index Per Article: 3.3] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/29/2022]
Affiliation(s)
- Siwei Chen
- Sichuan Engineering Research Center for Biomimetic Synthesis of Natural Drugs, School of Life Science and Engineering, Southwest Jiaotong University, Chengdu 610031, China
| | - Lan Zhang
- Sichuan Engineering Research Center for Biomimetic Synthesis of Natural Drugs, School of Life Science and Engineering, Southwest Jiaotong University, Chengdu 610031, China
| | - Yi Chen
- Department of Gastrointestinal Surgery, West China Hospital, Sichuan University, Chengdu 610041, China
| | - Leilei Fu
- Sichuan Engineering Research Center for Biomimetic Synthesis of Natural Drugs, School of Life Science and Engineering, Southwest Jiaotong University, Chengdu 610031, China
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Suresh M, Menne S. Recent Drug Development in the Woodchuck Model of Chronic Hepatitis B. Viruses 2022; 14:v14081711. [PMID: 36016334 PMCID: PMC9416195 DOI: 10.3390/v14081711] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/17/2022] [Revised: 07/22/2022] [Accepted: 07/31/2022] [Indexed: 11/24/2022] Open
Abstract
Infection with hepatitis B virus (HBV) is responsible for the increasing global hepatitis burden, with an estimated 296 million people being carriers and living with the risk of developing chronic liver disease and cancer. While the current treatment options for chronic hepatitis B (CHB), including oral nucleos(t)ide analogs and systemic interferon-alpha, are deemed suboptimal, the path to finding an ultimate cure for this viral disease is rather challenging. The lack of suitable laboratory animal models that support HBV infection and associated liver disease progression is one of the major hurdles in antiviral drug development. For more than four decades, experimental infection of the Eastern woodchuck with woodchuck hepatitis virus has been applied for studying the immunopathogenesis of HBV and developing new antiviral therapeutics against CHB. There are several advantages to this animal model that are beneficial for performing both basic and translational HBV research. Previous review articles have focused on the value of this animal model in regard to HBV replication, pathogenesis, and immune response. In this article, we review studies of drug development and preclinical evaluation of direct-acting antivirals, immunomodulators, therapeutic vaccines, and inhibitors of viral entry, gene expression, and antigen release in the woodchuck model of CHB since 2014 until today and discuss their significance for clinical trials in patients.
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26
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Atorvastatin Rapidly Reduces Hepatitis B Viral Load in Combination with Tenofovir: A Prospective Clinical Trial. CANADIAN JOURNAL OF INFECTIOUS DISEASES AND MEDICAL MICROBIOLOGY 2022; 2022:3443813. [PMID: 35873362 PMCID: PMC9303483 DOI: 10.1155/2022/3443813] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Subscribe] [Scholar Register] [Received: 02/09/2022] [Revised: 06/14/2022] [Accepted: 06/24/2022] [Indexed: 11/18/2022]
Abstract
Objective and Aim. Atorvastatin inhibits cholesterol synthesis which is critically important in the formation of the viral envelope and secretion. The efficacy and safety of giving atorvastatin (40 mg/day) as an adjunct to tenofovir in the treatment of hepatitis B (HBV) were assessed. Method. In this single-blind clinical trial, 40 patients with active chronic hepatitis B were randomly allocated to treatment or control groups. The treatment group received the standard treatment for chronic HBV (300 mg tenofovir twice a day) along with 40 mg/day atorvastatin for 12 months, while the control group received a placebo once daily in addition to the standard tenofovir regimen. Serum aspartate aminotransferase (AST), alanine aminotransferase (ALT), and HBV DNA copy numbers were measured at the beginning of the treatment and 1, 3, 6, 9, 12 months later. Results. One month after starting the treatment, the HBV copy number in the atorvastatin + tenofovir-treated group was significantly lower, by 200×, compared with the control group. After three months of the treatment, there was no detectable HBV DNA in 50% of the atorvastatin + tenofovir-treated group compared with 30% in the control group. The half-life of plasma viral load was 2.03 and 3.32 months in the atorvastatin + tenofovir-treated and control groups, respectively. No adverse events due to taking atorvastatin were observed. Conclusions. The combination of atorvastatin with tenofovir increased antiviral activity and led to a faster recovery from viral infection. Therefore, this modality can be recommended as a safe combination therapy for chronic hepatitis B patients.
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27
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Huang Y, Liu N, Ning Q, Zhou M, Zang N, Liang T, Wei W. Design, synthesis, and biological evaluation of novel (E)-1-arylethan-1-one O-((3-arylisoxazol-5-yl) methyl) oxime derivatives as potent non-nucleoside HBV inhibitors. J Mol Struct 2022. [DOI: 10.1016/j.molstruc.2022.132789] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/29/2022]
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28
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Boettler T, Gill US, Allweiss L, Pollicino T, Tavis JE, Zoulim F. Assessing immunological and virological responses in the liver: Implications for the cure of chronic hepatitis B virus infection. JHEP Rep 2022; 4:100480. [PMID: 35493765 PMCID: PMC9039841 DOI: 10.1016/j.jhepr.2022.100480] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 12/28/2021] [Revised: 03/08/2022] [Accepted: 03/09/2022] [Indexed: 11/11/2022] Open
Abstract
Cure from chronic HBV infection is rare with current therapies. Basic research has helped to fundamentally improve our knowledge of the viral life cycle and virus-host interactions, and provided the basis for several novel drug classes that are currently being developed or are being tested in clinical trials. While these novel compounds targeting the viral life cycle or antiviral immune responses hold great promise, we are still lacking a comprehensive understanding of the immunological and virological processes that occur at the site of infection, the liver. At the International Liver Congress 2021 (ILC 2021), a research think tank on chronic HBV infection focused on mechanisms within the liver that facilitate persistent infection and looked at the research questions that need to be addressed to fill knowledge gaps and identify novel therapeutic strategies. Herein, we summarise the discussion by the think tank and identify the key basic research questions that must be addressed in order to develop more effective strategies for the functional cure of HBV infection.
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Affiliation(s)
- Tobias Boettler
- Department of Medicine II (Gastroenterology, Hepatology, Endocrinology and Infectious Diseases), Freiburg University Medical Center, Faculty of Medicine, University of Freiburg, Freiburg, Germany
| | - Upkar S. Gill
- Blizard Institute, Centre for Immunobiology, Barts and the London School of Medicine and Dentistry, Queen Mary University of London, London, UK
| | - Lena Allweiss
- I. Medical Clinic and Polyclinic, University Medical Center Hamburg-Eppendorf, Hamburg, Germany
- German Center for Infection Research (DZIF), Hamburg-Lübeck-Borstel-Riems sites, Germany
| | - Teresa Pollicino
- Department of Human Pathology, University Hospital "G. Martino" of Messina, Messina, Italy
| | - John E. Tavis
- Department of Molecular Microbiology and Immunology and Institute for Drug and Biotherapeutic Innovation, Saint Louis University, Saint Louis MO USA
| | - Fabien Zoulim
- INSERM Unit 1052 – Cancer Research Center of Lyon, Department of Hepatology Hospices Civils de Lyon, Lyon University, France
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29
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Mitosis of Hepatitis B virus-infected cells in vitro results in uninfected daughter cells. JHEP Rep 2022; 4:100514. [PMID: 35898957 PMCID: PMC9309680 DOI: 10.1016/j.jhepr.2022.100514] [Citation(s) in RCA: 6] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 04/13/2022] [Accepted: 05/16/2022] [Indexed: 11/24/2022] Open
Abstract
Background & Aims The chronicity of HBV (and resultant liver disease) is determined by intrahepatic persistence of the HBV covalently closed circular DNA (cccDNA), an episomal form that encodes all viral transcripts. Therefore, cccDNA is a key target for new treatments, with the ultimate therapeutic aim being its complete elimination. Although established cccDNA molecules are known to be stable in resting hepatocytes, we aimed to understand their fate in dividing cells using in vitro models. Methods We infected HepG2-NTCP and HepaRG-NTCP cells with HBV and induced mitosis by passaging cells. We measured cccDNA copy number (by precise PCR assays) and HBV-expressing cells (by immunofluorescence) with wild-type HBV. We used reporter viruses expressing luciferase or RFP to track number of HBV-expressing cells over time after mitosis induction using luciferase assays and live imaging, respectively. Results In all cases, we observed dramatic reductions in cccDNA levels, HBV-positive cell numbers, and cccDNA-dependent protein expression after each round of cell mitosis. The rates of reduction were highly consistent with mathematical models of a complete cccDNA loss in (as opposed to dilution into) daughter cells. Conclusions Our results are concordant with previous animal models of HBV infection and show that HBV persistence can be efficiently overcome by inducing cell mitosis. These results support therapeutic approaches that induce liver turnover (e.g. immune modulators) in addition to direct-acting antiviral therapies to achieve hepatitis B cure. Lay summary Chronic hepatitis B affects 300 million people (killing 884,000 per year) and is incurable. To cure it, we need to clear the HBV genome from the liver. In this study, we looked at how the virus behaves after a cell divides. We found that it completely clears the virus, making 2 new uninfected cells. Our work informs new approaches to develop cures for chronic hepatitis B infections.
HBV persists over decades in the liver, leading to chronic inflammation and serious liver disease. Controversy exists over the fate of viral DNA after cell mitosis, which is crucial to understanding viral persistence. We find here that 2 completely uninfected daughter cells are generated when infected cells undergo mitosis. Our results suggest that therapies that induce turnover of infected cells could facilitate the clearance of chronic HBV infection.
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30
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Park JH, Iwamoto M, Yun JH, Uchikubo-Kamo T, Son D, Jin Z, Yoshida H, Ohki M, Ishimoto N, Mizutani K, Oshima M, Muramatsu M, Wakita T, Shirouzu M, Liu K, Uemura T, Nomura N, Iwata S, Watashi K, Tame JRH, Nishizawa T, Lee W, Park SY. Structural insights into the HBV receptor and bile acid transporter NTCP. Nature 2022; 606:1027-1031. [PMID: 35580630 PMCID: PMC9242859 DOI: 10.1038/s41586-022-04857-0] [Citation(s) in RCA: 62] [Impact Index Per Article: 20.7] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/29/2021] [Accepted: 05/11/2022] [Indexed: 01/05/2023]
Abstract
Around 250 million people are infected with hepatitis B virus (HBV) worldwide1, and 15 million may also carry the satellite virus hepatitis D virus (HDV), which confers even greater risk of severe liver disease2. The HBV receptor has been identified as sodium taurocholate co-transporting polypeptide (NTCP), which interacts directly with the first 48 amino acid residues of the N-myristoylated N-terminal preS1 domain of the viral large protein3. Despite the pressing need for therapeutic agents to counter HBV, the structure of NTCP remains unsolved. This 349-residue protein is closely related to human apical sodium-dependent bile acid transporter (ASBT), another member of the solute carrier family SLC10. Crystal structures have been reported of similar bile acid transporters from bacteria4,5, and these models are believed to resemble closely both NTCP and ASBT. Here we have used cryo-electron microscopy to solve the structure of NTCP bound to an antibody, clearly showing that the transporter has no equivalent of the first transmembrane helix found in other SLC10 proteins, and that the N terminus is exposed on the extracellular face. Comparison of our structure with those of related proteins indicates a common mechanism of bile acid transport, but the NTCP structure displays an additional pocket formed by residues that are known to interact with preS1, presenting new opportunities for structure-based drug design. Cryo-electron structures of the hepatitis B virus receptor NTCP show a distinct membrane topology compared with other SLC10 proteins, but a common bile acid transport mechanism that is shared with related mammalian and bacterial proteins.
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Affiliation(s)
- Jae-Hyun Park
- Drug Design Laboratory, Graduate School of Medical Life Science, Yokohama City University, Yokohama, Japan
| | - Masashi Iwamoto
- Department of Virology II, National Institute of Infectious Diseases, Tokyo, Japan
| | - Ji-Hye Yun
- Department of Biochemistry, College of Life Science and Biotechnology, Yonsei University, Seoul, South Korea.,PCG-Biotech, Seoul, South Korea
| | - Tomomi Uchikubo-Kamo
- Laboratory for Protein Functional and Structural Biology, RIKEN Center for Biosystems Dynamics Research, Yokohama, Japan
| | - Donghwan Son
- Department of Biochemistry, College of Life Science and Biotechnology, Yonsei University, Seoul, South Korea
| | - Zeyu Jin
- Drug Design Laboratory, Graduate School of Medical Life Science, Yokohama City University, Yokohama, Japan.,Department of Biochemistry, College of Life Science and Biotechnology, Yonsei University, Seoul, South Korea
| | - Hisashi Yoshida
- Drug Design Laboratory, Graduate School of Medical Life Science, Yokohama City University, Yokohama, Japan
| | - Mio Ohki
- Drug Design Laboratory, Graduate School of Medical Life Science, Yokohama City University, Yokohama, Japan
| | - Naito Ishimoto
- Drug Design Laboratory, Graduate School of Medical Life Science, Yokohama City University, Yokohama, Japan
| | - Kenji Mizutani
- Drug Design Laboratory, Graduate School of Medical Life Science, Yokohama City University, Yokohama, Japan
| | - Mizuki Oshima
- Department of Virology II, National Institute of Infectious Diseases, Tokyo, Japan.,Department of Biological Sciences, Tokyo University of Science, Noda, Japan
| | - Masamichi Muramatsu
- Department of Virology II, National Institute of Infectious Diseases, Tokyo, Japan
| | - Takaji Wakita
- Department of Virology II, National Institute of Infectious Diseases, Tokyo, Japan
| | - Mikako Shirouzu
- Laboratory for Protein Functional and Structural Biology, RIKEN Center for Biosystems Dynamics Research, Yokohama, Japan
| | - Kehong Liu
- Department of Cell Biology, Graduate School of Medicine, Kyoto University, Kyoto, Japan
| | - Tomoko Uemura
- Department of Cell Biology, Graduate School of Medicine, Kyoto University, Kyoto, Japan
| | - Norimichi Nomura
- Department of Cell Biology, Graduate School of Medicine, Kyoto University, Kyoto, Japan
| | - So Iwata
- Department of Cell Biology, Graduate School of Medicine, Kyoto University, Kyoto, Japan.,RIKEN SPring-8 Center, Sayo-gun, Japan
| | - Koichi Watashi
- Department of Virology II, National Institute of Infectious Diseases, Tokyo, Japan.,Department of Biological Sciences, Tokyo University of Science, Noda, Japan.,Research Center for Drug and Vaccine Development, Tokyo, Japan
| | - Jeremy R H Tame
- Drug Design Laboratory, Graduate School of Medical Life Science, Yokohama City University, Yokohama, Japan
| | - Tomohiro Nishizawa
- Laboratory of Biomembrane Dynamics, Graduate School of Medical Life Science, Yokohama City University, Yokohama, Japan
| | - Weontae Lee
- Department of Biochemistry, College of Life Science and Biotechnology, Yonsei University, Seoul, South Korea. .,PCG-Biotech, Seoul, South Korea.
| | - Sam-Yong Park
- Drug Design Laboratory, Graduate School of Medical Life Science, Yokohama City University, Yokohama, Japan.
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Li Y, Zhou J, Li T. Regulation of the HBV Entry Receptor NTCP and its Potential in Hepatitis B Treatment. Front Mol Biosci 2022; 9:879817. [PMID: 35495620 PMCID: PMC9039015 DOI: 10.3389/fmolb.2022.879817] [Citation(s) in RCA: 11] [Impact Index Per Article: 3.7] [Reference Citation Analysis] [Abstract] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/20/2022] [Accepted: 03/14/2022] [Indexed: 11/13/2022] Open
Abstract
Hepatitis B virus (HBV) is a globally prevalent human DNA virus responsible for more than 250 million cases of chronic liver infection, a condition that can lead to liver inflammation, cirrhosis, and hepatocellular carcinoma. Sodium taurocholate co-transporting polypeptide (NTCP), a transmembrane protein highly expressed in human hepatocytes and a mediator of bile acid transport, has been identified as the receptor responsible for the cellular entry of both HBV and its satellite, hepatitis delta virus (HDV). This has led to significant advances in our understanding of the HBV life cycle, especially the early steps of infection. HepG2-NTCP cells and human NTCP-expressing transgenic mice have been employed as the primary cell culture and animal models, respectively, for the study of HBV, and represent valuable approaches for investigating its basic biology and developing treatments for infection. However, the mechanisms involved in the regulation of NTCP transcription, translation, post-translational modification, and transport are still largely elusive. Improvements in our understanding of NTCP biology would likely facilitate the design of new therapeutic drugs for the prevention of the de novo infection of naïve hepatocytes. In this review, we provide critical findings regarding NTCP biology and discuss important questions that remain unanswered.
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Affiliation(s)
- Yan Li
- *Correspondence: Yan Li, ; Tianliang Li,
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Colombatto P, Coco B, Bonino F, Brunetto MR. Management and Treatment of Patients with Chronic Hepatitis B: Towards Personalized Medicine. Viruses 2022; 14:701. [PMID: 35458431 PMCID: PMC9027850 DOI: 10.3390/v14040701] [Citation(s) in RCA: 8] [Impact Index Per Article: 2.7] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/13/2022] [Revised: 03/22/2022] [Accepted: 03/23/2022] [Indexed: 02/07/2023] Open
Abstract
The currently available antiviral treatments (Peg-Interferon-α and Nucleos(t)ide Analogues, NA) for chronic hepatitis B (CHB) achieve a functional cure (serum HBsAg and HDV-DNA clearance) of HBV infection in a limited number of patients. Nevertheless, the continuous pharmacological suppression of viral replication by NA halts liver disease progression lowering the risk of HCC development and improving the survival. In the near future, to fully exploit the potential of old and new drugs for HBV treatment a personalized approach to the patients will be required according to an accurate definition of their virologic, immunologic and clinical profile.
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Affiliation(s)
- Piero Colombatto
- Hepatology Unit and Laboratory of Molecular Genetics and Pathology of Hepatitis Viruses, Reference Center of the Tuscany Region for Chronic Liver Disease and Cancer, Department of Medical Specialties, University Hospital of Pisa, Via Paradisa 2, 56124 Pisa, Italy; (P.C.); (B.C.)
| | - Barbara Coco
- Hepatology Unit and Laboratory of Molecular Genetics and Pathology of Hepatitis Viruses, Reference Center of the Tuscany Region for Chronic Liver Disease and Cancer, Department of Medical Specialties, University Hospital of Pisa, Via Paradisa 2, 56124 Pisa, Italy; (P.C.); (B.C.)
| | - Ferruccio Bonino
- Institute of Biostructure and Bioimaging, National Research Council, Via De Amicis 95, 80145 Naples, Italy;
| | - Maurizia R. Brunetto
- Hepatology Unit and Laboratory of Molecular Genetics and Pathology of Hepatitis Viruses, Reference Center of the Tuscany Region for Chronic Liver Disease and Cancer, Department of Medical Specialties, University Hospital of Pisa, Via Paradisa 2, 56124 Pisa, Italy; (P.C.); (B.C.)
- Institute of Biostructure and Bioimaging, National Research Council, Via De Amicis 95, 80145 Naples, Italy;
- Internal Medicine, Department of Clinical and Experimental Medicine, University of Pisa, Via Savi 10, 56127 Pisa, Italy
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33
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Gad SA, Sugiyama M, Tsuge M, Wakae K, Fukano K, Oshima M, Sureau C, Watanabe N, Kato T, Murayama A, Li Y, Shoji I, Shimotohno K, Chayama K, Muramatsu M, Wakita T, Nozaki T, Aly HH. The kinesin KIF4 mediates HBV/HDV entry through the regulation of surface NTCP localization and can be targeted by RXR agonists in vitro. PLoS Pathog 2022; 18:e1009983. [PMID: 35312737 PMCID: PMC8970526 DOI: 10.1371/journal.ppat.1009983] [Citation(s) in RCA: 4] [Impact Index Per Article: 1.3] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/25/2021] [Revised: 03/31/2022] [Accepted: 03/04/2022] [Indexed: 11/30/2022] Open
Abstract
Intracellular transport via microtubule-based dynein and kinesin family motors plays a key role in viral reproduction and transmission. We show here that Kinesin Family Member 4 (KIF4) plays an important role in HBV/HDV infection. We intended to explore host factors impacting the HBV life cycle that can be therapeutically addressed using siRNA library transfection and HBV/NLuc (HBV/NL) reporter virus infection in HepG2-hNTCP cells. KIF4 silencing resulted in a 3-fold reduction in luciferase activity following HBV/NL infection. KIF4 knockdown suppressed both HBV and HDV infection. Transient KIF4 depletion reduced surface and raised intracellular NTCP (HBV/HDV entry receptor) levels, according to both cellular fractionation and immunofluorescence analysis (IF). Overexpression of wild-type KIF4 but not ATPase-null KIF4 mutant regained the surface localization of NTCP and significantly restored HBV permissiveness in these cells. IF revealed KIF4 and NTCP colocalization across microtubule filaments, and a co-immunoprecipitation study revealed that KIF4 interacts with NTCP. KIF4 expression is regulated by FOXM1. Interestingly, we discovered that RXR agonists (Bexarotene, and Alitretinoin) down-regulated KIF4 expression via FOXM1-mediated suppression, resulting in a substantial decrease in HBV-Pre-S1 protein attachment to HepG2-hNTCP cell surface and subsequent HBV infection in both HepG2-hNTCP and primary human hepatocyte (PXB) (Bexarotene, IC50 1.89 ± 0.98 μM) cultures. Overall, our findings show that human KIF4 is a critical regulator of NTCP surface transport and localization, which is required for NTCP to function as a receptor for HBV/HDV entry. Furthermore, small molecules that suppress or alleviate KIF4 expression would be potential antiviral candidates targeting HBV and HDV entry. Understanding HBV/HDV entry machinery and the mechanism by which NTCP (HBV/HDV entry receptor) surface expression is regulated is crucial to develop antiviral entry inhibitors. We found that NTCP surface transport is mainly controlled by the motor kinesin KIF4. Surprisingly, KIF4 was negatively regulated by RXR receptors through FOXM1-mediated suppression. This study not only mechanistically correlated the role of RXR receptors in regulating HBV/HDV entry but also suggested a novel approach to develop therapeutic rexinoids for preventing HBV and/or HDV infections in important clinical situations, such as in patients undergoing liver transplantation or those who are at a high risk of HBV infection and unresponsive to HBV vaccination.
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Affiliation(s)
- Sameh A. Gad
- Department of Virology II, National Institute of Infectious Diseases, Tokyo, Japan
- Department of Biomedical Chemistry, Graduate School of Medicine, The University of Tokyo, Tokyo, Japan
- Department of Microbiology and Immunology, Faculty of Pharmacy, Minia University, Minia, Egypt
| | - Masaya Sugiyama
- Genome Medical Sciences Project, National Center for Global Health and Medicine, Chiba, Japan
| | - Masataka Tsuge
- Natural Science Center for Basic Research and Development, Hiroshima University, Hiroshima, Japan
| | - Kosho Wakae
- Department of Virology II, National Institute of Infectious Diseases, Tokyo, Japan
| | - Kento Fukano
- Department of Virology II, National Institute of Infectious Diseases, Tokyo, Japan
| | - Mizuki Oshima
- Department of Virology II, National Institute of Infectious Diseases, Tokyo, Japan
- Graduate School of Science and Technology, Tokyo University of Science, Noda, Japan
| | - Camille Sureau
- Institut National de la Transfusion Sanguine, Paris, France
| | - Noriyuki Watanabe
- Department of Virology II, National Institute of Infectious Diseases, Tokyo, Japan
| | - Takanobu Kato
- Department of Virology II, National Institute of Infectious Diseases, Tokyo, Japan
| | - Asako Murayama
- Department of Virology II, National Institute of Infectious Diseases, Tokyo, Japan
| | - Yingfang Li
- Department of Virology II, National Institute of Infectious Diseases, Tokyo, Japan
| | - Ikuo Shoji
- Center for Infectious Diseases, Kobe University Graduate School of Medicine, Kobe, Japan
| | - Kunitada Shimotohno
- Center for Hepatitis and Immunology, National Center for Global Health and Medicine, Chiba, Japan
| | - Kazuaki Chayama
- Collaborative Research Laboratory of Medical Innovation, Graduate School of Biomedical and Health Sciences, Hiroshima University, Hiroshima, Japan
- Research Center for Hepatology and Gastroenterology, Graduate School of Biomedical and Health Sciences, Hiroshima University, Hiroshima, Japan
- RIKEN Center for Integrative Medical Sciences, Yokohama, Japan
| | - Masamichi Muramatsu
- Department of Virology II, National Institute of Infectious Diseases, Tokyo, Japan
| | - Takaji Wakita
- Department of Virology II, National Institute of Infectious Diseases, Tokyo, Japan
- * E-mail: (TW); (HHA)
| | - Tomoyoshi Nozaki
- Department of Biomedical Chemistry, Graduate School of Medicine, The University of Tokyo, Tokyo, Japan
| | - Hussein H. Aly
- Department of Virology II, National Institute of Infectious Diseases, Tokyo, Japan
- * E-mail: (TW); (HHA)
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Michelet M, Alfaiate D, Chardès B, Pons C, Faure-Dupuy S, Engleitner T, Farhat R, Riedl T, Legrand AF, Rad R, Rivoire M, Zoulim F, Heikenwälder M, Salvetti A, Durantel D, Lucifora J. Inducers of the NF-κB pathways impair hepatitis delta virus replication and strongly decrease progeny infectivity in vitro. JHEP Rep 2022; 4:100415. [PMID: 35141510 PMCID: PMC8792426 DOI: 10.1016/j.jhepr.2021.100415] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 07/07/2021] [Revised: 11/03/2021] [Accepted: 12/02/2021] [Indexed: 10/26/2022] Open
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Kamiya N, Sugimoto T, Abe-Chayama H, Akiyama R, Tsuboi Y, Mogami A, Imamura M, Hayes CN, Chayama K. Untying relaxed circular DNA of hepatitis B virus by polymerase reaction provides a new option for accurate quantification and visualization of covalently closed circular DNA. J Gen Virol 2022; 103. [PMID: 35130138 DOI: 10.1099/jgv.0.001591] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/18/2022] Open
Abstract
Hepatitis B virus (HBV) is a small hepatotropic DNA virus that replicates via an RNA intermediate. After entry, the virus capsid carries relaxed circular DNA (rcDNA) into the nucleus where the viral genome is converted into covalently closed circular DNA (cccDNA), which serves as the template for all viral transcripts. To monitor cccDNA levels, preprocessing methods to eliminate rcDNA have emerged for quantitative PCR, although Southern blotting is still the only method to discriminate cccDNA from other DNA intermediates. In this study, we have established a robust method for untying mature rcDNA into double stranded linear DNA using specific polymerases. Untying rcDNA provides not only an alternative method for cccDNA quantification but also a sensitive method for visualizing cccDNA. We combined this method with plasmid-safe DNase and T5 exonuclease preprocessing and revealed that accurate quantification requires cccDNA digestion by a restriction enzyme because heat stability of cccDNA increases after T5 exonuclease treatment. In digital PCR using duplex TaqMan probes, fewer than 1000 copies of cccDNA were successfully visualized as double positive spots that were distinct from single positives derived from untied rcDNA. This method was further applied to the infection model of primary hepatocytes treated with nucleoside analogues and a core protein allosteric modulator to monitor cccDNA levels. Relative quantification of cccDNA by human genome copy demonstrated the possibility of precise evaluation of cccDNA level per nucleus. These results clearly indicate that the sequential reaction from untying rcDNA is useful to investigate cccDNA fates in a small fraction of nuclei.
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Affiliation(s)
- Naohiro Kamiya
- Research Unit/Immunology & Inflammation, Sohyaku, Innovative Research Division, Mitsubishi Tanabe Pharma Corporation, Yokohama, Kanagawa, Japan.,Department of Gastroenterology and Metabolism, Graduate School of Biomedical and Health Sciences, Hiroshima University, Hiroshima, Japan
| | - Takahiko Sugimoto
- Research Unit/Immunology & Inflammation, Sohyaku, Innovative Research Division, Mitsubishi Tanabe Pharma Corporation, Yokohama, Kanagawa, Japan
| | - Hiromi Abe-Chayama
- Department of Gastroenterology and Metabolism, Graduate School of Biomedical and Health Sciences, Hiroshima University, Hiroshima, Japan.,Research Center for Hepatology and Gastroenterology, Hiroshima University, Hiroshima, Japan
| | - Rie Akiyama
- Department of Gastroenterology and Metabolism, Graduate School of Biomedical and Health Sciences, Hiroshima University, Hiroshima, Japan.,Research Center for Hepatology and Gastroenterology, Hiroshima University, Hiroshima, Japan
| | - Yasunori Tsuboi
- Research Unit/Immunology & Inflammation, Sohyaku, Innovative Research Division, Mitsubishi Tanabe Pharma Corporation, Yokohama, Kanagawa, Japan
| | - Akira Mogami
- Research Unit/Immunology & Inflammation, Sohyaku, Innovative Research Division, Mitsubishi Tanabe Pharma Corporation, Yokohama, Kanagawa, Japan
| | - Michio Imamura
- Department of Gastroenterology and Metabolism, Graduate School of Biomedical and Health Sciences, Hiroshima University, Hiroshima, Japan.,Research Center for Hepatology and Gastroenterology, Hiroshima University, Hiroshima, Japan
| | - C Nelson Hayes
- Department of Gastroenterology and Metabolism, Graduate School of Biomedical and Health Sciences, Hiroshima University, Hiroshima, Japan.,Research Center for Hepatology and Gastroenterology, Hiroshima University, Hiroshima, Japan
| | - Kazuaki Chayama
- Department of Gastroenterology and Metabolism, Graduate School of Biomedical and Health Sciences, Hiroshima University, Hiroshima, Japan.,Research Center for Hepatology and Gastroenterology, Hiroshima University, Hiroshima, Japan.,Institute of Physical and Chemical Research (RIKEN) Center for Integrative Medical Sciences, Yokohama, Japan
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36
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Kang B, Yi DY, Choe BH. Translational Strategies to Eliminate Chronic Hepatitis B in Children: Prophylaxis and Management in East Asian Countries. Front Pediatr 2022; 9:809838. [PMID: 35186829 PMCID: PMC8854863 DOI: 10.3389/fped.2021.809838] [Citation(s) in RCA: 3] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 11/05/2021] [Accepted: 12/27/2021] [Indexed: 12/14/2022] Open
Abstract
Translational medical research on hepatitis B virus (HBV) infection and chronic hepatitis B (CHB) pathogenesis provides guidance on strengthening the treatment and prevention strategies of CHB. Preventing vertical transmission is the key to eliminating HBV infection in children. The understanding of HBV replication, hepatocyte turnover, and the fate of covalently closed circular DNA (cccDNA) would help establish a personalized application of the guidelines, especially concerning the discontinuation of nucleos(t)ide analog (NA) treatment in children. Transplacental leakage of HBV-infected maternal blood is suggested as the leading cause of vertical transmission. Prenatal maternal prophylaxis could diminish maternal HBV viremia at delivery, to reduce the risk of neonatal HBV infection. The meaning of the expression "no additional risk of breast milk feeding" is thereby explained. Understanding the untreated natural course of CHB in children and the course changeable by treatment is important to apply individualistic strategies and avoid the immoral selection of treatment indications. NAs with potent efficacy and a high barrier to drug resistance should be used as first-line treatment to reduce the likelihood of NA-resistant HBV development because the rate of mutant HBV emergence might count on the infected hepatocyte turnover rate in chronic HBV infection. Although elimination of intranuclear cccDNA is difficult by NAs alone, a cure is possible by human immunity and hepatocyte turnover. The reduction of intranuclear cccDNA occurs after the destruction of HBV-infected hepatocytes, non-cytolytic immune response, apoptosis of hepatocytes, and compensatory cell proliferation. Therefore, consolidation therapy after NA-induced hepatitis B e-antigen seroconversion must be necessary for a sufficient period. This review also summarizes the treatment strategies of CHB in children based on the practical application of translational research.
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Affiliation(s)
- Ben Kang
- Department of Pediatrics, School of Medicine, Kyungpook National University, Daegu, South Korea
| | - Dae Yong Yi
- Department of Pediatrics, College of Medicine, Chung-Ang University Hospital, Chung-Ang University, Seoul, South Korea
| | - Byung-Ho Choe
- Department of Pediatrics, School of Medicine, Kyungpook National University, Daegu, South Korea
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Allweiss L, Giersch K, Pirosu A, Volz T, Muench RC, Beran RK, Urban S, Javanbakht H, Fletcher SP, Lütgehetmann M, Dandri M. Therapeutic shutdown of HBV transcripts promotes reappearance of the SMC5/6 complex and silencing of the viral genome in vivo. Gut 2022; 71:372-381. [PMID: 33509930 PMCID: PMC8762019 DOI: 10.1136/gutjnl-2020-322571] [Citation(s) in RCA: 55] [Impact Index Per Article: 18.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 07/23/2020] [Revised: 01/12/2021] [Accepted: 01/14/2021] [Indexed: 12/13/2022]
Abstract
OBJECTIVE Therapeutic strategies silencing and reducing the hepatitis B virus (HBV) reservoir, the covalently closed circular DNA (cccDNA), have the potential to cure chronic HBV infection. We aimed to investigate the impact of small interferring RNA (siRNA) targeting all HBV transcripts or pegylated interferon-α (peg-IFNα) on the viral regulatory HBx protein and the structural maintenance of chromosome 5/6 complex (SMC5/6), a host factor suppressing cccDNA transcription. In particular, we assessed whether interventions lowering HBV transcripts can achieve and maintain silencing of cccDNA transcription in vivo. DESIGN HBV-infected human liver chimeric mice were treated with siRNA or peg-IFNα. Virological and host changes were analysed at the end of treatment and during the rebound phase by qualitative PCR, ELISA, immunoblotting and chromatin immunoprecipitation. RNA in situ hybridisation was combined with immunofluorescence to detect SMC6 and HBV RNAs at single cell level. The entry inhibitor myrcludex-B was used during the rebound phase to avoid new infection events. RESULTS Both siRNA and peg-IFNα strongly reduced all HBV markers, including HBx levels, thus enabling the reappearance of SMC5/6 in hepatocytes that achieved HBV-RNA negativisation and SMC5/6 association with the cccDNA. Only IFN reduced cccDNA loads and enhanced IFN-stimulated genes. However, the antiviral effects did not persist off treatment and SMC5/6 was again degraded. Remarkably, the blockade of viral entry that started at the end of treatment hindered renewed degradation of SMC5/6. CONCLUSION These results reveal that therapeutics abrogating all HBV transcripts including HBx promote epigenetic suppression of the HBV minichromosome, whereas strategies protecting the human hepatocytes from reinfection are needed to maintain cccDNA silencing.
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Affiliation(s)
- Lena Allweiss
- Department of Internal Medicine, University Medical Center Hamburg-Eppendorf, Hamburg, Germany
| | - Katja Giersch
- Department of Internal Medicine, University Medical Center Hamburg-Eppendorf, Hamburg, Germany
| | - Andrea Pirosu
- Department of Internal Medicine, University Medical Center Hamburg-Eppendorf, Hamburg, Germany
| | - Tassilo Volz
- Department of Internal Medicine, University Medical Center Hamburg-Eppendorf, Hamburg, Germany,German Center for Infection Research (DZIF), Hamburg-Lübeck-Borstel-Riems and Heidelberg sites, Germany
| | | | | | - Stephan Urban
- German Center for Infection Research (DZIF), Hamburg-Lübeck-Borstel-Riems and Heidelberg sites, Germany,Department of Infectious Diseases, Molecular Virology, University Hospital Heidelberg, Heidelberg, Germany
| | | | | | - Marc Lütgehetmann
- German Center for Infection Research (DZIF), Hamburg-Lübeck-Borstel-Riems and Heidelberg sites, Germany,Department of Medical Microbiology, Virology and Hygiene, University Medical Center Hamburg-Eppendorf, Hamburg, Germany
| | - Maura Dandri
- Department of Internal Medicine, University Medical Center Hamburg-Eppendorf, Hamburg, Germany .,German Center for Infection Research (DZIF), Hamburg-Lübeck-Borstel-Riems and Heidelberg sites, Germany
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38
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Nosaka T, Naito T, Murata Y, Matsuda H, Ohtani M, Hiramatsu K, Nishizawa T, Okamoto H, Nakamoto Y. Regulatory function of interferon-inducible 44-like for hepatitis B virus covalently closed circular DNA in primary human hepatocytes. Hepatol Res 2022; 52:141-152. [PMID: 34697871 DOI: 10.1111/hepr.13722] [Citation(s) in RCA: 5] [Impact Index Per Article: 1.7] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 05/24/2021] [Revised: 10/13/2021] [Accepted: 10/17/2021] [Indexed: 12/12/2022]
Abstract
AIM Curing hepatitis B virus (HBV) infection requires elimination of covalently closed circular DNA (cccDNA). Interferon (IFN)-γ has noncytolytic antiviral potential; however, elimination of cccDNA could not be achieved. To enhance the regulatory effect, we comprehensively analyzed the host factors associated with cccDNA amplification and IFN-γ and IFN-α effects using an in vitro HBV infection system showing various transcription levels. METHODS Primary human hepatocytes were infected with HBV using genomic plasmids carrying the basic core promoter mutation A1762T/G1764A and/or the precore mutation G1896A and treated with IFN-γ and IFN-α. Comprehensive and functional studies involving microarray and small interfering RNA analysis revealed the host factors related to cccDNA regulation. RESULTS The HBV infection system reproduced the HBV life cycle and showed various propagation levels. Microarray analysis revealed 53 genes correlated with the cccDNA levels. Of the 53 genes, expression of IFN-induced protein 44-like (IFI44L) was significantly upregulated by IFN-γ and IFN-α. The anti-HBV effect of IFI44L is exerted regardless of IFN-γ or IFN-α by inhibiting the activation of nuclear factor-κB and signal transducer and activator of transcription 1 pathways. CONCLUSIONS Using the in vitro HBV infection system, an IFN-inducible molecule, IFI44L, associated with cccDNA amplification, was identified. These results suggest an innovative molecular strategy for the regulation of HBV cccDNA by controlling a novel host factor, IFI44L.
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Affiliation(s)
- Takuto Nosaka
- Second Department of Internal Medicine, Faculty of Medical Sciences, University of Fukui, Fukui, Japan
| | - Tatsushi Naito
- Second Department of Internal Medicine, Faculty of Medical Sciences, University of Fukui, Fukui, Japan
| | - Yosuke Murata
- Second Department of Internal Medicine, Faculty of Medical Sciences, University of Fukui, Fukui, Japan
| | - Hidetaka Matsuda
- Second Department of Internal Medicine, Faculty of Medical Sciences, University of Fukui, Fukui, Japan
| | - Masahiro Ohtani
- Second Department of Internal Medicine, Faculty of Medical Sciences, University of Fukui, Fukui, Japan
| | - Katsushi Hiramatsu
- Second Department of Internal Medicine, Faculty of Medical Sciences, University of Fukui, Fukui, Japan
| | - Tsutomu Nishizawa
- Division of Virology, Department of Infection and Immunity, Jichi Medical University School of Medicine, Tochigi, Japan
| | - Hiroaki Okamoto
- Division of Virology, Department of Infection and Immunity, Jichi Medical University School of Medicine, Tochigi, Japan
| | - Yasunari Nakamoto
- Second Department of Internal Medicine, Faculty of Medical Sciences, University of Fukui, Fukui, Japan
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Tricot T, Thibaut HJ, Abbasi K, Boon R, Helsen N, Kumar M, Neyts J, Verfaillie C. Metabolically Improved Stem Cell Derived Hepatocyte-Like Cells Support HBV Life Cycle and Are a Promising Tool for HBV Studies and Antiviral Drug Screenings. Biomedicines 2022; 10:biomedicines10020268. [PMID: 35203482 PMCID: PMC8869365 DOI: 10.3390/biomedicines10020268] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.7] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/23/2021] [Revised: 01/21/2022] [Accepted: 01/23/2022] [Indexed: 11/16/2022] Open
Abstract
More than 300 million people worldwide are diagnosed with a chronic hepatitis B virus (HBV) infection. Nucleos(t)ide viral polymerase inhibitors are available on the market and can efficiently treat patients with chronic HBV. However, life-long treatment is needed as covalently closed circular DNA (cccDNA) persists in the hepatocyte nucleus. Hence, there is a high demand for novel therapeutics that can eliminate cccDNA from the hepatocyte nucleus and cure chronically infected HBV patients. The gold standard for in vitro HBV studies is primary human hepatocytes (PHHs). However, alternatives are needed due to donor organ shortage and high batch-to-batch variability. Therefore, human pluripotent stem cell (hPSC)-derived hepatocyte-like cells (HLCs) are being explored as an in vitro HBV infection model. We recently generated hPSC lines that overexpress three transcription factors (HC3x) and that, upon differentiation in a high amino-acid supplemented maturation medium, generate a more mature hepatocyte progeny (HC3x-AA-HLCs). Here, we demonstrate that HBV can efficiently infect these HC3x-AA-HLCs, as was shown by the presence of HBV core (HBc) and surface antigens. A clear increasing release of HBV surface and e antigens was detected, indicating the formation of functional cccDNA. Moreover, back-titration of culture supernatant of HBV-infected HC3x-AA-HLCs on HepG2-NTCP cells revealed the production of novel infectious HBV particles. Additionally, an increasing number of HBc-positive HC3x-AA-HLCs over time suggests viral spreading is occurring. Finally, the HC3x-AA-HLC model was validated for use in antiviral drug studies using the nucleoside reverse-transcriptase inhibitor, lamivudine, and the HBV entry inhibitor, Myrcludex B.
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Affiliation(s)
- Tine Tricot
- Stem Cell Institute, Rega Institute KU Leuven, 3000 Leuven, Belgium; (R.B.); (N.H.); (M.K.)
- Correspondence: (T.T.); (H.J.T.); (C.V.); Tel.: +32-16-37-71-09 (T.T.); +32-16-32-16-82 (H.J.T.); +32-16-37-26-54 (C.V.)
| | - Hendrik Jan Thibaut
- Department of Microbiology, Immunology and Transplantation, Virology and Chemotherapy, Rega Institute KU Leuven, 3000 Leuven, Belgium; (K.A.); (J.N.)
- Department of Microbiology, Immunology and Transplantation, Translational Platform Virology and Chemotherapy (TPVC), Rega Institute KU Leuven, 3000 Leuven, Belgium
- Correspondence: (T.T.); (H.J.T.); (C.V.); Tel.: +32-16-37-71-09 (T.T.); +32-16-32-16-82 (H.J.T.); +32-16-37-26-54 (C.V.)
| | - Kayvan Abbasi
- Department of Microbiology, Immunology and Transplantation, Virology and Chemotherapy, Rega Institute KU Leuven, 3000 Leuven, Belgium; (K.A.); (J.N.)
| | - Ruben Boon
- Stem Cell Institute, Rega Institute KU Leuven, 3000 Leuven, Belgium; (R.B.); (N.H.); (M.K.)
- Laboratory for Functional Epigenetics, Department of Human Genetics, Rega Institute KU Leuven, 3000 Leuven, Belgium
| | - Nicky Helsen
- Stem Cell Institute, Rega Institute KU Leuven, 3000 Leuven, Belgium; (R.B.); (N.H.); (M.K.)
- Ismar Healthcare NV, 2500 Lier, Belgium
| | - Manoj Kumar
- Stem Cell Institute, Rega Institute KU Leuven, 3000 Leuven, Belgium; (R.B.); (N.H.); (M.K.)
| | - Johan Neyts
- Department of Microbiology, Immunology and Transplantation, Virology and Chemotherapy, Rega Institute KU Leuven, 3000 Leuven, Belgium; (K.A.); (J.N.)
| | - Catherine Verfaillie
- Stem Cell Institute, Rega Institute KU Leuven, 3000 Leuven, Belgium; (R.B.); (N.H.); (M.K.)
- Correspondence: (T.T.); (H.J.T.); (C.V.); Tel.: +32-16-37-71-09 (T.T.); +32-16-32-16-82 (H.J.T.); +32-16-37-26-54 (C.V.)
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Leowattana W, Leowattana T. Chronic hepatitis B: New potential therapeutic drugs target. World J Virol 2022; 11:57-72. [PMID: 35117971 PMCID: PMC8788212 DOI: 10.5501/wjv.v11.i1.57] [Citation(s) in RCA: 7] [Impact Index Per Article: 2.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 05/31/2021] [Revised: 08/13/2021] [Accepted: 01/05/2022] [Indexed: 02/06/2023] Open
Abstract
Chronic hepatitis B (CHB) infection remains the most causative agent of liver-related morbidity and mortality worldwide. It impacts nearly 300 million people. The current treatment for chronic infection with the hepatitis B virus (HBV) is complex and lacks a durable treatment response, especially hepatitis B surface antigen (HBsAg) loss, necessitating indefinite treatment in most CHB patients due to the persistence of HBV covalently closed circular DNA (cccDNA). New drugs that target distinct steps of the HBV life cycle have been investigated, which comprise inhibiting the entry of HBV into hepatocytes, disrupting or silencing HBV cccDNA, modulating nucleocapsid assembly, interfering HBV transcription, and inhibiting HBsAg release. The achievement of a functional cure or sustained HBsAg loss in CHB patients represents the following approach towards HBV eradication. This review will explore the up-to-date advances in the development of new direct-acting anti-HBV drugs. Hopefully, with the combination of the current antiviral drugs and the newly developed direct-acting antiviral drugs targeting the different steps of the HBV life cycle, the ultimate eradication of CHB infection will soon be achieved.
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Affiliation(s)
- Wattana Leowattana
- Clinical Tropical Medicine, Faculty of Tropical Medicine, Mahidol University, Bangkok 10400, Thailand
| | - Tawithep Leowattana
- Department of Medicine, Faculty of Medicine, Srinakharinwirot University, Bangkok 10110, Thailand
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Sintusek P, Thanapirom K, Komolmit P, Poovorawan Y. Eliminating viral hepatitis in children after liver transplants: How to reach the goal by 2030. World J Gastroenterol 2022; 28:290-309. [PMID: 35110951 PMCID: PMC8771616 DOI: 10.3748/wjg.v28.i3.290] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 06/03/2021] [Revised: 08/12/2021] [Accepted: 01/06/2022] [Indexed: 02/06/2023] Open
Abstract
Viral hepatitis infections are a great burden in children who have received liver transplant. Hepatotropic viruses can cause liver inflammation that can develop into liver graft fibrosis and cirrhosis over the long term. Immunological reactions due to viral hepatitis infections are associated with or can mimic graft rejection, rendering the condition difficult to manage. Prevention strategies using vaccinations are agreeable to patients, safe, cost-effective and practical. Hence, strategies to eliminate viral hepatitis A and B focus mainly on immunization programmes for children who have received a liver transplant. Although a vaccine has been developed to prevent hepatitis C and E viruses, its use is not licensed worldwide. Consequently, eliminating hepatitis C and E viruses mainly involves early detection in children with suspected cases and effective treatment with antiviral therapy. Good hygiene and sanitation are also important to prevent hepatitis A and E infections. Donor blood products and liver grafts should be screened for hepatitis B, C and E in children who are undergoing liver transplantation. Future research on early detection of viral hepatitis infections should include molecular techniques for detecting hepatitis B and E. Moreover, novel antiviral drugs for eradicating viral hepatitis that are highly effective and safe are needed for children who have undergone liver transplantation.
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Affiliation(s)
- Palittiya Sintusek
- The Thai Pediatric Gastroenterology, Hepatology and Immunology (TPGHAI) Research Unit, Chulalongkorn University, Bangkok 10330, Thailand
- Division of Gastroenterology, Department of Pediatrics, Faculty of Medicine, King Chulalongkorn Memorial Hospital, Chulalongkorn University, Bangkok 10330, Thailand
| | - Kessarin Thanapirom
- Division of Gastroenterology, Department of Medicine, Liver Fibrosis and Cirrhosis Research Unit, Faculty of Medicine, Chulalongkorn University and King Chulalongkorn Memorial Hospital, Thai Red Cross Society, Bangkok 10330, Thailand
- Center of Excellence in Liver Diseases, King Chulalongkorn Memorial Hospital, Chulalongkorn University, Bangkok 10330, Thailand
| | - Piyawat Komolmit
- Division of Gastroenterology, Department of Medicine, Liver Fibrosis and Cirrhosis Research Unit, Faculty of Medicine, Chulalongkorn University and King Chulalongkorn Memorial Hospital, Thai Red Cross Society, Bangkok 10330, Thailand
- Center of Excellence in Liver Diseases, King Chulalongkorn Memorial Hospital, Chulalongkorn University, Bangkok 10330, Thailand
| | - Yong Poovorawan
- Center of Excellence in Clinical Virology, Faculty of Medicine, Chulalongkorn University, Bangkok 10330, Thailand
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Düzgüneş N, Fernandez-Fuentes N, Konopka K. Inhibition of Viral Membrane Fusion by Peptides and Approaches to Peptide Design. Pathogens 2021; 10:1599. [PMID: 34959554 PMCID: PMC8709411 DOI: 10.3390/pathogens10121599] [Citation(s) in RCA: 11] [Impact Index Per Article: 2.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/23/2021] [Revised: 12/06/2021] [Accepted: 12/06/2021] [Indexed: 12/29/2022] Open
Abstract
Fusion of lipid-enveloped viruses with the cellular plasma membrane or the endosome membrane is mediated by viral envelope proteins that undergo large conformational changes following binding to receptors. The HIV-1 fusion protein gp41 undergoes a transition into a "six-helix bundle" after binding of the surface protein gp120 to the CD4 receptor and a co-receptor. Synthetic peptides that mimic part of this structure interfere with the formation of the helix structure and inhibit membrane fusion. This approach also works with the S spike protein of SARS-CoV-2. Here we review the peptide inhibitors of membrane fusion involved in infection by influenza virus, HIV-1, MERS and SARS coronaviruses, hepatitis viruses, paramyxoviruses, flaviviruses, herpesviruses and filoviruses. We also describe recent computational methods used for the identification of peptide sequences that can interact strongly with protein interfaces, with special emphasis on SARS-CoV-2, using the PePI-Covid19 database.
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Affiliation(s)
- Nejat Düzgüneş
- Department of Biomedical Sciences, Arthur A. Dugoni School of Dentistry, University of the Pacific, San Francisco, CA 94103, USA;
| | - Narcis Fernandez-Fuentes
- Institute of Biological, Environmental and Rural Sciences, Aberystwyth University, Aberystwyth SY23 3EE, UK;
| | - Krystyna Konopka
- Department of Biomedical Sciences, Arthur A. Dugoni School of Dentistry, University of the Pacific, San Francisco, CA 94103, USA;
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Efficient Inhibition of Hepatitis B Virus (HBV) Replication and cccDNA Formation by HBV Ribonuclease H Inhibitors during Infection. Antimicrob Agents Chemother 2021; 65:e0146021. [PMID: 34516242 DOI: 10.1128/aac.01460-21] [Citation(s) in RCA: 10] [Impact Index Per Article: 2.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/12/2022] Open
Abstract
The hepatitis B virus (HBV) ribonuclease H (RNase H) is an attractive but unexploited drug target. Here, we addressed three limitations to the current state of RNase H inhibitor development: (a) Efficacy has been assessed only in transfected cell lines. (b) Cytotoxicity data are from transformed cell lines rather than primary cells. (c) It is unknown how the compounds work against nucleos(t)ide analog resistant HBV strains. Three RNase H inhibitors from different chemotypes, 110 (α-hydroxytropolone), 1133 (N-hydroxypyridinedione), and 1073 (N-hydroxynapthyridinone), were tested in HBV-infected HepG2-NTCP cells for inhibition of cccDNA accumulation and HBV product formation. 50% effective concentrations (EC50s) were 0.049-0.078 μM in the infection studies compared to 0.29-1.6 μM in transfected cells. All compounds suppressed cccDNA formation by >98% at 5 μM when added shortly after infection. HBV RNA, intracellular and extracellular DNA, and HBsAg secretion were all robustly suppressed. The greater efficacy of the inhibitors when added shortly after infection is presumably due to blocking amplification of the HBV cccDNA, which suppresses events downstream of cccDNA formation. The compounds had 50% cytotoxic concentrations (CC50s) of 16-100 μM in HepG2-derived cell lines but were nontoxic in primary human hepatocytes, possibly due to the quiescent state of the hepatocytes. The compounds had similar EC50s against replication of wild-type, lamivudine-resistant, and adefovir/lamivudine-resistant HBV, as expected because the RNase H inhibitors do not target the viral reverse transcriptase active site. These studies expand confidence in inhibiting the HBV RNase H as a drug strategy and support inclusion of RNase H inhibitors in novel curative drug combinations for HBV.
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Korolowicz KE, Suresh M, Li B, Huang X, Yon C, Kallakury BV, Lee KP, Park S, Kim YW, Menne S. Combination Treatment with the Vimentin-Targeting Antibody hzVSF and Tenofovir Suppresses Woodchuck Hepatitis Virus Infection in Woodchucks. Cells 2021; 10:2321. [PMID: 34571970 PMCID: PMC8466705 DOI: 10.3390/cells10092321] [Citation(s) in RCA: 5] [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: 07/16/2021] [Revised: 08/24/2021] [Accepted: 08/27/2021] [Indexed: 02/07/2023] Open
Abstract
Current treatment options for patients infected with hepatitis B virus (HBV) are suboptimal, because the approved drugs rarely induce cure due to the persistence of the viral DNA genome in the nucleus of infected hepatocytes, and are associated with either severe side effects (pegylated interferon-alpha) or require life-long administration (nucleos(t)ide analogs). We report here the evaluation of the safety and therapeutic efficacy of a novel, humanized antibody (hzVSF) in the woodchuck model of HBV infection. hzVSF has been shown to act as a viral entry inhibitor, most likely by suppressing vimentin-mediated endocytosis of virions. Targeting the increased vimentin expression on liver cells by hzVSF after infection with HBV or woodchuck hepatitis virus (WHV) was demonstrated initially. Thereafter, hzVSF safety was assessed in eight woodchucks naïve for WHV infection. Antiviral efficacy of hzVSF was evaluated subsequently in 24 chronic WHV carrier woodchucks by monotreatment with three ascending doses and in combination with tenofovir alafenamide fumarate (TAF). Consistent with the proposed blocking of WHV reinfection, intravenous hzVSF administration for 12 weeks resulted in a modest but transient reduction of viral replication and associated liver inflammation. In combination with oral TAF dosing, the antiviral effect of hzVSF was enhanced and sustained in half of the woodchucks with an antibody response to viral proteins. Thus, hzVSF safely but modestly alters chronic WHV infection in woodchucks; however, as a combination partner to TAF, its antiviral efficacy is markedly increased. The results of this preclinical study support future evaluation of this novel anti-HBV drug in patients.
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Affiliation(s)
- Kyle E. Korolowicz
- Department of Microbiology & Immunology, Georgetown University Medical Center, Washington, DC 20057, USA; (K.E.K.); (M.S.); (B.L.); (X.H.); (C.Y.)
| | - Manasa Suresh
- Department of Microbiology & Immunology, Georgetown University Medical Center, Washington, DC 20057, USA; (K.E.K.); (M.S.); (B.L.); (X.H.); (C.Y.)
| | - Bin Li
- Department of Microbiology & Immunology, Georgetown University Medical Center, Washington, DC 20057, USA; (K.E.K.); (M.S.); (B.L.); (X.H.); (C.Y.)
| | - Xu Huang
- Department of Microbiology & Immunology, Georgetown University Medical Center, Washington, DC 20057, USA; (K.E.K.); (M.S.); (B.L.); (X.H.); (C.Y.)
| | - Changsuek Yon
- Department of Microbiology & Immunology, Georgetown University Medical Center, Washington, DC 20057, USA; (K.E.K.); (M.S.); (B.L.); (X.H.); (C.Y.)
| | - Bhaskar V. Kallakury
- Department of Pathology, Georgetown University Medical Center, Washington, DC 20057, USA;
| | - Kyoung-pil Lee
- ImmuneMed, Inc., Chuncheon BioTown, Soyanggang ro 32, Chuncheon-si 24232, Gangwon-do, Korea; (K.-p.L.); (S.P.); (Y.-W.K.)
| | - Sungman Park
- ImmuneMed, Inc., Chuncheon BioTown, Soyanggang ro 32, Chuncheon-si 24232, Gangwon-do, Korea; (K.-p.L.); (S.P.); (Y.-W.K.)
| | - Yoon-Won Kim
- ImmuneMed, Inc., Chuncheon BioTown, Soyanggang ro 32, Chuncheon-si 24232, Gangwon-do, Korea; (K.-p.L.); (S.P.); (Y.-W.K.)
| | - Stephan Menne
- Department of Microbiology & Immunology, Georgetown University Medical Center, Washington, DC 20057, USA; (K.E.K.); (M.S.); (B.L.); (X.H.); (C.Y.)
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Thangamani L, Balasubramanian B, Easwaran M, Natarajan J, Pushparaj K, Meyyazhagan A, Piramanayagam S. GalNAc-siRNA conjugates: Prospective tools on the frontier of anti-viral therapeutics. Pharmacol Res 2021; 173:105864. [PMID: 34474100 PMCID: PMC8405237 DOI: 10.1016/j.phrs.2021.105864] [Citation(s) in RCA: 10] [Impact Index Per Article: 2.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 08/03/2021] [Revised: 08/27/2021] [Accepted: 08/27/2021] [Indexed: 12/19/2022]
Abstract
The growing use of short-interfering RNA (siRNA)-based therapeutics for viral diseases reflects the most recent innovations in anti-viral vaccines and drugs. These drugs play crucial roles in the fight against many hitherto incurable diseases, the causes, pathophysiologies, and molecular processes of which remain unknown. Targeted liver drug delivery systems are in clinical trials. The receptor-mediated endocytosis approach involving the abundant asialoglycoprotein receptors (ASGPRs) on the surfaces of liver cells show great promise. We here review N-acetylgalactosamine (GalNAc)-siRNA conjugates that treat viral diseases such as hepatitis B infection, but we also mention that novel, native conjugate-based, targeted siRNA anti-viral drugs may also cure several life-threatening diseases such as hemorrhagic cystitis, multifocal leukoencephalopathy, and severe acute respiratory syndrome caused by coronaviruses and human herpes virus.
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Affiliation(s)
- Lokesh Thangamani
- Computational Biology Lab, Department of Bioinformatics, Bharathiar University, Coimbatore 641046, Tamil Nadu, India
| | | | - Murugesh Easwaran
- International Centre for Genetic Engineering and Biotechnology, New Delhi 110067, India
| | - Jeyakumar Natarajan
- Data Mining and Text Mining Laboratory, Department of Bioinformatics, Bharathiar University, Coimbatore 641046, Tamil Nadu, India
| | - Karthika Pushparaj
- Department of Zoology, School of Biosciences, Avinashilingam Institute for Home Science and Higher Education for Women, Coimbatore 641043, Tamil Nadu, India
| | - Arun Meyyazhagan
- Department of Life Sciences, CHRIST (Deemed to be University), Bengaluru 560029, Karnataka, India.
| | - Shanmughavel Piramanayagam
- Computational Biology Lab, Department of Bioinformatics, Bharathiar University, Coimbatore 641046, Tamil Nadu, India.
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Naresh P, Pottabatula SS, Selvaraj J. Dengue virus entry/fusion inhibition by small bioactive molecules; A critical review. Mini Rev Med Chem 2021; 22:484-497. [PMID: 34353253 DOI: 10.2174/1389557521666210805105146] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/02/2020] [Revised: 02/14/2021] [Accepted: 04/19/2021] [Indexed: 11/22/2022]
Abstract
Many flaviviruses are remarkable human pathogens that can be transmitted by mosquitoes and ticks. Despite the availability of vaccines for viral infections such as yellow fever, Japanese encephalitis, and tick-borne encephalitis, flavivirus-like dengue is still a significant life-threatening illness worldwide. To date, there is no antiviral treatment for dengue therapy. Industry and the research community have been taking ongoing steps to improve anti-flavivirus treatment to meet this clinical need. The successful activity has been involved in the inhibition of the virus entry fusion process in the last two decades. In this study, the latest understanding of the use of small molecules used as fusion inhibitors has been comprehensively presented. We summarized the structure, the process of fusion of dengue virus E protein (DENV E), and the amino acids involved in the fusion process. Special attention has been given to small molecules that allow conformational changes to DENV E protein viz. blocking the pocket of βOG, which is important for fusion.
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Affiliation(s)
- Podila Naresh
- Department of Pharmaceutical Chemistry JSS College of Pharmacy, JSS Academy of Higher Education & Research, Ooty, Nilgiris, Tamilnadu. India
| | - Shyam Sunder Pottabatula
- Department of Pharmaceutical Chemistry JSS College of Pharmacy, JSS Academy of Higher Education & Research, Ooty, Nilgiris, Tamilnadu. India
| | - Jubie Selvaraj
- Department of Pharmaceutical Chemistry JSS College of Pharmacy, JSS Academy of Higher Education & Research, Ooty, Nilgiris, Tamilnadu. India
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Abstract
Bulevirtide (Hepcludex®), a first-in-class entry inhibitor, is being developed by MYR GmbH for the treatment of chronic hepatitis delta virus (HDV) and chronic hepatitis B virus (HBV) infections. Bulevirtide was recently approved in the European Union (EU) for the treatment of chronic HDV infection in HDV RNA positive adult patients with compensated liver disease. This article summarizes the milestones in the development of bulevirtide leading to this first approval for chronic HDV.
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Qu B, Brown RJP. Strategies to Inhibit Hepatitis B Virus at the Transcript Level. Viruses 2021; 13:v13071327. [PMID: 34372533 PMCID: PMC8310268 DOI: 10.3390/v13071327] [Citation(s) in RCA: 9] [Impact Index Per Article: 2.3] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/05/2021] [Revised: 07/02/2021] [Accepted: 07/06/2021] [Indexed: 12/11/2022] Open
Abstract
Approximately 240 million people are chronically infected with hepatitis B virus (HBV), despite four decades of effective HBV vaccination. During chronic infection, HBV forms two distinct templates responsible for viral transcription: (1) episomal covalently closed circular (ccc)DNA and (2) host genome-integrated viral templates. Multiple ubiquitous and liver-specific transcription factors are recruited onto these templates and modulate viral gene transcription. This review details the latest developments in antivirals that inhibit HBV gene transcription or destabilize viral transcripts. Notably, nuclear receptor agonists exhibit potent inhibition of viral gene transcription from cccDNA. Small molecule inhibitors repress HBV X protein-mediated transcription from cccDNA, while small interfering RNAs and single-stranded oligonucleotides result in transcript degradation from both cccDNA and integrated templates. These antivirals mediate their effects by reducing viral transcripts abundance, some leading to a loss of surface antigen expression, and they can potentially be added to the arsenal of drugs with demonstrable anti-HBV activity. Thus, these candidates deserve special attention for future repurposing or further development as anti-HBV therapeutics.
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Affiliation(s)
- Bingqian Qu
- Division of Veterinary Medicine, Paul Ehrlich Institute, 63225 Langen, Germany
- European Virus Bioinformatics Center, 07743 Jena, Germany
- Correspondence: (B.Q.); (R.J.P.B.)
| | - Richard J. P. Brown
- Division of Veterinary Medicine, Paul Ehrlich Institute, 63225 Langen, Germany
- Correspondence: (B.Q.); (R.J.P.B.)
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Ito K, Okumura A, Takeuchi JS, Watashi K, Inoue R, Yamauchi T, Sakamoto K, Yamashita Y, Iguchi Y, Une M, Wakita T, Umezawa K, Yoneda M. Dual Agonist of Farnesoid X Receptor and Takeda G Protein-Coupled Receptor 5 Inhibits Hepatitis B Virus Infection In Vitro and In Vivo. Hepatology 2021; 74:83-98. [PMID: 33434356 DOI: 10.1002/hep.31712] [Citation(s) in RCA: 21] [Impact Index Per Article: 5.3] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 06/12/2020] [Revised: 11/24/2020] [Accepted: 12/21/2020] [Indexed: 12/15/2022]
Abstract
BACKGROUND AND AIMS Chronic HBV infection is a major health problem worldwide. Currently, the first-line treatment for HBV is nucleos(t)ide analogs or interferons; however, efficient therapeutic approaches that enable cure are lacking. Therefore, anti-HBV agents with mechanisms distinct from those of current drugs are needed. Sodium taurocholate cotransporting polypeptide (NTCP) was previously identified as an HBV receptor that is inhibited by several compounds. Farnesoid X receptor (FXR) activation also inhibits NTCP function. APPROACH AND RESULTS In this study, we investigated the inhibitory effect of bile acid (BA) derivatives-namely obeticholic acid (OCA), 6α-ethyl-24-nor-5β-cholane-3α,7α,23-triol-23 sulfate sodium salt (INT-767; a dual agonist of FXR and Takeda G protein-coupled receptor [TGR5]), and 6α-ethyl-23(S)-methyl-cholic acid (INT-777; a TGR5 agonist)-3-(2,6-dichlorophenyl)-4-(3'-carboxy-2-chlorostilben-4-yl)oxymethyl-5-isopropylisoxazole (GW4064; a FXR agonist), cyclosporin A, and irbesartan. OCA and INT-777 suppressed HBV infection in HepG2-human NTCP-C4 cells. Interestingly, INT-767 showed potent inhibition by attaching to HBV particles rather than binding to NTCP. As an entry inhibitor, INT-767 was stronger than various natural BAs. Furthermore, in chimeric mice with humanized liver, INT-767 markedly delayed the initial rise of HBsAg, HBeAg, and HBV DNA and reduced covalently closed circular DNA. The strong inhibitory effect of INT-767 may be due to the cumulative effect of its ability to inhibit the entry of HBV and to stimulate FXR downstream signaling, which affects the postentry step. CONCLUSIONS Our results suggest that BA derivatives, particularly INT-767, are prospective candidate anti-HBV agents. Clarifying the underlying mechanisms of BA derivatives would facilitate the development of anti-HBV agents.
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Affiliation(s)
- Kiyoaki Ito
- Department of Gastroenterology, Aichi Medical University, Nagakute, Japan
| | - Akinori Okumura
- Department of Gastroenterology, Aichi Medical University, Nagakute, Japan
| | - Junko S Takeuchi
- Department of Virology II, National Institute of Infectious Diseases, Tokyo, Japan
| | - Koichi Watashi
- Department of Virology II, National Institute of Infectious Diseases, Tokyo, Japan
| | - Rieko Inoue
- Department of Gastroenterology, Aichi Medical University, Nagakute, Japan
| | - Taeko Yamauchi
- Department of Gastroenterology, Aichi Medical University, Nagakute, Japan
| | - Kazumasa Sakamoto
- Department of Gastroenterology, Aichi Medical University, Nagakute, Japan
| | - Yukiko Yamashita
- Faculty of Pharmaceutical Sciences, Hiroshima International University, Kure, Japan
| | - Yusuke Iguchi
- Faculty of Pharmaceutical Sciences, Hiroshima International University, Kure, Japan
| | - Mizuho Une
- Faculty of Pharmaceutical Sciences, Hiroshima International University, Kure, Japan
| | - Takaji Wakita
- Department of Virology II, National Institute of Infectious Diseases, Tokyo, Japan
| | - Kazuo Umezawa
- Department of Molecular Target Medicine Screening, Aichi Medical University School of Medicine, Nagakute, Japan
| | - Masashi Yoneda
- Department of Gastroenterology, Aichi Medical University, Nagakute, Japan
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50
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Current management & future directions in post-liver transplant recurrence of viral hepatitis. JOURNAL OF LIVER TRANSPLANTATION 2021. [DOI: 10.1016/j.liver.2021.100027] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 12/17/2022] Open
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