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Kissi-Twum AA, Pionek K, Loeb DD. The HBV variant CpF97L supports the secretion of pgRNA-containing virions at a level much greater than WT HBV. J Virol 2025; 99:e0010025. [PMID: 40231820 PMCID: PMC12090816 DOI: 10.1128/jvi.00100-25] [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: 01/16/2025] [Accepted: 03/21/2025] [Indexed: 04/16/2025] Open
Abstract
Viruses in the Hepadnaviridae family, including hepatitis B virus (HBV), replicate their double-stranded DNA (dsDNA) genomes through reverse transcription of an RNA intermediate, the pregenomic RNA (pgRNA), in the viral capsid within an infected cell. In the cell, capsids containing pgRNA, single-stranded DNA (ssDNA), and dsDNA are present. However, capsids containing dsDNA (referred to as mature genomes) are preferentially secreted in virions while only small amounts of capsids with pgRNA and ssDNA (referred to as immature genomes) are enveloped and secreted. The naturally occurring HBV core protein variant, CpF97L, is an exception; HBV CpF97L secretes high levels of ssDNA-containing virions in addition to dsDNA virions. We asked whether HBV CpF97L is capable of secreting pgRNA-containing virions as well. We found that HBV CpF97L secretes high levels of pgRNA-containing virions compared to wild-type (WT) HBV when reverse transcription was inhibited by entecavir or by the Y63F change in P protein. We detected pgRNA virions in Huh7 and HepG2 cell lines, indicating that RNA virion secretion was independent of the cell line used in virion propagation. More importantly, pgRNA virions were detected when dsDNA virions were synthesized as well. Our findings suggest that the capsids of CpF97L are constitutively matured, allowing for virions with immature genomes (ssDNA and pgRNA) to be secreted in addition to dsDNA virions.IMPORTANCEFinding a cure for hepatitis B is critical, as over 250 million people live with a hepatitis B virus (HBV) infection. HBV replicates through a series of nascent RNA and DNA intermediates in capsids, resulting in the secretion of a DNA virion to propagate the infection. HBV infections have been managed with nucleos(t)ide analogs (NAs), which terminate DNA synthesis during replication. During NA treatment, DNA levels plummet, RNA-containing capsids accumulate in infected cells and are secreted, albeit inefficiently, as virions. RNA virions in serum have therefore been proposed to be used as an indicator for covalently closed circular DNA (cccDNA) (HBV's minichromosome in hepatocytes) to determine patients who can be withdrawn from NAs without virological rebound. However, it is unknown if RNA virions are efficiently secreted by the frequent HBV variants that secrete high levels of ssDNA-containing virions, as these will lead to an erroneous overestimate of the cccDNA reservoir; hence, the need for our study.
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Affiliation(s)
- Abena Adomah Kissi-Twum
- McArdle Laboratory for Cancer Research, School of Medicine and Public Health, University of Wisconsin-Madison, Madison, Wisconsin, USA
- Microbiology Doctoral Training Program, University of Wisconsin-Madison, Madison, Wisconsin, USA
| | - Karolyn Pionek
- McArdle Laboratory for Cancer Research, School of Medicine and Public Health, University of Wisconsin-Madison, Madison, Wisconsin, USA
| | - Daniel D. Loeb
- McArdle Laboratory for Cancer Research, School of Medicine and Public Health, University of Wisconsin-Madison, Madison, Wisconsin, USA
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Li G, Liu X, Lu F. What is the real sword of HBV precore and basal core promoter mutations piercing the hepatocyte homeostasis: HBcAg or HBsAg? J Med Virol 2024; 96:e29745. [PMID: 38884414 DOI: 10.1002/jmv.29745] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/05/2024] [Revised: 05/24/2024] [Accepted: 06/08/2024] [Indexed: 06/18/2024]
Affiliation(s)
- Guixin Li
- Peking University People's Hospital, Peking University Hepatology Institute, Infectious Disease and Hepatology Center of Peking University People's Hospital，Beijing Key Laboratory of Hepatitis C and Immunotherapy for Liver Diseases, Beijing International Cooperation Base for Science and Technology on NAFLD Diagnosis, Beijing, China
| | - Xin Liu
- Department of Microbiology and Infectious Disease Center, School of Basic Medical Sciences, Peking University, Beijing, China
| | - Fengmin Lu
- Peking University People's Hospital, Peking University Hepatology Institute, Infectious Disease and Hepatology Center of Peking University People's Hospital，Beijing Key Laboratory of Hepatitis C and Immunotherapy for Liver Diseases, Beijing International Cooperation Base for Science and Technology on NAFLD Diagnosis, Beijing, China
- Department of Microbiology and Infectious Disease Center, School of Basic Medical Sciences, Peking University, Beijing, China
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Lazarevic I, Banko A, Miljanovic D, Cupic M. Hepatitis B Surface Antigen Isoforms: Their Clinical Implications, Utilisation in Diagnosis, Prevention and New Antiviral Strategies. Pathogens 2024; 13:46. [PMID: 38251353 PMCID: PMC10818932 DOI: 10.3390/pathogens13010046] [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: 12/02/2023] [Revised: 12/27/2023] [Accepted: 01/02/2024] [Indexed: 01/23/2024] Open
Abstract
The hepatitis B surface antigen (HBsAg) is a multifunctional glycoprotein composed of large (LHB), middle (MHB), and small (SHB) subunits. HBsAg isoforms have numerous biological functions during HBV infection-from initial and specific viral attachment to the hepatocytes to initiating chronic infection with their immunomodulatory properties. The genetic variability of HBsAg isoforms may play a role in several HBV-related liver phases and clinical manifestations, from occult hepatitis and viral reactivation upon immunosuppression to fulminant hepatitis and hepatocellular carcinoma (HCC). Their immunogenic properties make them a major target for developing HBV vaccines, and in recent years they have been recognised as valuable targets for new therapeutic approaches. Initial research has already shown promising results in utilising HBsAg isoforms instead of quantitative HBsAg for correctly evaluating chronic infection phases and predicting functional cures. The ratio between surface components was shown to indicate specific outcomes of HBV and HDV infections. Thus, besides traditional HBsAg detection and quantitation, HBsAg isoform quantitation can become a useful non-invasive biomarker for assessing chronically infected patients. This review summarises the current knowledge of HBsAg isoforms, their potential usefulness and aspects deserving further research.
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Affiliation(s)
- Ivana Lazarevic
- Institute of Microbiology and Immunology, Faculty of Medicine, University of Belgrade, 11000 Belgrade, Serbia; (A.B.); (D.M.); (M.C.)
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Romero S, Unchwaniwala N, Evans EL, Eliceiri KW, Loeb DD, Sherer NM. Live Cell Imaging Reveals HBV Capsid Translocation from the Nucleus To the Cytoplasm Enabled by Cell Division. mBio 2023; 14:e0330322. [PMID: 36809075 PMCID: PMC10127671 DOI: 10.1128/mbio.03303-22] [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: 11/29/2022] [Accepted: 01/17/2023] [Indexed: 02/23/2023] Open
Abstract
Hepatitis B virus (HBV) capsid assembly is traditionally thought to occur predominantly in the cytoplasm, where the virus gains access to the virion egress pathway. To better define sites of HBV capsid assembly, we carried out single cell imaging of HBV Core protein (Cp) subcellular trafficking over time under conditions supporting genome packaging and reverse transcription in Huh7 hepatocellular carcinoma cells. Time-course analyses including live cell imaging of fluorescently tagged Cp derivatives showed Cp to accumulate in the nucleus at early time points (~24 h), followed by a marked re-distribution to the cytoplasm at 48 to 72 h. Nucleus-associated Cp was confirmed to be capsid and/or high-order assemblages using a novel dual label immunofluorescence strategy. Nuclear-to-cytoplasmic re-localization of Cp occurred predominantly during nuclear envelope breakdown in conjunction with cell division, followed by strong cytoplasmic retention of Cp. Blocking cell division resulted in strong nuclear entrapment of high-order assemblages. A Cp mutant, Cp-V124W, predicted to exhibit enhanced assembly kinetics, also first trafficked to the nucleus to accumulate at nucleoli, consistent with the hypothesis that Cp's transit to the nucleus is a strong and constitutive process. Taken together, these results provide support for the nucleus as an early-stage site of HBV capsid assembly, and provide the first dynamic evidence of cytoplasmic retention after cell division as a mechanism underpinning capsid nucleus-to-cytoplasm relocalization. IMPORTANCE Hepatitis B virus (HBV) is an enveloped, reverse-transcribing DNA virus that is a major cause of liver disease and hepatocellular carcinoma. Subcellular trafficking events underpinning HBV capsid assembly and virion egress remain poorly characterized. Here, we developed a combination of fixed and long-term (>24 h) live cell imaging technologies to study the single cell trafficking dynamics of the HBV Core Protein (Cp). We demonstrate that Cp first accumulates in the nucleus, and forms high-order structures consistent with capsids, with the predominant route of nuclear egress being relocalization to the cytoplasm during cell division in conjunction with nuclear membrane breakdown. Single cell video microscopy demonstrated unequivocally that Cp's localization to the nucleus is constitutive. This study represents a pioneering application of live cell imaging to study HBV subcellular transport, and demonstrates links between HBV Cp and the cell cycle.
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Affiliation(s)
- Sofia Romero
- McArdle Laboratory for Cancer Research (Department of Oncology), University of Wisconsin-Madison School of Medicine and Public Health, Madison, Wisconsin, USA
- Institute for Molecular Virology, University of Wisconsin-Madison, Madison, Wisconsin, USA
- Microbiology Doctoral Training Program, University of Wisconsin-Madison, Madison, Wisconsin, USA
- Carbone Cancer Center, University of Wisconsin-Madison School of Medicine and Public Health, Madison, Wisconsin, USA
| | - Nuruddin Unchwaniwala
- McArdle Laboratory for Cancer Research (Department of Oncology), University of Wisconsin-Madison School of Medicine and Public Health, Madison, Wisconsin, USA
- Institute for Molecular Virology, University of Wisconsin-Madison, Madison, Wisconsin, USA
- Carbone Cancer Center, University of Wisconsin-Madison School of Medicine and Public Health, Madison, Wisconsin, USA
| | - Edward L. Evans
- Laboratory for Optical and Computational Instrumentation, Center for Quantitative Cell Imaging, University of Wisconsin-Madison, Madison, Wisconsin, USA
- Morgridge Institute for Research, Madison, Wisconsin, USA
| | - Kevin W. Eliceiri
- Laboratory for Optical and Computational Instrumentation, Center for Quantitative Cell Imaging, University of Wisconsin-Madison, Madison, Wisconsin, USA
- Morgridge Institute for Research, Madison, Wisconsin, USA
| | - Daniel D. Loeb
- McArdle Laboratory for Cancer Research (Department of Oncology), University of Wisconsin-Madison School of Medicine and Public Health, Madison, Wisconsin, USA
- Carbone Cancer Center, University of Wisconsin-Madison School of Medicine and Public Health, Madison, Wisconsin, USA
| | - Nathan M. Sherer
- McArdle Laboratory for Cancer Research (Department of Oncology), University of Wisconsin-Madison School of Medicine and Public Health, Madison, Wisconsin, USA
- Institute for Molecular Virology, University of Wisconsin-Madison, Madison, Wisconsin, USA
- Carbone Cancer Center, University of Wisconsin-Madison School of Medicine and Public Health, Madison, Wisconsin, USA
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Watanabe T, Hayashi S, Tanaka Y. Drug Discovery Study Aimed at a Functional Cure for HBV. Viruses 2022; 14:1393. [PMID: 35891374 PMCID: PMC9321005 DOI: 10.3390/v14071393] [Citation(s) in RCA: 9] [Impact Index Per Article: 3.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/31/2022] [Revised: 06/19/2022] [Accepted: 06/23/2022] [Indexed: 02/04/2023] Open
Abstract
Hepatitis B virus (HBV) causes acute and, most importantly, chronic hepatitis B worldwide. Antiviral treatments have been developed to reduce viral loads but few patients with chronic hepatitis B (CHB) achieve a functional cure. The development of new therapeutic agents is desirable. Recently, many novel agents have been developed, including drugs targeting HBV-DNA and HBV-RNA. This review provides an overview of the developmental status of these drugs, especially direct acting antiviral agents (DAAs). Serological biomarkers of HBV infection are essential for predicting the clinical course of CHB. It is also important to determine the amount and activity of covalently closed circular DNA (cccDNA) in the nuclei of infected hepatocytes. Hepatitis B core-associated antigen (HBcrAg) is a new HBV marker that has an important role in reflecting cccDNA in CHB, because it is associated with hepatic cccDNA, as well as serum HBV DNA. The highly sensitive HBcrAg (iTACT-HBcrAg) assay could be a very sensitive HBV activation marker and an alternative to HBV DNA testing for monitoring reactivation. Many of the drugs currently in clinical trials have shown efficacy in reducing hepatitis B surface antigen (HBsAg) levels. Combination therapies with DAAs and boost immune response are also under development; finding the best combinations will be important for therapeutic development.
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Affiliation(s)
| | | | - Yasuhito Tanaka
- Department of Gastroenterology and Hepatology, Faculty of Life Sciences, Kumamoto University, Kumamoto 860-8556, Japan; (T.W.); (S.H.)
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Abstract
Hepatitis B virus (HBV) is a hepatotropic virus and an important human pathogen. There are an estimated 296 million people in the world that are chronically infected by this virus, and many of them will develop severe liver diseases including hepatitis, cirrhosis and hepatocellular carcinoma (HCC). HBV is a small DNA virus that replicates via the reverse transcription pathway. In this review, we summarize the molecular pathways that govern the replication of HBV and its interactions with host cells. We also discuss viral and non-viral factors that are associated with HBV-induced carcinogenesis and pathogenesis, as well as the role of host immune responses in HBV persistence and liver pathogenesis.
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Affiliation(s)
- Yu-Chen Chuang
- Department of Molecular Microbiology and Immunology, University of Southern California Keck School of Medicine, Los Angeles, CA 90089, USA
| | - Kuen-Nan Tsai
- Department of Molecular Microbiology and Immunology, University of Southern California Keck School of Medicine, Los Angeles, CA 90089, USA
| | - Jing-Hsiung James Ou
- Department of Molecular Microbiology and Immunology, University of Southern California Keck School of Medicine, Los Angeles, CA 90089, USA
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Regulation of Hepatitis B Virus Virion Release and Envelopment Timing by Nucleocapsid and Envelope Interactions. J Virol 2021; 96:e0130521. [PMID: 34643434 DOI: 10.1128/jvi.01305-21] [Citation(s) in RCA: 6] [Impact Index Per Article: 1.5] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/30/2023] Open
Abstract
Interactions between the N-terminal (assembly) domain (NTD) and the linker region of the hepatitis B virus (HBV) capsid protein and the large (L) envelope protein are required for virion formation, which occurs via budding of cytoplasmic mature nucleocapsids (NCs) containing the relaxed circular (RC) DNA genome into an intracellular membrane compartment containing viral envelope proteins. L-capsid interactions also negatively regulates covalently closed circular (CCC) DNA formation, which occurs after RC DNA release from mature NCs and nuclear import. We have now found that L could increase RC DNA in cytoplasmic mature NCs that are destabilized due to mutations in the NTD or the linker, even in those that apparently fail to support secretion of complete virions extracellularly. Other mutations in the capsid linker could block the effects of L on both cytoplasmic NC DNA and nuclear CCC DNA. Furthermore, the maturity of RC DNA in cytoplasmic NCs that was enhanced by L or found in secreted virions was modulated by the capsid linker sequence. The level and maturity of the cytoplasmic RC DNA was further influenced by the efficiency of extracellular virion secretion dependent on viral genotype-specific envelope proteins. These results suggest that interactions between the capsid and envelope proteins regulate one or more steps during virion secretion beyond initial capsid envelopment, and highlights the critical role of the capsid linker in regulating capsid-envelope interaction, including the timing of envelopment during NC maturation. Importance Hepatitis B virus (HBV) is a major human pathogen causing serious liver diseases including cancer. The interactions between the HBV capsid and the large (L) envelope protein is required for formation of infectious viral particles and also negatively regulate formation of an HBV DNA episome in the host cell nucleus, which serves as the sole transcriptional template capable of supporting all viral gene expression to sustain HBV replication and therefore, is the molecular basis of HBV persistence. Here, we report evidence indicating that L-capsid interactions modulate the timing of formation of infectious HBV particles during replication and facilitate extracellular release following their formation. Furthermore, a short linker sequence in the capsid protein plays a critical role in these processes as well as controls the amplification of the nuclear episome. These findings inform fundamental mechanisms of HBV replication as well as antiviral development targeting the HBV capsid and DNA episome.
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Yamasaki M, Matsuda N, Matoba K, Kondo S, Kanegae Y, Saito I, Nomoto A. Acetophenone 4-nitrophenylhydrazone inhibits Hepatitis B virus replication by modulating capsid assembly. Virus Res 2021; 306:198565. [PMID: 34555437 DOI: 10.1016/j.virusres.2021.198565] [Citation(s) in RCA: 3] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/24/2021] [Revised: 09/08/2021] [Accepted: 09/09/2021] [Indexed: 02/07/2023]
Abstract
Hepatitis B virus (HBV) is the causative agent of chronic liver disease and is correlated with the development of subsequent hepatic cirrhosis and hepatocellular carcinoma. Current antiviral therapy using nucleos(t)ide analogs is effective in suppressing viral replication and interrupting disease progression, but HBV is rarely cured completely. Thus, there remains an unmet need for the development of novel anti-HBV drugs. Here, we report the identification of N-(4-Nitrophenyl)-1-phenylethanone hydrazone (ANPH) as a novel structural class of selective inhibitors targeting the replication of the HBV genome using adenovirus vector-mediated HBV genome transduction. ANPH inhibited viral genome replication in HepG2.2.15 cells by inducing the formation of empty capsids devoid of pregenomic RNA without affecting its transcription and translation. Biochemical assays using a truncated core protein consisting of the assembly domain showed that ANPH accelerates the formation of morphologically intact capsids. Taken together, we propose that ANPH might provide a new structural scaffold to design a new anti-HBV drug in medicinal chemistry as well as chemical probes for HBV core protein functions in the future.
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Affiliation(s)
- Manabu Yamasaki
- Institute of Microbial Chemistry (BIKAKEN), Shinagawa-ku, Tokyo, Japan.
| | - Norie Matsuda
- Institute of Microbial Chemistry (BIKAKEN), Shinagawa-ku, Tokyo, Japan
| | - Kazuaki Matoba
- Institute of Microbial Chemistry (BIKAKEN), Shinagawa-ku, Tokyo, Japan
| | - Saki Kondo
- Laboratory of Molecular Genetics, The Institute of Medical Science, The University of Tokyo, Minato-ku, Tokyo, Japan
| | - Yumi Kanegae
- Laboratory of Molecular Genetics, The Institute of Medical Science, The University of Tokyo, Minato-ku, Tokyo, Japan; Core Research Facilities of Basic Science (Molecular Genetics), Research Center for Medical Science, Jikei University School of Medicine
| | - Izumu Saito
- Laboratory of Molecular Genetics, The Institute of Medical Science, The University of Tokyo, Minato-ku, Tokyo, Japan
| | - Akio Nomoto
- Institute of Microbial Chemistry (BIKAKEN), Shinagawa-ku, Tokyo, Japan
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Wei L, Ploss A. Mechanism of Hepatitis B Virus cccDNA Formation. Viruses 2021; 13:v13081463. [PMID: 34452329 PMCID: PMC8402782 DOI: 10.3390/v13081463] [Citation(s) in RCA: 67] [Impact Index Per Article: 16.8] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/11/2021] [Revised: 07/14/2021] [Accepted: 07/21/2021] [Indexed: 02/06/2023] Open
Abstract
Hepatitis B virus (HBV) remains a major medical problem affecting at least 257 million chronically infected patients who are at risk of developing serious, frequently fatal liver diseases. HBV is a small, partially double-stranded DNA virus that goes through an intricate replication cycle in its native cellular environment: human hepatocytes. A critical step in the viral life-cycle is the conversion of relaxed circular DNA (rcDNA) into covalently closed circular DNA (cccDNA), the latter being the major template for HBV gene transcription. For this conversion, HBV relies on multiple host factors, as enzymes capable of catalyzing the relevant reactions are not encoded in the viral genome. Combinations of genetic and biochemical approaches have produced findings that provide a more holistic picture of the complex mechanism of HBV cccDNA formation. Here, we review some of these studies that have helped to provide a comprehensive picture of rcDNA to cccDNA conversion. Mechanistic insights into this critical step for HBV persistence hold the key for devising new therapies that will lead not only to viral suppression but to a cure.
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Zhang X, Wang X, Wu M, Ghildyal R, Yuan Z. Animal Models for the Study of Hepatitis B Virus Pathobiology and Immunity: Past, Present, and Future. Front Microbiol 2021; 12:715450. [PMID: 34335553 PMCID: PMC8322840 DOI: 10.3389/fmicb.2021.715450] [Citation(s) in RCA: 12] [Impact Index Per Article: 3.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/27/2021] [Accepted: 06/18/2021] [Indexed: 12/11/2022] Open
Abstract
Hepatitis B virus (HBV) infection is a global public health problem that plagues approximately 240 million people. Chronic hepatitis B (CHB) often leads to liver inflammation and aberrant repair which results in diseases ranging from liver fibrosis, cirrhosis, to hepatocellular carcinoma. Despite its narrow species tropism, researchers have established various in vivo models for HBV or its related viruses which have provided a wealth of knowledge on viral lifecycle, pathogenesis, and immunity. Here we briefly revisit over five decades of endeavor in animal model development for HBV and summarize their advantages and limitations. We also suggest directions for further improvements that are crucial for elucidation of the viral immune-evasion strategies and for development of novel therapeutics for a functional cure.
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Affiliation(s)
- Xiaonan Zhang
- Centre for Research in Therapeutic Solutions, Faculty of Science and Technology, University of Canberra, Canberra, ACT, Australia.,Shanghai Public Health Clinical Center, Fudan University, Shanghai, China
| | - Xiaomeng Wang
- Shanghai Public Health Clinical Center, Fudan University, Shanghai, China
| | - Min Wu
- Shanghai Public Health Clinical Center, Fudan University, Shanghai, China
| | - Reena Ghildyal
- Centre for Research in Therapeutic Solutions, Faculty of Science and Technology, University of Canberra, Canberra, ACT, Australia
| | - Zhenghong Yuan
- Key Laboratory of Medical Molecular Virology, School of Basic Medical Sciences, Shanghai Medical College, Fudan University, Shanghai, China
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Tsounis EP, Tourkochristou E, Mouzaki A, Triantos C. Toward a new era of hepatitis B virus therapeutics: The pursuit of a functional cure. World J Gastroenterol 2021; 27:2727-2757. [PMID: 34135551 PMCID: PMC8173382 DOI: 10.3748/wjg.v27.i21.2727] [Citation(s) in RCA: 32] [Impact Index Per Article: 8.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 01/20/2021] [Revised: 02/23/2021] [Accepted: 04/13/2021] [Indexed: 02/06/2023] Open
Abstract
Hepatitis B virus (HBV) infection, although preventable by vaccination, remains a global health problem and a major cause of chronic liver disease. Although current treatment strategies suppress viral replication very efficiently, the optimal endpoint of hepatitis B surface antigen (HBsAg) clearance is rarely achieved. Moreover, the thorny problems of persistent chromatin-like covalently closed circular DNA and the presence of integrated HBV DNA in the host genome are ignored. Therefore, the scientific community has focused on developing innovative therapeutic approaches to achieve a functional cure of HBV, defined as undetectable HBV DNA and HBsAg loss over a limited treatment period. A deeper understanding of the HBV life cycle has led to the introduction of novel direct-acting antivirals that exert their function through multiple mechanisms, including inhibition of viral entry, transcriptional silencing, epigenetic manipulation, interference with capsid assembly, and disruption of HBsAg release. In parallel, another category of new drugs aims to restore dysregulated immune function in chronic hepatitis B accompanied by lethargic cellular and humoral responses. Stimulation of innate immunity by pattern-recognition receptor agonists leads to upregulation of antiviral cytokine expression and appears to contribute to HBV containment. Immune checkpoint inhibitors and adoptive transfer of genetically engineered T cells are breakthrough technologies currently being explored that may elicit potent HBV-specific T-cell responses. In addition, several clinical trials are attempting to clarify the role of therapeutic vaccination in this setting. Ultimately, it is increasingly recognized that elimination of HBV requires a treatment regimen based on a combination of multiple drugs. This review describes the rationale for progressive therapeutic interventions and discusses the latest findings in the field of HBV therapeutics.
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Affiliation(s)
- Efthymios P Tsounis
- Division of Gastroenterology, Department of Internal Medicine, Medical School, University of Patras, Patras 26504, Greece
| | - Evanthia Tourkochristou
- Division of Gastroenterology, Department of Internal Medicine, Medical School, University of Patras, Patras 26504, Greece
| | - Athanasia Mouzaki
- Division of Hematology, Department of Internal Medicine, Medical School, University of Patras, Patras 26504, Greece
| | - Christos Triantos
- Division of Gastroenterology, Department of Internal Medicine, Medical School, University of Patras, Patras 26504, Greece
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12
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Goyal A. Modeling reveals no direct role of the extent of HBV DNA integrations on the outcome of infection. J Theor Biol 2021; 526:110793. [PMID: 34087271 DOI: 10.1016/j.jtbi.2021.110793] [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: 03/17/2021] [Revised: 05/15/2021] [Accepted: 05/30/2021] [Indexed: 11/27/2022]
Abstract
Hepatitis B virus (HBV) with its high prevalence and death toll is one of the most important infectious diseases to study. Yet, there is very little progress in the development of within-host models for HBV, which has subsequently hindered our understanding of this virus. The uncertainty around the proliferation of infected hepatocytes has been studied but never in association with other important biological continuous events such as integrations and superinfections. This is despite the fact that these processes affect the diversity and composition of infected cell population in the liver and an improved understanding of the cellular composition will undoubtedly assist in strategizing against this viral infection. Here, we developed novel mathematical models that incorporate these key biological processes and analyzed them both analytically and numerically. Unaffected by the extent of integrated DNA (IDNA), the outcome of HBV infection was primarily dictated by the balance between processes generating and killing infected hepatocytes containing covalent closed circular DNA (cccDNA). The superinfection was found to be a key process in the spread of HBV infection as its exclusion could not reproduce experimentally observed composition of infected hepatocytes at peak of acute HBV infection, a stage where our model predicts that infected hepatocytes most likely carry both cccDNA and IDNA. Our analysis further suggested the existence of some form of selective advantage of infected hepatocytes containing only IDNA to explain the viral dynamics observed during antiviral treatment and the transition from peak to acute infection. Finally, the fine line between liver destruction and resolution of acute HBV infection was found to be highly influenced by the fate of cccDNA during cellular proliferation.
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Affiliation(s)
- Ashish Goyal
- Vaccine and Infectious Disease Division, Fred Hutchinson Cancer Research Center, United States
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13
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Regulation of Hepatitis B Virus Replication by Cyclin Docking Motifs in Core Protein. J Virol 2021; 95:JVI.00230-21. [PMID: 33789995 DOI: 10.1128/jvi.00230-21] [Citation(s) in RCA: 8] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/09/2021] [Accepted: 03/27/2021] [Indexed: 12/16/2022] Open
Abstract
Hepatitis B virus (HBV) capsid or core protein (HBc) consists of an N-terminal domain (NTD) and a C-terminal domain (CTD) connected by a short linker peptide. Dynamic phosphorylation and dephosphorylation of HBc regulate its multiple functions in capsid assembly and viral replication. The cellular cyclin-dependent kinase 2 (CDK2) plays a major role in HBc phosphorylation and, furthermore, is incorporated into the viral capsid, accounting for most of the "endogenous kinase" activity associated with the capsid. The packaged CDK2 is thought to play a role in phosphorylating HBc to trigger nucleocapsid disassembly (uncoating), an essential step during viral infection. However, little is currently known on how CDK2 is recruited and packaged into the capsid. We have now identified three RXL motifs in the HBc NTD known as cyclin docking motifs (CDMs), which mediate the interactions of various CDK substrates/regulators with CDK/cyclin complexes. Mutations of the CDMs in the HBc NTD reduced CTD phosphorylation and diminished CDK2 packaging into the capsid. Also, the CDM mutations showed little effects on capsid assembly and pregenomic RNA (pgRNA) packaging but impaired the integrity of mature nucleocapsids. Furthermore, the CDM mutations blocked covalently closed circular DNA (CCC DNA) formation during infection while having no effect on or enhancing CCC DNA formation via intracellular amplification. These results indicate that the HBc NTD CDMs play a role in CDK2 recruitment and packaging, which, in turn, is important for productive infection.IMPORTANCE Hepatitis B virus (HBV) is an important global human pathogen and persistently infects hundreds of millions of people, who are at high risk of cirrhosis and liver cancer. HBV capsid packages a host cell protein kinase, the cyclin-dependent kinase 2 (CDK2), which is thought to be required to trigger disassembly of the viral nucleocapsid during infection by phosphorylating the capsid protein, a prerequisite for successful infection. We have identified docking sites on the capsid protein for recruiting CDK2, in complex with its cyclin partner, to facilitate capsid protein phosphorylation and CDK2 packaging. Mutations of these docking sites reduced capsid protein phosphorylation, impaired CDK2 packaging into HBV capsids, and blocked HBV infection. These results provide novel insights regarding CDK2 packaging into HBV capsids and the role of CDK2 in HBV infection and should facilitate the development of antiviral drugs that target the HBV capsid protein.
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Xi J, Luckenbaugh L, Hu J. Multiple roles of PP2A binding motif in hepatitis B virus core linker and PP2A in regulating core phosphorylation state and viral replication. PLoS Pathog 2021; 17:e1009230. [PMID: 33493210 PMCID: PMC7861550 DOI: 10.1371/journal.ppat.1009230] [Citation(s) in RCA: 32] [Impact Index Per Article: 8.0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/09/2020] [Revised: 02/04/2021] [Accepted: 12/08/2020] [Indexed: 12/18/2022] Open
Abstract
Hepatitis B virus (HBV) capsid or core protein (HBc) contains an N-terminal domain (NTD) and a C-terminal domain (CTD) connected by a short linker peptide. HBc plays a critical role in virtually every step of viral replication, which is further modulated by dynamic phosphorylation and dephosphorylation of its CTD. While several cellular kinases have been identified that mediate HBc CTD phosphorylation, there is little information on the cellular phosphatases that mediate CTD dephosphorylation. Herein, a consensus binding motif for the protein phosphatase 2A (PP2A) regulatory subunit B56 was recognized within the HBc linker peptide. Mutations within this motif designed to block or enhance B56 binding showed pleiotropic effects on CTD phosphorylation state as well as on viral RNA packaging, reverse transcription, and virion secretion. Furthermore, linker mutations affected the HBV nuclear episome (the covalently closed circular or CCC DNA) differentially during intracellular amplification vs. infection. The effects of linker mutations on CTD phosphorylation state varied with different phosphorylation sites and were only partially consistent with the linker motif serving to recruit PP2A-B56, specifically, to dephosphorylate CTD, suggesting that multiple phosphatases and/or kinases may be recruited to modulate CTD (de)phosphorylation. Furthermore, pharmacological inhibition of PP2A could decrease HBc CTD dephosphorylation and increase the nuclear HBV episome. These results thus strongly implicate the HBc linker in recruiting PP2A and other host factors to regulate multiple stages of HBV replication. Hepatitis B virus (HBV) causes acute and chronic viral hepatitis, liver fibrosis, cirrhosis and cancer. The dynamic phosphorylation and dephosphorylation of the viral capsid protein (HBc), which are controlled by host cell protein kinases and phosphatases, play a critical role in regulating multiple stages of HBV replication. While a number of cellular kinases have been identified that mediate HBc phosphorylation, there is little information on cellular phosphatases that mediate its dephosphorylation. Herein we have identified a consensus binding motif in HBc for one of the major cellular phosphatases, the protein phosphatase 2A (PP2A). Genetic analysis of this motif revealed that it played multiple roles in regulating CTD phosphorylation state, as well as viral RNA packaging, reverse transcription, virion secretion, and formation of the nuclear HBV episome responsible for viral persistence. Furthermore, pharmacological inhibition of PP2A decreased HBc dephosphorylation and increased the nuclear episome, further supporting a role of PP2A in HBc dephosphorylation and HBV persistence. These results thus suggest that HBc recruits PP2A, among other host factors, to regulate HBc phosphorylation and dephosphorylation dynamics and HBV replication and persistence.
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Affiliation(s)
- Ji Xi
- Department of Microbiology and Immunology, The Pennsylvania State University College of Medicine, Hershey, Pennsylvania, United States of America
| | - Laura Luckenbaugh
- Department of Microbiology and Immunology, The Pennsylvania State University College of Medicine, Hershey, Pennsylvania, United States of America
| | - Jianming Hu
- Department of Microbiology and Immunology, The Pennsylvania State University College of Medicine, Hershey, Pennsylvania, United States of America
- * E-mail:
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15
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Sun H, Chang L, Yan Y, Wang L. Hepatitis B virus pre-S region: Clinical implications and applications. Rev Med Virol 2020; 31. [PMID: 33314434 DOI: 10.1002/rmv.2201] [Citation(s) in RCA: 5] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/03/2020] [Revised: 11/22/2020] [Accepted: 11/29/2020] [Indexed: 12/12/2022]
Abstract
Hepatitis B virus (HBV) infection is a major threat to global public health, which can result in many acute and chronic liver diseases. HBV, a member of the family Hepadnaviridae, is a small enveloped DNA virus containing a circular genome of 3.2 kb. Located upstream of the S-open-reading frame of the HBV genome is the pre-S region, which is vital to the viral life cycle. The pre-S region has high variability and many mutations in the pre-S region are associated with several liver diseases, such as fulminant hepatitis (FH), liver cirrhosis (LC), and hepatocellular carcinoma (HCC). In addition, the pre-S region has been applied in the development of several pre-S-based materials and systems to prevent or treat HBV infection. In conclusion, the pre-S region plays an essential role in the occurrence, diagnosis, and treatment of HBV-related liver diseases, which may provide a novel perspective for the study of HBV infection and relevant diseases.
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Affiliation(s)
- Huizhen Sun
- National Center for Clinical Laboratories, Beijing Hospital, National Center of Gerontology; Institute of Geriatric Medicine, Chinese Academy of Medical Sciences, Beijing, PR China
- Beijing Engineering Research Center of Laboratory Medicine, Beijing Hospital, Beijing, PR China
- Graduate School, Peking Union Medical College, Chinese Academy of Medical Sciences, Beijing, PR China
| | - Le Chang
- National Center for Clinical Laboratories, Beijing Hospital, National Center of Gerontology; Institute of Geriatric Medicine, Chinese Academy of Medical Sciences, Beijing, PR China
- Beijing Engineering Research Center of Laboratory Medicine, Beijing Hospital, Beijing, PR China
| | - Ying Yan
- National Center for Clinical Laboratories, Beijing Hospital, National Center of Gerontology; Institute of Geriatric Medicine, Chinese Academy of Medical Sciences, Beijing, PR China
- Beijing Engineering Research Center of Laboratory Medicine, Beijing Hospital, Beijing, PR China
| | - Lunan Wang
- National Center for Clinical Laboratories, Beijing Hospital, National Center of Gerontology; Institute of Geriatric Medicine, Chinese Academy of Medical Sciences, Beijing, PR China
- Beijing Engineering Research Center of Laboratory Medicine, Beijing Hospital, Beijing, PR China
- Graduate School, Peking Union Medical College, Chinese Academy of Medical Sciences, Beijing, PR China
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16
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Marchetti AL, Guo H. New Insights on Molecular Mechanism of Hepatitis B Virus Covalently Closed Circular DNA Formation. Cells 2020; 9:cells9112430. [PMID: 33172220 PMCID: PMC7694973 DOI: 10.3390/cells9112430] [Citation(s) in RCA: 33] [Impact Index Per Article: 6.6] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/12/2020] [Revised: 11/03/2020] [Accepted: 11/04/2020] [Indexed: 12/15/2022] Open
Abstract
The chronic factor of the Hepatitis B Virus (HBV), specifically the covalently closed circular DNA (cccDNA), is a highly stable and active viral episomal genome established in the livers of chronic hepatitis B patients as a constant source of disease. Being able to target and eliminate cccDNA is the end goal for a genuine cure for HBV. Yet how HBV cccDNA is formed from the viral genomic relaxed circular DNA (rcDNA) and by what host factors had been long-standing research questions. It is generally acknowledged that HBV hijacks cellular functions to turn the open circular DNA conformation of rcDNA into cccDNA through DNA repair mechanisms. With great efforts from the HBV research community, there have been several recent leaps in our understanding of cccDNA formation. It is our goal in this review to analyze the recent reports showing evidence of cellular factor's involvement in the molecular pathway of cccDNA biosynthesis.
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Affiliation(s)
- Alexander L. Marchetti
- Department of Microbiology and Immunology, School of Medicine, Indiana University, Indianapolis, IN 46202, USA;
- Cancer Virology Program, Hillman Cancer Center, School of Medicine, University of Pittsburgh, Pittsburgh, PA 15213, USA
| | - Haitao Guo
- Cancer Virology Program, Hillman Cancer Center, School of Medicine, University of Pittsburgh, Pittsburgh, PA 15213, USA
- Department of Microbiology and Molecular Genetics, University of Pittsburgh, Pittsburgh, PA 15213, USA
- Correspondence:
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17
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He JH, Xia Q, Weng S, He J, Xu X. Identification of infectious spleen and kidney necrosis virus (ISKNV)-encoded microRNAs. Virus Genes 2020; 56:724-733. [PMID: 33033882 DOI: 10.1007/s11262-020-01798-6] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/05/2020] [Accepted: 09/25/2020] [Indexed: 10/23/2022]
Abstract
MicroRNAs (miRNAs) are small noncoding RNAs that post-transcriptionally regulate gene expression by complementary binding to target mRNAs. Virus-encoded miRNAs play important roles in virus life cycle and virus-host interactions. Viruses from the Megalocytivirus genus, family Iridoviridae, infect a wide range of fishes, bringing great challenges to aquaculture. Infectious spleen and kidney necrosis virus (ISKNV) is the type species of the Megalocytivirus genus. In this study, using Illumina sequencing coupled with miRNA precursor prediction and stem-loop real-time PCR, 14 putative ISKNV-encoded miRNAs were preliminarily identified from ISKNV-infected mandarin fish MFF-1 cells. To initially study their functions, inhibitors of the 14 viral miRNAs were synthesized and transfected into MFF-1 cells, which were further infected with ISKNV. The results showed that these viral miRNAs could affect the virus titers in the supernatant of ISKNV-infected cells and the expression of major capsid protein (MCP). Moreover, we observed that inhibition of several ISKNV miRNAs had different effects on MCP expression and on titer of released virus, suggesting complex roles of viral miRNAs in ISKNV infection. The current study may provide a fundamental information for further identification and functional studies on miRNAs encoded by Megalocytivirus.
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Affiliation(s)
- Jian-Hui He
- State Key Laboratory of Biocontrol, School of Life Sciences, Sun Yat-Sen University, Guangzhou, 510275, People's Republic of China
| | - Qiong Xia
- State Key Laboratory of Biocontrol, School of Life Sciences, Sun Yat-Sen University, Guangzhou, 510275, People's Republic of China
| | - Shaoping Weng
- State Key Laboratory of Biocontrol, School of Life Sciences, Sun Yat-Sen University, Guangzhou, 510275, People's Republic of China.,Southern Marine Science and Engineering Guangdong Laboratory (Zhuhai), Sun Yat-Sen University, Guangzhou, People's Republic of China.,Institute of Aquatic Economic Animals and Guangdong Provice Key Laboratory for Aquatic Economic Animals, Sun Yat-Sen University, Guangzhou, People's Republic of China
| | - Jianguo He
- State Key Laboratory of Biocontrol, School of Life Sciences, Sun Yat-Sen University, Guangzhou, 510275, People's Republic of China. .,Southern Marine Science and Engineering Guangdong Laboratory (Zhuhai), Sun Yat-Sen University, Guangzhou, People's Republic of China. .,Institute of Aquatic Economic Animals and Guangdong Provice Key Laboratory for Aquatic Economic Animals, Sun Yat-Sen University, Guangzhou, People's Republic of China.
| | - Xiaopeng Xu
- State Key Laboratory of Biocontrol, School of Life Sciences, Sun Yat-Sen University, Guangzhou, 510275, People's Republic of China. .,Southern Marine Science and Engineering Guangdong Laboratory (Zhuhai), Sun Yat-Sen University, Guangzhou, People's Republic of China. .,Institute of Aquatic Economic Animals and Guangdong Provice Key Laboratory for Aquatic Economic Animals, Sun Yat-Sen University, Guangzhou, People's Republic of China.
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18
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Xia Y, Guo H. Hepatitis B virus cccDNA: Formation, regulation and therapeutic potential. Antiviral Res 2020; 180:104824. [PMID: 32450266 PMCID: PMC7387223 DOI: 10.1016/j.antiviral.2020.104824] [Citation(s) in RCA: 125] [Impact Index Per Article: 25.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/29/2020] [Revised: 05/03/2020] [Accepted: 05/18/2020] [Indexed: 02/06/2023]
Abstract
Hepatitis B virus (HBV) infection remains a major public health concern worldwide with about 257 million individuals chronically infected. Current therapies can effectively control HBV replication and slow down disease progress, but cannot cure HBV infection. Upon infection, HBV establishes a pool of covalently closed circular DNA (cccDNA) in the nucleus of infected hepatocytes. The cccDNA exists as a minichromosome and resists to antivirals, thus a therapeutic eradication of cccDNA from the infected cells remains unattainable. In this review, we summarize the state of knowledge on the mechanisms underlying cccDNA formation and regulation, and discuss the possible strategies that may contribute to the eradication of HBV through targeting cccDNA.
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Affiliation(s)
- Yuchen Xia
- State Key Laboratory of Virology and Hubei Province Key Laboratory of Allergy and Immunology, School of Basic Medical Sciences, Wuhan University, Wuhan, China.
| | - Haitao Guo
- UPMC Hillman Cancer Center, Department of Microbiology and Molecular Genetics, University of Pittsburgh, Pittsburgh, PA, USA.
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19
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Tu T, Zehnder B, Qu B, Ni Y, Main N, Allweiss L, Dandri M, Shackel N, George J, Urban S. A novel method to precisely quantify hepatitis B virus covalently closed circular (ccc)DNA formation and maintenance. Antiviral Res 2020; 181:104865. [PMID: 32726641 DOI: 10.1016/j.antiviral.2020.104865] [Citation(s) in RCA: 17] [Impact Index Per Article: 3.4] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/30/2020] [Revised: 06/21/2020] [Accepted: 06/23/2020] [Indexed: 01/05/2023]
Abstract
Hepatitis B virus (HBV) is the major cause of virus-associated liver disease. Persistent HBV infection is maintained by its episomal genome (covalently closed circular DNA, cccDNA), which acts as a template for viral transcripts. The formation of cccDNA is poorly characterised due to limited ability to quantify it accurately in the presence of replicative intermediates. Here, we describe a novel cccDNA quantification assay (cccDNA inversion quantitative PCR, cinqPCR), which uses restriction enzymes to invert a DNA sequence close to the gap region of Genotype D HBV strains, including the isolate widely used in experimental studies. Importantly, cinqPCR allows simultaneous normalisation to cellular DNA in a single reaction, provides absolute copy numbers without requiring a standard curve, and has high precision, sensitivity, and specificity for cccDNA compared to previous assays. We first established that cinqPCR gives values consistent with classical approaches in both in vitro and in vivo (humanised mice) HBV infections. We then used cinqPCR to find that cccDNA is formed within 12 h post-inoculation (hpi). cccDNA formation slowed by 28 hpi despite de novo synthesis of HBV DNA, indicating inefficient conversion of new viral genomes to cccDNA within infected cells. Finally, we show that cinqPCR can be used as a 96-well screening assay. Thus, we have developed an ideal method for testing current and future anti-cccDNA therapeutics with high precision and sensitivity.
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Affiliation(s)
- Thomas Tu
- Department of Infectious Diseases, Molecular Virology, University Hospital Heidelberg, Heidelberg, Germany; German Center for Infection Research (DZIF), Heidelberg Partner Site, Heidelberg, Germany; Storr Liver Centre, Westmead Institute for Medical Research, Westmead Hospital and University of Sydney, Westmead, NSW, Australia.
| | - Benno Zehnder
- Department of Infectious Diseases, Molecular Virology, University Hospital Heidelberg, Heidelberg, Germany
| | - Bingqian Qu
- Department of Infectious Diseases, Molecular Virology, University Hospital Heidelberg, Heidelberg, Germany
| | - Yi Ni
- Department of Infectious Diseases, Molecular Virology, University Hospital Heidelberg, Heidelberg, Germany
| | - Nathan Main
- Gastroenterology and Liver Laboratory, Ingham Institute for Applied Medical Research, Sydney, Australia; Department of Gastroenterology and Hepatology, Liverpool Hospital, Sydney, New South Wales, Australia
| | - Lena Allweiss
- Department of Internal Medicine, Center for Internal Medicine, University Medical Center Hamburg-Eppendorf, Hamburg, Germany
| | - Maura Dandri
- Department of Internal Medicine, Center for Internal Medicine, University Medical Center Hamburg-Eppendorf, Hamburg, Germany; DZIF, Hamburg-Lübeck-Borstel Partner Site, Germany
| | - Nicholas Shackel
- Gastroenterology and Liver Laboratory, Ingham Institute for Applied Medical Research, Sydney, Australia; Department of Gastroenterology and Hepatology, Liverpool Hospital, Sydney, New South Wales, Australia; South Western Sydney Clinical School, University of New South Wales, Australia
| | - Jacob George
- Storr Liver Centre, Westmead Institute for Medical Research, Westmead Hospital and University of Sydney, Westmead, NSW, Australia
| | - Stephan Urban
- Department of Infectious Diseases, Molecular Virology, University Hospital Heidelberg, Heidelberg, Germany; German Center for Infection Research (DZIF), Heidelberg Partner Site, Heidelberg, Germany
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20
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Lazarevic I, Banko A, Miljanovic D, Cupic M. Biological features of hepatitis B virus strains associated with fulminant hepatitis. Future Virol 2020. [DOI: 10.2217/fvl-2020-0011] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/21/2022]
Abstract
Accumulating evidence suggests that hepatitis B virus (HBV) biological features may influence the course and clinical manifestations of infection and possibly the development of fulminant hepatitis (FH). Since HBV is not a cytocidal virus, virus-induced liver damage results from an interplay between the virus replication and the host's defense. Therefore, viral factors contributing to enhanced replication, induction of a stronger immune attack or apoptosis of hepatocytes could be crucial in development of FH. Numerous mutations in basal core promoter, pre-C, C and S regions of the HBV genome contribute to development of FH by different mechanisms, including enhanced viral replication, the loss of a decoy for immune response, unbalanced expression of viral proteins and retention of unprocessed cytotoxic proteins in hepatocytes.
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Affiliation(s)
- Ivana Lazarevic
- Institute of Microbiology & Immunology, Faculty of Medicine, University of Belgrade, Dr Subotica 1, 11000 Belgrade, Serbia
| | - Ana Banko
- Institute of Microbiology & Immunology, Faculty of Medicine, University of Belgrade, Dr Subotica 1, 11000 Belgrade, Serbia
| | - Danijela Miljanovic
- Institute of Microbiology & Immunology, Faculty of Medicine, University of Belgrade, Dr Subotica 1, 11000 Belgrade, Serbia
| | - Maja Cupic
- Institute of Microbiology & Immunology, Faculty of Medicine, University of Belgrade, Dr Subotica 1, 11000 Belgrade, Serbia
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21
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Cavallone D, Ricco G, Oliveri F, Colombatto P, Moriconi F, Coco B, Romagnoli V, Salvati A, Surace L, Bonino F, Brunetto MR. Do the circulating Pre-S/S quasispecies influence hepatitis B virus surface antigen levels in the HBeAg negative phase of HBV infection? Aliment Pharmacol Ther 2020; 51:1406-1416. [PMID: 32390175 DOI: 10.1111/apt.15753] [Citation(s) in RCA: 4] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 10/25/2019] [Revised: 11/19/2019] [Accepted: 04/06/2020] [Indexed: 12/18/2022]
Abstract
BACKGROUND Virus, host factors and their interplay influence Hepatitis B surface Antigen serum levels during Hepatitis B Virus (HBV) infection course and treatment. AIM To study the Pre-S/S circulating quasispecies in a cohort of untreated, HBeAg negative, genotype-D, HBsAg carriers. METHODS We studied 260 carriers: 71 with HBeAg negative infection (ENI; HBV-DNA ≤2000 IU/mL); 42 Grey Zone (GZ; HBV-DNA ≤20 000 IU/mL); 82 chronic hepatitis (CH) and 65 cirrhosis (CI) (HBV-DNA > 20 000 IU/mL). Population sequencing was applied to identify Pre-S/S gene mutations responsible for any amino acid substitution or potential biological/antigenic implications (M-muts) on HBsAg. RESULTS HBsAg serum levels were lower in ENI + GZ than in CH + CI (2.61 [-1.10/4.06] vs 3.62 [2.41/4.92] log10 IU/mL, P < 0.001) and in CI than CH (3.48 [2.41/4.38] vs 3.66 [2.57/4.92] log10 IU/mL, P < 0.001). M-muts were found in 73 (28.1%) cases: 5 (7.0%) ENI, 3 (7.1%) GZ, 26 (31.7%) CH, 39 (60.0%) CI (P < 0.001) and mostly in Pre-S2 (17.6%) than Pre-S1 (5.8%) and Small-S (10.8%; P < 0.001). Overall HBsAg serum levels were higher in carriers with M-muts (3.56 [0.95/4.38] vs 3.17 [-1.10/4.92] log10 IU/mL, P < 0.001), but comparable in carriers with or without M-mut when considering separately ENI + GZ (2.84 [0.95/3.89] vs 2.61 [-1.10/4.06] log10 IU/mL, P = 0.330] and CH + CI (3.57 [2.67/4.38] vs 3.63 [2.41/4.92] log10 IU/mL, P = 0.37). Infection phase (β: 0.422, P < 0.001), age (β: -0.260, P < 0.001), ALT (β: -0.103, P = 0.045), liver stiffness (β: -0.118, P = 0.039) and HBV-DNA (β: 0.384, P < 0.001), but not M-mut were independently associated with HBsAg serum levels. CONCLUSIONS In HBeAg negative, genotype-D, carriers Pre-S/S heterogeneity increases with severity of liver disease, but does not influence HBsAg serum levels, that in low viraemic carriers are associated with an effective control of HBV.
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Affiliation(s)
- Daniela Cavallone
- Department of Clinical and Experimental Medicine, University of Pisa, Pisa, Italy
- Hepatology Unit and Laboratory of Molecular Genetics and Pathology of Hepatitis Viruses, Pisa University Hospital, Pisa, Italy
| | - Gabriele Ricco
- Department of Clinical and Experimental Medicine, University of Pisa, Pisa, Italy
- Hepatology Unit and Laboratory of Molecular Genetics and Pathology of Hepatitis Viruses, Pisa University Hospital, Pisa, Italy
| | - Filippo Oliveri
- Hepatology Unit and Laboratory of Molecular Genetics and Pathology of Hepatitis Viruses, Pisa University Hospital, Pisa, Italy
| | - Piero Colombatto
- Hepatology Unit and Laboratory of Molecular Genetics and Pathology of Hepatitis Viruses, Pisa University Hospital, Pisa, Italy
| | - Francesco Moriconi
- Hepatology Unit and Laboratory of Molecular Genetics and Pathology of Hepatitis Viruses, Pisa University Hospital, Pisa, Italy
| | - Barbara Coco
- Hepatology Unit and Laboratory of Molecular Genetics and Pathology of Hepatitis Viruses, Pisa University Hospital, Pisa, Italy
| | - Veronica Romagnoli
- Hepatology Unit and Laboratory of Molecular Genetics and Pathology of Hepatitis Viruses, Pisa University Hospital, Pisa, Italy
| | - Antonio Salvati
- Hepatology Unit and Laboratory of Molecular Genetics and Pathology of Hepatitis Viruses, Pisa University Hospital, Pisa, Italy
| | - Lidia Surace
- Hepatology Unit and Laboratory of Molecular Genetics and Pathology of Hepatitis Viruses, Pisa University Hospital, Pisa, Italy
| | - Ferruccio Bonino
- Biostructure and Bio-imaging Institute of National Research Council of Italy, Naples, Italy
| | - Maurizia Rossana Brunetto
- Department of Clinical and Experimental Medicine, University of Pisa, Pisa, Italy
- Hepatology Unit and Laboratory of Molecular Genetics and Pathology of Hepatitis Viruses, Pisa University Hospital, Pisa, Italy
- Biostructure and Bio-imaging Institute of National Research Council of Italy, Naples, Italy
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22
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Gerasi M, Frakolaki E, Papadakis G, Chalari A, Lougiakis N, Marakos P, Pouli N, Vassilaki N. Design, synthesis and anti-HBV activity evaluation of new substituted imidazo[4,5-b]pyridines. Bioorg Chem 2020; 98:103580. [DOI: 10.1016/j.bioorg.2020.103580] [Citation(s) in RCA: 7] [Impact Index Per Article: 1.4] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/26/2019] [Revised: 12/18/2019] [Accepted: 01/10/2020] [Indexed: 02/08/2023]
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23
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Luo J, Xi J, Gao L, Hu J. Role of Hepatitis B virus capsid phosphorylation in nucleocapsid disassembly and covalently closed circular DNA formation. PLoS Pathog 2020; 16:e1008459. [PMID: 32226051 PMCID: PMC7145273 DOI: 10.1371/journal.ppat.1008459] [Citation(s) in RCA: 40] [Impact Index Per Article: 8.0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/26/2019] [Revised: 04/09/2020] [Accepted: 03/05/2020] [Indexed: 12/16/2022] Open
Abstract
Hepatitis B virus (HBV) delivers a partially double-stranded, relaxed circular (RC) DNA genome in complete virions to the host cell nucleus for conversion to the covalently closed circular (CCC) DNA, which establishes and sustains viral infection. An overlength pregenomic RNA (pgRNA) is then transcribed from CCC DNA and packaged into immature nucleocapsids (NCs) by the viral core (HBc) protein. pgRNA is reverse transcribed to produce RC DNA in mature NCs, which are then enveloped and secreted as complete virions, or delivered to the nucleus to replenish the nuclear CCC DNA pool. RC DNA, whether originating from extracellular virions or intracellular mature NCs, must be released upon NC disassembly (uncoating) for CCC DNA formation. HBc is known to undergo dynamic phosphorylation and dephosphorylation at its C-terminal domain (CTD) to facilitate pgRNA packaging and reverse transcription. Here, two putative phosphorylation sites in the HBc N-terminal domain (NTD), S44 and S49, were targeted for genetic and biochemical analysis to assess their potential roles in viral replication. The NTD mutant that mimics the non-phosphorylated state (N2A) was competent in all steps of viral replication tested from capsid assembly, pgRNA packaging, reverse transcription, to virion secretion, except for a decrease in CCC DNA formation. On the other hand, the phosphor-mimetic mutant N2E showed a defect in the early step of pgRNA packaging but enhanced the late step of mature NC uncoating and consequently, increased CCC DNA formation. N2E also enhanced phosphorylation in CTD and possibly elsewhere in HBc. Furthermore, inhibition of the cyclin-dependent kinase 2 (CDK2), which is packaged into viral capsids, could block CCC DNA formation. These results prompted us to propose a model whereby rephosphorylation of HBc at both NTD and CTD by the packaged CDK2, following CTD dephosphorylation during NC maturation, facilitates uncoating and CCC DNA formation by destabilizing mature NCs.
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Affiliation(s)
- Jun Luo
- Department of Microbiology and Immunology, The Pennsylvania State University College of Medicine, Hershey, Pennsylvania, United States of America
| | - Ji Xi
- Department of Microbiology and Immunology, The Pennsylvania State University College of Medicine, Hershey, Pennsylvania, United States of America
| | - Lu Gao
- Roche Pharma Research and Early Development, Roche Innovation Center Shanghai, Shanghai, China
| | - Jianming Hu
- Department of Microbiology and Immunology, The Pennsylvania State University College of Medicine, Hershey, Pennsylvania, United States of America
- * E-mail:
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24
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Abstract
With a yearly death toll of 880,000, hepatitis B virus (HBV) remains a major health problem worldwide, despite an effective prophylactic vaccine and well-tolerated, effective antivirals. HBV causes chronic hepatitis, fibrosis, cirrhosis, and hepatocellular carcinoma. The viral genome persists in infected hepatocytes even after long-term antiviral therapy, and its integration, though no longer able to support viral replication, destabilizes the host genome. HBV is a DNA virus that utilizes a virus-encoded reverse transcriptase to convert an RNA intermediate, termed pregenomic RNA, into the relaxed circular DNA genome, which is subsequently converted into a covalently closed circular DNA (cccDNA) in the host cell nucleus. cccDNA is maintained in the nucleus of the infected hepatocyte as a stable minichromosome and functions as the viral transcriptional template for the production of all viral gene products, and thus, it is the molecular basis of HBV persistence. The nuclear cccDNA pool can be replenished through recycling of newly synthesized, DNA-containing HBV capsids. Licensed antivirals target the HBV reverse transcriptase activity but fail to eliminate cccDNA, which would be required to cure HBV infection. Elimination of HBV cccDNA is so far only achieved by antiviral immune responses. Thus, this review will focus on possible curative strategies aimed at eliminating or crippling the viral cccDNA. Newer insights into the HBV life cycle and host immune response provide novel, potentially curative therapeutic opportunities and targets.
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25
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Lazarevic I, Banko A, Miljanovic D, Cupic M. Immune-Escape Hepatitis B Virus Mutations Associated with Viral Reactivation upon Immunosuppression. Viruses 2019; 11:v11090778. [PMID: 31450544 PMCID: PMC6784188 DOI: 10.3390/v11090778] [Citation(s) in RCA: 68] [Impact Index Per Article: 11.3] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/25/2019] [Revised: 08/21/2019] [Accepted: 08/22/2019] [Indexed: 02/07/2023] Open
Abstract
Hepatitis B virus (HBV) reactivation occurs as a major complication of immunosuppressive therapy among persons who have recovered from acute hepatitis and those who have controlled chronic infection. Recent literature data emphasize the presence of a high degree of S gene variability in HBV isolates from patients who developed reactivation. In reactivated HBV, the most frequently detected mutations belong to the second loop of “a” determinant in HBsAg. These mutations were identified to be immune escape and responsible for vaccine- and diagnostic-escape phenomena. Their emergence clearly provides survival in the presence of a developed humoral immune response and is often associated with impaired serological diagnosis of HBV reactivation. The knowledge of their existence and roles can elucidate the process of reactivation and strongly highlights the importance of HBV DNA detection in monitoring all patients with a history of HBV infection who are undergoing immunosuppression. This review discusses the possible influence of the most frequently found immune-escape mutations on HBV reactivation.
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Affiliation(s)
- Ivana Lazarevic
- Institute of Microbiology and Immunology, Faculty of Medicine, University of Belgrade, Dr Subotica 1, 11000 Belgrade, Serbia.
| | - Ana Banko
- Institute of Microbiology and Immunology, Faculty of Medicine, University of Belgrade, Dr Subotica 1, 11000 Belgrade, Serbia
| | - Danijela Miljanovic
- Institute of Microbiology and Immunology, Faculty of Medicine, University of Belgrade, Dr Subotica 1, 11000 Belgrade, Serbia
| | - Maja Cupic
- Institute of Microbiology and Immunology, Faculty of Medicine, University of Belgrade, Dr Subotica 1, 11000 Belgrade, Serbia
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26
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Ogura S, Tameda M, Sugimoto K, Ikejiri M, Usui M, Ito M, Takei Y. A substitution in the pre-S1 promoter region is associated with the viral regulation of hepatitis B virus. Virol J 2019; 16:59. [PMID: 31046787 PMCID: PMC6498540 DOI: 10.1186/s12985-019-1169-x] [Citation(s) in RCA: 3] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/24/2018] [Accepted: 04/23/2019] [Indexed: 12/19/2022] Open
Abstract
Background Much evidence has demonstrated the influence of Hepatitis B virus (HBV) mutations on the clinical course of HBV infection. As large (L) protein plays a crucial role for viral entry, we hypothesized that mutations in the pre-S1 promoter region might affect the expression of L protein and subsequently change the biological characters of virus. Methods Patients infected with genotype C HBV were enrolled for analysis. HBV DNA sequences were inserted into a TA cloning vector and analyzed. To evaluate the effects of mutations in the pre-S1 promoter region, promoter activity and the expression of mRNA and L protein were analyzed using HepG2 cells. Results In total, 35 patients were enrolled and 13 patients (37.1%) had a single base substitution in the pre-S1 promoter region; the most frequent substitution was a G-to-A substitution at the 2765th base (G2765A) in the Sp1 region. The HBV viral load showed a negative correlation with the substitution ratio of the Sp1 region or G2765A (r = − 0.493 and − 0.473, respectively). Among those with a viral load ≤5.0 log IU/ml, patients with the G2765A substitution showed a significantly lower HBV viral load than those with the wild-type sequence. HepG2 cells transfected with the G2765A substitution vector showed reduced luciferase activity of the pre-S1 promoter, as well as reduced expression of pre-S1 mRNA and L protein. Furthermore, the G2765A substitution greatly reduced the L protein expression level of vector-produced virus particles. Conclusion G2765A substitution in the pre-S1 promoter reduced the expression of L protein and resulted in a low viral load and less severe disease in chronic HBV infections.
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Affiliation(s)
- Suguru Ogura
- Department of Gastroenterology and Hepatology, Mie University Graduate School of Medicine, 2-174 Edobashi, Tsu, Mie, 514-8507, Japan
| | - Masahiko Tameda
- Department of Gastroenterology and Hepatology, Mie University Graduate School of Medicine, 2-174 Edobashi, Tsu, Mie, 514-8507, Japan
| | - Kazushi Sugimoto
- Department of Gastroenterology and Hepatology, Mie University Graduate School of Medicine, 2-174 Edobashi, Tsu, Mie, 514-8507, Japan. .,Department of Central Laboratory, Mie University Hospital, Tsu, Japan.
| | - Makoto Ikejiri
- Department of Central Laboratory, Mie University Hospital, Tsu, Japan
| | - Masanobu Usui
- Department of Hepatobiliary Pancreatic and Transplant Surgery, Mie University Graduate School of Medicine, Tsu, Japan
| | - Masaaki Ito
- Department of Cardiology and Nephrology, Mie University Graduate School of Medicine, Tsu, Japan
| | - Yoshiyuki Takei
- Department of Gastroenterology and Hepatology, Mie University Graduate School of Medicine, 2-174 Edobashi, Tsu, Mie, 514-8507, Japan
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Xia Y, Liang TJ. Development of Direct-acting Antiviral and Host-targeting Agents for Treatment of Hepatitis B Virus Infection. Gastroenterology 2019; 156:311-324. [PMID: 30243618 PMCID: PMC6340783 DOI: 10.1053/j.gastro.2018.07.057] [Citation(s) in RCA: 81] [Impact Index Per Article: 13.5] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 05/17/2018] [Revised: 07/13/2018] [Accepted: 07/23/2018] [Indexed: 02/06/2023]
Abstract
Hepatitis B virus (HBV) infection affects approximately 300 million people worldwide. Although antiviral therapies have improved the long-term outcomes, patients often require life-long treatment and there is no cure for HBV infection. New technologies can help us learn more about the pathogenesis of HBV infection and develop therapeutic agents to reduce its burden. We review recent advances in development of direct-acting antiviral and host-targeting agents, some of which have entered clinical trials. We also discuss strategies for unbiased high-throughput screens to identify compounds that inhibit HBV and for repurposing existing drugs.
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Affiliation(s)
- Yuchen Xia
- Liver Diseases Branch, National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, Bethesda, Maryland, 20892
| | - T Jake Liang
- Liver Diseases Branch, National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, Bethesda, Maryland, 20892.
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Effect of Plasma Protein Binding on the Anti-Hepatitis B Virus Activity and Pharmacokinetic Properties of NVR 3-778. Antimicrob Agents Chemother 2018; 62:AAC.01497-18. [PMID: 30181376 DOI: 10.1128/aac.01497-18] [Citation(s) in RCA: 3] [Impact Index Per Article: 0.4] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/20/2018] [Accepted: 08/29/2018] [Indexed: 02/07/2023] Open
Abstract
High plasma protein binding (PPB) levels not only affect drug-target engagement but can also impact exposure of hepatocytes to antivirals and thereby affect antiviral activity. In this study, we assessed the effect of PPB on the antiviral activity of NVR 3-778, a sulfamoylbenzamide capsid assembly modulator (CAM). To this end, primary human hepatocyte (PHH) medium was spiked with plasma proteins. First, the effect of plasma proteins on the hepatitis B virus (HBV) infection assay was evaluated. The addition of plasma proteins neither decreased cell viability nor affected HBV DNA secretion or intracellular HBV RNA accumulation. In contrast, the secretion and intracellular amount of HBV proteins were induced with increasing amounts of plasma proteins. Next, the antiviral activity of NVR 3-778 was demonstrated by multiple assays while PPB and the time-dependent disappearance of the parent drug were quantified by liquid chromatography-tandem mass spectrometry (LC-MS/MS). Plasma proteins strongly decreased the free fraction of NVR 3-778, resulting in a physiologically relevant in vitro hepatocyte exposure. NVR 3-778 displayed a high PPB level, while the antiviral activity was reduced approximately only 4-fold. The disconnect between the high PPB level and the only moderate shift of the antiviral activity was explained by the rapid hepatic clearance of NVR 3-778 in the absence of plasma proteins. This study highlights the use of PHHs as a model to accurately determine the antiviral activity by capturing PPB, clearance, and liver distribution. It is advantageous to consider both pharmacokinetics and pharmacodynamics for selection of HBV antiviral drug candidates and for successful extrapolation of in vitro data to clinical studies.
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29
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Weinman SA, Gudima SO. What roles HBV integrant-derived RNAs could play in the life cycles of HBV and HDV? Future Virol 2018. [DOI: 10.2217/fvl-2018-0130] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/21/2022]
Affiliation(s)
- Steven A Weinman
- Department of Internal Medicine, University of Kansas Medical Center, Kansas City, KS 66160, USA
- Department of Microbiology, Molecular Genetics & Immunology, University of Kansas Medical Center, Kansas City, KS 66160, USA
| | - Severin O Gudima
- Department of Microbiology, Molecular Genetics & Immunology, University of Kansas Medical Center, Kansas City, KS 66160, USA
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30
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Wu CC, Chen YS, Cao L, Chen XW, Lu MJ. Hepatitis B virus infection: Defective surface antigen expression and pathogenesis. World J Gastroenterol 2018; 24:3488-3499. [PMID: 30131655 PMCID: PMC6102499 DOI: 10.3748/wjg.v24.i31.3488] [Citation(s) in RCA: 27] [Impact Index Per Article: 3.9] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 05/03/2018] [Revised: 06/01/2018] [Accepted: 06/25/2018] [Indexed: 02/06/2023] Open
Abstract
Hepatitis B virus (HBV) infection is a global public health concern. HBV causes chronic infection in patients and can lead to liver cirrhosis, hepatocellular carcinoma, and other severe liver diseases. Thus, understanding HBV-related pathogenesis is of particular importance for prevention and clinical intervention. HBV surface antigens are indispensable for HBV virion formation and are useful viral markers for diagnosis and clinical assessment. During chronic HBV infection, HBV genomes may acquire and accumulate mutations and deletions, leading to the expression of defective HBV surface antigens. These defective HBV surface antigens have been found to play important roles in the progression of HBV-associated liver diseases. In this review, we focus our discussion on the nature of defective HBV surface antigen mutations and their contribution to the pathogenesis of fulminant hepatitis B. The relationship between defective surface antigens and occult HBV infection are also discussed.
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MESH Headings
- DNA, Viral/genetics
- DNA, Viral/isolation & purification
- Disease Progression
- Genome, Viral/genetics
- Hepatitis B Surface Antigens/genetics
- Hepatitis B Surface Antigens/immunology
- Hepatitis B Surface Antigens/metabolism
- Hepatitis B virus/genetics
- Hepatitis B virus/immunology
- Hepatitis B, Chronic/immunology
- Hepatitis B, Chronic/pathology
- Hepatitis B, Chronic/prevention & control
- Hepatitis B, Chronic/virology
- Humans
- Liver/immunology
- Liver/pathology
- Liver/virology
- Liver Failure, Acute/immunology
- Liver Failure, Acute/pathology
- Liver Failure, Acute/prevention & control
- Liver Failure, Acute/virology
- Mutation
- Virus Replication/genetics
- Virus Replication/immunology
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Affiliation(s)
- Chun-Chen Wu
- State Key Laboratory of Virology, Wuhan Institute of Virology, Chinese Academy of Sciences, Wuhan 430071, Hubei Province, China
| | - Ying-Shan Chen
- State Key Laboratory of Virology, Wuhan Institute of Virology, Chinese Academy of Sciences, Wuhan 430071, Hubei Province, China
| | - Liang Cao
- State Key Laboratory of Virology, Wuhan Institute of Virology, Chinese Academy of Sciences, Wuhan 430071, Hubei Province, China
- Department of Microbiology and Immunology, Feinberg School of Medicine Northwestern University, Chicago, IL 60611, United States
| | - Xin-Wen Chen
- State Key Laboratory of Virology, Wuhan Institute of Virology, Chinese Academy of Sciences, Wuhan 430071, Hubei Province, China
| | - Meng-Ji Lu
- Institute of Virology, University Hospital of Essen, Essen 45122, Germany
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31
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Karayiannis P. Hepatitis B virus: virology, molecular biology, life cycle and intrahepatic spread. Hepatol Int 2017; 11:500-508. [PMID: 29098564 DOI: 10.1007/s12072-017-9829-7] [Citation(s) in RCA: 84] [Impact Index Per Article: 10.5] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 07/11/2017] [Accepted: 10/05/2017] [Indexed: 12/18/2022]
Abstract
Hepatitis B virus is a member of the Hepadnaviridae family and responsible for causing acute and chronic hepatitis in humans. The current estimates of people chronically infected with the virus are put at 250 million worldwide. Immune-mediated liver damage in these individuals may lead to the development of cirrhosis and hepatocellular carcinoma later in life. This review deals with our current understanding of the virology, molecular biology, life cycle and cell-to-cell spread of this very important pathogen, all of which are considered essential for current and future approaches to antiviral treatment.
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Affiliation(s)
- P Karayiannis
- Medical School, University of Nicosia, 21 Ilia Papakyriakou, 2414 Engomi, P.O. Box 24005, CY-1700, Nicosia, Cyprus.
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32
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Chen J, Xu CR, Xi M, Hu WW, Tang ZH, Zang GQ. Predictors of liver histological changes and a sustained virological response to peginterferon among chronic hepatitis B e antigen-positive patients with normal or minimally elevated alanine aminotransferase levels. J Viral Hepat 2017; 24:573-579. [PMID: 28107601 DOI: 10.1111/jvh.12679] [Citation(s) in RCA: 6] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 11/03/2016] [Accepted: 12/30/2016] [Indexed: 12/25/2022]
Abstract
A proportion of chronic hepatitis B patients with normal or only minimally elevated alanine aminotransferase (ALT) levels display significant histologic changes and would benefit from antiviral therapy. We aim to evaluate the histologic abnormalities seen in these patients and then determine which of them would most likely respond to peginterferon therapy. One hundred and thirteen hepatitis B e antigen (HBeAg)-positive patients with a normal or minimally elevated ALT level and moderate-to-severe histologic changes in their liver tissue were selected to receive peginterferon monotherapy and participate in a follow-up analysis. A multiple logistic regression analysis indicated that increasing age (P=.049) and lower hepatitis B virus (HBV) DNA levels (P=.038) were associated with significant histological abnormalities in patients with a normal or minimally elevated ALT. Our predictive model which incorporated HBeAg testing at treatment week 12 combined with hepatitis B surface antigen (HBsAg) testing at treatment week 24 was able to identify which patients with a normal ALT level would achieve a sustained virological response (SVR) (positive predictive value [PPV]: 66.7%, negative predictive value [NPV]: 90.0%). Lower HBsAg and HBeAg levels at treatment week 24 were associated with a SVR in patients with a minimally elevated ALT level (PPV: 100.0%, NPV: 100.0%). A liver biopsy and antiviral therapy should be strongly considered when treating HBeAg-positive patients with a normal or minimally elevated ALT level, low HBV DNA level, and aged >35 years. On-treatment quantification of combined HBsAg and HBeAg test results may be useful for predicting a SVR to peginterferon monotherapy in these patients.
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Affiliation(s)
- J Chen
- Department of Infectious Disease, The Sixth People's Hospital affiliated with Shanghai Jiao Tong University, Shanghai, China
| | - C R Xu
- Department of Infectious Disease, Southeast Hospital affiliated with Xiamen University, Zhangzhou, China
| | - M Xi
- Department of Infectious Disease, The Sixth People's Hospital affiliated with Shanghai Jiao Tong University, Shanghai, China
| | - W W Hu
- Department of Infectious Disease, The Sixth People's Hospital affiliated with Shanghai Jiao Tong University, Shanghai, China
| | - Z H Tang
- Department of Infectious Disease, The Sixth People's Hospital affiliated with Shanghai Jiao Tong University, Shanghai, China
| | - G Q Zang
- Department of Infectious Disease, The Sixth People's Hospital affiliated with Shanghai Jiao Tong University, Shanghai, China
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33
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Schreiner S, Nassal M. A Role for the Host DNA Damage Response in Hepatitis B Virus cccDNA Formation-and Beyond? Viruses 2017; 9:v9050125. [PMID: 28531167 PMCID: PMC5454437 DOI: 10.3390/v9050125] [Citation(s) in RCA: 69] [Impact Index Per Article: 8.6] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/10/2017] [Revised: 05/16/2017] [Accepted: 05/18/2017] [Indexed: 12/12/2022] Open
Abstract
Chronic hepatitis B virus (HBV) infection puts more than 250 million people at a greatly increased risk to develop end-stage liver disease. Like all hepadnaviruses, HBV replicates via protein-primed reverse transcription of a pregenomic (pg) RNA, yielding an unusually structured, viral polymerase-linked relaxed-circular (RC) DNA as genome in infectious particles. Upon infection, RC-DNA is converted into nuclear covalently closed circular (ccc) DNA. Associating with cellular proteins into an episomal minichromosome, cccDNA acts as template for new viral RNAs, ensuring formation of progeny virions. Hence, cccDNA represents the viral persistence reservoir that is not directly targeted by current anti-HBV therapeutics. Eliminating cccDNA will thus be at the heart of a cure for chronic hepatitis B. The low production of HBV cccDNA in most experimental models and the associated problems in reliable cccDNA quantitation have long hampered a deeper understanding of cccDNA molecular biology. Recent advancements including cccDNA-dependent cell culture systems have begun to identify select host DNA repair enzymes that HBV usurps for RC-DNA to cccDNA conversion. While this list is bound to grow, it may represent just one facet of a broader interaction with the cellular DNA damage response (DDR), a network of pathways that sense and repair aberrant DNA structures and in the process profoundly affect the cell cycle, up to inducing cell death if repair fails. Given the divergent interactions between other viruses and the DDR it will be intriguing to see how HBV copes with this multipronged host system.
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Affiliation(s)
- Sabrina Schreiner
- Institute of Virology, Technische Universität München/Helmholtz Zentrum München, Ingolstädter Landstr. 1, Neuherberg, D-85764 Munich, Germany.
| | - Michael Nassal
- Dept. of Internal Medicine II/Molecular Biology, University Hospital Freiburg, Hugstetter Str. 55, D-79106 Freiburg, Germany.
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34
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Liu W, Bai M, Chen F, Zhu H, Zheng M, Zhou C, Yang Y, Hu Y, Wang J, Chen Z. CMV promoter and the mouse albumin promoter and enhancer triggered the elevation of hepatitis B virus envelope polypeptides. Acta Biochim Biophys Sin (Shanghai) 2017; 49:374-377. [PMID: 28338773 DOI: 10.1093/abbs/gmx007] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/26/2016] [Indexed: 11/14/2022] Open
Affiliation(s)
- Weixia Liu
- State Key Laboratory for Diagnosis and Treatment of Infectious Diseases, Collaborative Innovation Center for Diagnosis and Treatment of Infectious Diseases, The First Affiliated Hospital, College of Medicine, Zhejiang University, Hangzhou 310003, China
| | - Minghui Bai
- State Key Laboratory for Diagnosis and Treatment of Infectious Diseases, Collaborative Innovation Center for Diagnosis and Treatment of Infectious Diseases, The First Affiliated Hospital, College of Medicine, Zhejiang University, Hangzhou 310003, China
- Hangzhou Xixi Hospital, Hangzhou 310023, China
| | - Feng Chen
- State Key Laboratory for Diagnosis and Treatment of Infectious Diseases, Collaborative Innovation Center for Diagnosis and Treatment of Infectious Diseases, The First Affiliated Hospital, College of Medicine, Zhejiang University, Hangzhou 310003, China
| | - Haihong Zhu
- State Key Laboratory for Diagnosis and Treatment of Infectious Diseases, Collaborative Innovation Center for Diagnosis and Treatment of Infectious Diseases, The First Affiliated Hospital, College of Medicine, Zhejiang University, Hangzhou 310003, China
| | - Min Zheng
- State Key Laboratory for Diagnosis and Treatment of Infectious Diseases, Collaborative Innovation Center for Diagnosis and Treatment of Infectious Diseases, The First Affiliated Hospital, College of Medicine, Zhejiang University, Hangzhou 310003, China
| | - Cheng Zhou
- State Key Laboratory for Diagnosis and Treatment of Infectious Diseases, Collaborative Innovation Center for Diagnosis and Treatment of Infectious Diseases, The First Affiliated Hospital, College of Medicine, Zhejiang University, Hangzhou 310003, China
| | - Ying Yang
- State Key Laboratory for Diagnosis and Treatment of Infectious Diseases, Collaborative Innovation Center for Diagnosis and Treatment of Infectious Diseases, The First Affiliated Hospital, College of Medicine, Zhejiang University, Hangzhou 310003, China
| | - Ying Hu
- State Key Laboratory for Diagnosis and Treatment of Infectious Diseases, Collaborative Innovation Center for Diagnosis and Treatment of Infectious Diseases, The First Affiliated Hospital, College of Medicine, Zhejiang University, Hangzhou 310003, China
| | - Jing Wang
- State Key Laboratory for Diagnosis and Treatment of Infectious Diseases, Collaborative Innovation Center for Diagnosis and Treatment of Infectious Diseases, The First Affiliated Hospital, College of Medicine, Zhejiang University, Hangzhou 310003, China
| | - Zhi Chen
- State Key Laboratory for Diagnosis and Treatment of Infectious Diseases, Collaborative Innovation Center for Diagnosis and Treatment of Infectious Diseases, The First Affiliated Hospital, College of Medicine, Zhejiang University, Hangzhou 310003, China
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35
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Efficient genome replication of hepatitis B virus using adenovirus vector: a compact pregenomic RNA-expression unit. Sci Rep 2017; 7:41851. [PMID: 28157182 PMCID: PMC5291108 DOI: 10.1038/srep41851] [Citation(s) in RCA: 7] [Impact Index Per Article: 0.9] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/03/2016] [Accepted: 01/04/2017] [Indexed: 01/05/2023] Open
Abstract
The complicated replication mechanisms of hepatitis B virus (HBV) have impeded HBV studies and anti-HBV therapy development as well. Herein we report efficient genome replication of HBV applying adenovirus vectors (AdVs) showing high transduction efficiency. Even in primary hepatocytes derived from humanized mice the transduction efficiencies using AdVs were 450-fold higher compared than those using plasmids. By using an expression unit consisting of the CMV promoter, 1.03-copy HBV genome and foreign poly(A) signal, we successfully generated an improved AdV (HBV103-AdV) that efficiently provided 58 times more pregenomic RNA than previously reported AdVs. The HBV103-AdV-mediated HBV replication was easily and precisely detected using quantitative real-time PCR in primary hepatocytes as well as in HepG2 cells. Notably, when the AdV containing replication-defective HBV genome of 1.14 copy was transduced, we observed that HBV DNA-containing circular molecules (pseudo-ccc DNA) were produced, which were probably generated through homologous recombination. However, the replication-defective HBV103-AdV hardly yielded the pseudo-ccc, probably because the repeated sequences are vey short. Additionally, the efficacies of entecavir and lamivudine were quantitatively evaluated using this system at only 4 days postinfection with HBV103-AdVs. Therefore, this system offers high production of HBV genome replication and thus could become used widely.
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36
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Locatelli M, Testoni B. [Analysis of intrahepatic virological events associated to chronic hepatitis B infection]. Med Sci (Paris) 2017; 33:92-95. [PMID: 28120764 DOI: 10.1051/medsci/20173301017] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/14/2022] Open
Affiliation(s)
- Maëlle Locatelli
- Inserm U1052, 151, cours Albert Thomas, 69003 Lyon, France - Centre de recherche en cancérologie de Lyon (CRCL), 151, cours Albert Thomas, 69003 Lyon, France - Université de Lyon, UMR-S1052, UCBL, 151, cours Albert Thomas, 69003 Lyon, France
| | - Barbara Testoni
- Inserm U1052, 151, cours Albert Thomas, 69003 Lyon, France - Centre de recherche en cancérologie de Lyon (CRCL), 151, cours Albert Thomas, 69003 Lyon, France - Université de Lyon, UMR-S1052, UCBL, 151, cours Albert Thomas, 69003 Lyon, France
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37
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Zhou LY, Chen EQ, Wang ML, Chen LL, Liu CP, Zeng F, Tang H. Biological characteristics comparison of HBV rtA181T mutants with truncated or substituted HBsAg expression in vitro and in vivo model systems. Sci Rep 2016; 6:39260. [PMID: 27976732 PMCID: PMC5157016 DOI: 10.1038/srep39260] [Citation(s) in RCA: 6] [Impact Index Per Article: 0.7] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/16/2016] [Accepted: 11/22/2016] [Indexed: 02/05/2023] Open
Abstract
The hepatitis B virus(HBV) polymerase rtA181T mutation is selected during long-term antiviral therapy. As the polymerase gene completely overlaps with the envelope (S) gene, HBV rtA181T mutation also carries sW172 mutations. In this study, we investigated whether there were biological differences between rtA181T/sW172* (coding truncated HBsAg) and rtA181T/sW172L (coding substituted HBsAg) mutants. In cell experiments, a slight decline of viral replication was observed in both two mutants as compared to wild-type strains, but the levels of supernatant HBsAg and HBV DNA in rtA181T/sW172* were significantly lower than those in rtA181T/sW172L transfected cells. In animal experiments, we were amazed to find that viral replication in rtA181T/sW172* mutant increased and maintained significantly longer than that in rtA181T/sW172L mutant, while no significant difference was observed between rtA181T/sW172L and wild-type strains. Compared with wild-type strains, there were intracellular accumulations of HBsAg and HBcAg in rtA181/sW172* but none in rtA181/sW172L mutant strains. Importantly, we also found that truncated HBsAg could increase the activity of HBV core promoter, but substituted HBsAg could not. In summary, the characteristics of above two rtA181T mutants mentioned above were significantly different, and it is necessary and important for us to distinguish sW172* truncated mutation from sW172L substituted mutation.
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Affiliation(s)
- Ling-Yun Zhou
- Center of Infectious Diseases, West China Hospital, Sichuan University, Chengdu 610041, China
- Division of Infectious Diseases, State Key Laboratory of Biotherapy and Cancer Center, West China Hospital, Sichuan University, Chengdu 610041, China
| | - En-Qiang Chen
- Center of Infectious Diseases, West China Hospital, Sichuan University, Chengdu 610041, China
- Division of Infectious Diseases, State Key Laboratory of Biotherapy and Cancer Center, West China Hospital, Sichuan University, Chengdu 610041, China
| | - Meng-Lan Wang
- Center of Infectious Diseases, West China Hospital, Sichuan University, Chengdu 610041, China
- Division of Infectious Diseases, State Key Laboratory of Biotherapy and Cancer Center, West China Hospital, Sichuan University, Chengdu 610041, China
| | - Lan-Lan Chen
- Center of Infectious Diseases, West China Hospital, Sichuan University, Chengdu 610041, China
- Division of Infectious Diseases, State Key Laboratory of Biotherapy and Cancer Center, West China Hospital, Sichuan University, Chengdu 610041, China
| | - Cui-Ping Liu
- Center of Infectious Diseases, West China Hospital, Sichuan University, Chengdu 610041, China
- Division of Infectious Diseases, State Key Laboratory of Biotherapy and Cancer Center, West China Hospital, Sichuan University, Chengdu 610041, China
| | - Fan Zeng
- Center of Infectious Diseases, West China Hospital, Sichuan University, Chengdu 610041, China
- Division of Infectious Diseases, State Key Laboratory of Biotherapy and Cancer Center, West China Hospital, Sichuan University, Chengdu 610041, China
| | - Hong Tang
- Center of Infectious Diseases, West China Hospital, Sichuan University, Chengdu 610041, China
- Division of Infectious Diseases, State Key Laboratory of Biotherapy and Cancer Center, West China Hospital, Sichuan University, Chengdu 610041, China
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38
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Qi Y, Gao Z, Xu G, Peng B, Liu C, Yan H, Yao Q, Sun G, Liu Y, Tang D, Song Z, He W, Sun Y, Guo JT, Li W. DNA Polymerase κ Is a Key Cellular Factor for the Formation of Covalently Closed Circular DNA of Hepatitis B Virus. PLoS Pathog 2016; 12:e1005893. [PMID: 27783675 PMCID: PMC5081172 DOI: 10.1371/journal.ppat.1005893] [Citation(s) in RCA: 156] [Impact Index Per Article: 17.3] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/23/2016] [Accepted: 08/24/2016] [Indexed: 12/11/2022] Open
Abstract
Hepatitis B virus (HBV) infection of hepatocytes begins by binding to its cellular receptor sodium taurocholate cotransporting polypeptide (NTCP), followed by the internalization of viral nucleocapsid into the cytoplasm. The viral relaxed circular (rc) DNA genome in nucleocapsid is transported into the nucleus and converted into covalently closed circular (ccc) DNA to serve as a viral persistence reservoir that is refractory to current antiviral therapies. Host DNA repair enzymes have been speculated to catalyze the conversion of rcDNA to cccDNA, however, the DNA polymerase(s) that fills the gap in the plus strand of rcDNA remains to be determined. Here we conducted targeted genetic screening in combination with chemical inhibition to identify the cellular DNA polymerase(s) responsible for cccDNA formation, and exploited recombinant HBV with capsid coding deficiency which infects HepG2-NTCP cells with similar efficiency of wild-type HBV to assure cccDNA synthesis is exclusively from de novo HBV infection. We found that DNA polymerase κ (POLK), a Y-family DNA polymerase with maximum activity in non-dividing cells, substantially contributes to cccDNA formation during de novo HBV infection. Depleting gene expression of POLK in HepG2-NTCP cells by either siRNA knockdown or CRISPR/Cas9 knockout inhibited the conversion of rcDNA into cccDNA, while the diminished cccDNA formation in, and hence the viral infection of, the knockout cells could be effectively rescued by ectopic expression of POLK. These studies revealed that POLK is a crucial host factor required for cccDNA formation during a de novo HBV infection and suggest that POLK may be a potential target for developing antivirals against HBV. HBV chronically infects 240 million people worldwide. Persistent HBV infection relies on stable maintenance of the nuclear form of viral genome, the covalently closed circular (ccc) DNA. However, the molecular mechanism underlying the conversion of HBV genomic relaxed circular (rc) DNA into cccDNA remains elusive. Our studies reported herein provide unambiguous evidence suggesting that host DNA polymerase κ (POLK) is required for repairing the gap of rcDNA and formation of cccDNA in a de novo HBV infection. POLK is thus a potential therapeutic target for treatment of chronic hepatitis B.
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Affiliation(s)
- Yonghe Qi
- National Institute of Biological Sciences, Beijing, China
| | - Zhenchao Gao
- National Institute of Biological Sciences, Beijing, China
- Graduate program in School of Life Sciences, Peking University, Beijing, China
| | - Guangwei Xu
- National Institute of Biological Sciences, Beijing, China
| | - Bo Peng
- National Institute of Biological Sciences, Beijing, China
- Graduate program in School of Life Sciences, Peking University, Beijing, China
| | - Chenxuan Liu
- National Institute of Biological Sciences, Beijing, China
- College of Life Sciences Beijing Normal University, Beijing, China
| | - Huan Yan
- National Institute of Biological Sciences, Beijing, China
| | - Qiyan Yao
- National Institute of Biological Sciences, Beijing, China
| | - Guoliang Sun
- National Institute of Biological Sciences, Beijing, China
| | - Yang Liu
- National Institute of Biological Sciences, Beijing, China
- School of Life Science, Tsinghua University, Beijing, China
| | - Dingbin Tang
- National Institute of Biological Sciences, Beijing, China
- Graduate program in School of Life Sciences, Peking University, Beijing, China
| | - Zilin Song
- National Institute of Biological Sciences, Beijing, China
| | - Wenhui He
- National Institute of Biological Sciences, Beijing, China
| | - Yinyan Sun
- National Institute of Biological Sciences, Beijing, China
| | - Ju-Tao Guo
- Baruch S. Blumberg Institute, Doylestown, Pennsylvania, United States of America
| | - Wenhui Li
- National Institute of Biological Sciences, Beijing, China
- * E-mail: (WL)
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Liu C, Cai D, Zhang L, Tang W, Yan R, Guo H, Chen X. Identification of hydrolyzable tannins (punicalagin, punicalin and geraniin) as novel inhibitors of hepatitis B virus covalently closed circular DNA. Antiviral Res 2016; 134:97-107. [PMID: 27591143 DOI: 10.1016/j.antiviral.2016.08.026] [Citation(s) in RCA: 59] [Impact Index Per Article: 6.6] [Reference Citation Analysis] [Abstract] [Key Words] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/26/2016] [Revised: 08/24/2016] [Accepted: 08/29/2016] [Indexed: 02/07/2023]
Abstract
The development of new agents to target HBV cccDNA is urgently needed because of the limitations of current available drugs for treatment of hepatitis B. By using a cell-based assay in which the production of HBeAg is in a cccDNA-dependent manner, we screened a compound library derived from Chinese herbal remedies for inhibitors against HBV cccDNA. Three hydrolyzable tannins, specifically punicalagin, punicalin and geraniin, emerged as novel anti-HBV agents. These compounds significantly reduced the production of secreted HBeAg and cccDNA in a dose-dependent manner in our assay, without dramatic alteration of viral DNA replication. Furthermore, punicalagin did not affect precore/core promoter activity, pgRNA transcription, core protein expression, or HBsAg secretion. By employing the cell-based cccDNA accumulation and stability assay, we found that these tannins significantly inhibited the establishment of cccDNA and modestly facilitated the degradation of preexisting cccDNA. Collectively, our results suggest that hydrolyzable tannins inhibit HBV cccDNA production via a dual mechanism through preventing the formation of cccDNA and promoting cccDNA decay, although the latter effect is rather minor. These hydrolyzable tannins may serve as lead compounds for the development of new agents to cure HBV infection.
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Affiliation(s)
- Chunlan Liu
- State Key Laboratory of Virology, Wuhan Institute of Virology, Chinese Academy of Sciences, Wuhan 43001, Hubei, China
| | - Dawei Cai
- Department of Microbiology and Immunology, Indiana University School of Medicine, 635 Barnhill Drive, Indianapolis, IN 46202, USA
| | - Lin Zhang
- State Key Laboratory of Virology, Wuhan Institute of Virology, Chinese Academy of Sciences, Wuhan 43001, Hubei, China
| | - Wei Tang
- State Key Laboratory of Virology, Wuhan Institute of Virology, Chinese Academy of Sciences, Wuhan 43001, Hubei, China
| | - Ran Yan
- Department of Microbiology and Immunology, Indiana University School of Medicine, 635 Barnhill Drive, Indianapolis, IN 46202, USA
| | - Haitao Guo
- Department of Microbiology and Immunology, Indiana University School of Medicine, 635 Barnhill Drive, Indianapolis, IN 46202, USA.
| | - Xulin Chen
- State Key Laboratory of Virology, Wuhan Institute of Virology, Chinese Academy of Sciences, Wuhan 43001, Hubei, China.
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Hepatitis B Virus Middle Protein Enhances IL-6 Production via p38 MAPK/NF-κB Pathways in an ER Stress-Dependent Manner. PLoS One 2016; 11:e0159089. [PMID: 27434097 PMCID: PMC4951109 DOI: 10.1371/journal.pone.0159089] [Citation(s) in RCA: 16] [Impact Index Per Article: 1.8] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/01/2016] [Accepted: 06/27/2016] [Indexed: 02/07/2023] Open
Abstract
During hepatitis B virus (HBV) infection, three viral envelope proteins of HBV are overexpressed in the endoplasmic reticulum (ER). The large S protein (LHBs) and truncated middle S protein (MHBst) have been documented to play roles in regulating host gene expression and contribute to hepatic disease development. As a predominant protein at the ultrastructural level in biopsy samples taken from viremic patients, the role of the middle S protein (MHBs) remains to be understood despite its high immunogenicity. When we transfected hepatocytes with an enhanced green fluorescent protein (EGFP)-tagged MHBs expressing plasmid, the results showed that expression of MHBs cause an upregulation of IL-6 at the message RNA and protein levels through activating the p38 mitogen-activated protein kinase (p38 MAPK) and nuclear factor-kappa B (NF-κB) pathways. The use of specific inhibitors of the signaling pathways can diminish this upregulation. The use of BAPTA-AM attenuated the stimulation caused by MHBs. We further found that MHBs accumulated in the endoplasmic reticulum and increased the amount of glucose regulated protein 78 (GRP78/BiP). Our results provide a possibility that MHBs could be involved in liver disease progression.
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41
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Levrero M, Testoni B, Zoulim F. HBV cure: why, how, when? Curr Opin Virol 2016; 18:135-43. [PMID: 27447092 DOI: 10.1016/j.coviro.2016.06.003] [Citation(s) in RCA: 48] [Impact Index Per Article: 5.3] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/10/2016] [Revised: 06/06/2016] [Accepted: 06/06/2016] [Indexed: 12/17/2022]
Abstract
Current HBV treatments control replication and liver disease progression in the vast majority of treated patients. However, HBV patients often require lifelong therapies due to the persistence of transcriptionally active viral cccDNA mini-chromosome in the nucleus, which is not directly targeted by current antiviral therapies. A true complete cure of HBV would require clearance of intranuclear cccDNA from all infected hepatocytes. An intermediate but still relevant step forward that would allow treatment cessation would be reaching a functional cure, equivalent to resolved acute infection, with a durable HBsAg loss±anti-HBs seroconversion, undetectable serum DNA and persistence of cccDNA in a transcriptionally inactive status. Recent advances in technologies and pharmaceutical sciences, including the cloning of the mayor HBV receptor (i.e. the NTCP transporter) and the development in vitro HBV infection models, have heralded a new horizon of innovative antiviral and immune-therapeutic approaches.
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Affiliation(s)
- Massimo Levrero
- Cancer Research Center of Lyon (CRCL), Lyon 69008, France; INSERM, U1052, Lyon 69003, France; Hospices Civils de Lyon (HCL), 69002 Lyon, France; Department of Internal Medicine - DMISM, Sapienza University, 00161 Rome, Italy; CLNS@SAPIENZA, Istituto Italiano di Tecnologia (IIT), 00161 Rome, Italy.
| | - Barbara Testoni
- Cancer Research Center of Lyon (CRCL), Lyon 69008, France; INSERM, U1052, Lyon 69003, France
| | - Fabien Zoulim
- Cancer Research Center of Lyon (CRCL), Lyon 69008, France; INSERM, U1052, Lyon 69003, France; Hospices Civils de Lyon (HCL), 69002 Lyon, France; University of Lyon, UMR_S1052, UCBL, 69008 Lyon, France; Institut Universitaire de France (IUF), 75005 Paris, France
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42
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Lamontagne RJ, Bagga S, Bouchard MJ. Hepatitis B virus molecular biology and pathogenesis. HEPATOMA RESEARCH 2016; 2:163-186. [PMID: 28042609 PMCID: PMC5198785 DOI: 10.20517/2394-5079.2016.05] [Citation(s) in RCA: 115] [Impact Index Per Article: 12.8] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 12/11/2022]
Abstract
As obligate intracellular parasites, viruses need a host cell to provide a milieu favorable to viral replication. Consequently, viruses often adopt mechanisms to subvert host cellular signaling processes. While beneficial for the viral replication cycle, virus-induced deregulation of host cellular signaling processes can be detrimental to host cell physiology and can lead to virus-associated pathogenesis, including, for oncogenic viruses, cell transformation and cancer progression. Included among these oncogenic viruses is the hepatitis B virus (HBV). Despite the availability of an HBV vaccine, 350-500 million people worldwide are chronically infected with HBV, and a significant number of these chronically infected individuals will develop hepatocellular carcinoma (HCC). Epidemiological studies indicate that chronic infection with HBV is the leading risk factor for the development of HCC. Globally, HCC is the second highest cause of cancer-associated deaths, underscoring the need for understanding mechanisms that regulate HBV replication and the development of HBV-associated HCC. HBV is the prototype member of the Hepadnaviridae family; members of this family of viruses have a narrow host range and predominately infect hepatocytes in their respective hosts. The extremely small and compact hepadnaviral genome, the unique arrangement of open reading frames, and a replication strategy utilizing reverse transcription of an RNA intermediate to generate the DNA genome are distinguishing features of the Hepadnaviridae. In this review, we provide a comprehensive description of HBV biology, summarize the model systems used for studying HBV infections, and highlight potential mechanisms that link a chronic HBV-infection to the development of HCC. For example, the HBV X protein (HBx), a key regulatory HBV protein that is important for HBV replication, is thought to play a cofactor role in the development of HBV-induced HCC, and we highlight the functions of HBx that may contribute to the development of HBV-associated HCC.
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Affiliation(s)
- R. Jason Lamontagne
- Department of Biochemistry and Molecular Biology, Drexel University College of Medicine, Philadelphia, PA 19102, USA
- The Wistar Institute, Philadelphia, PA 19104, USA
| | - Sumedha Bagga
- Department of Biochemistry and Molecular Biology, Drexel University College of Medicine, Philadelphia, PA 19102, USA
| | - Michael J. Bouchard
- Department of Biochemistry and Molecular Biology, Drexel University College of Medicine, Philadelphia, PA 19102, USA
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Cai D, Wang X, Yan R, Mao R, Liu Y, Ji C, Cuconati A, Guo H. Establishment of an inducible HBV stable cell line that expresses cccDNA-dependent epitope-tagged HBeAg for screening of cccDNA modulators. Antiviral Res 2016; 132:26-37. [PMID: 27185623 DOI: 10.1016/j.antiviral.2016.05.005] [Citation(s) in RCA: 47] [Impact Index Per Article: 5.2] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/26/2016] [Revised: 04/26/2016] [Accepted: 05/11/2016] [Indexed: 02/07/2023]
Abstract
Hepatitis B virus (HBV) covalently closed circular (ccc) DNA is essential to the virus life cycle, its elimination during chronic infection is considered critical to a durable therapy but has not been achieved by current antivirals. Despite being essential, cccDNA has not been the major target of high throughput screening (HTS), largely because of the limitations of current HBV tissue culture systems, including the impracticality of detecting cccDNA itself. In response to this need, we have previously developed a proof-of-concept HepDE19 cell line in which the production of wildtype e antigen (HBeAg) is dependent upon cccDNA. However, the existing assay system is not ideal for HTS because the HBeAg ELISA cross reacts with a viral HBeAg homologue, which is the core antigen (HBcAg) expressed largely in a cccDNA-independent fashion in HepDE19 cells. To further improve the assay specificity, we report herein a "second-generation" cccDNA reporter cell line, termed HepBHAe82. In the similar principle of HepDE19 line, an in-frame HA epitope tag was introduced into the precore domain of HBeAg open reading frame in the transgene of HepBHAe82 cells without disrupting any cis-element critical for HBV replication and HBeAg secretion. A chemiluminescence ELISA assay (CLIA) for the detection of HA-tagged HBeAg with HA antibody serving as capture antibody and HBeAb serving as detection antibody has been developed to eliminate the confounding signal from HBcAg. The miniaturized HepBHAe82 cell based assay system exhibits high level of cccDNA-dependent HA-HBeAg production and high specific readout signals with low background. We have also established a HepHA-HBe4 cell line expressing transgene-dependent HA-HBeAg as a counter screen to identify HBeAg inhibitors. The HepBHAe82 system is amenable to antiviral HTS development, and can be used to identify host factors that regulate cccDNA metabolism and transcription.
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Affiliation(s)
- Dawei Cai
- Institute for Biotechnology and Virology Research, Drexel University College of Medicine, Doylestown, PA, 18902, USA; Department of Microbiology and Immunology, Indiana University School of Medicine, Indianapolis, IN, 46202, USA
| | - Xiaohe Wang
- Baruch S. Blumberg Institute, Hepatitis B Foundation, Doylestown, PA, 18902, USA
| | - Ran Yan
- Institute for Biotechnology and Virology Research, Drexel University College of Medicine, Doylestown, PA, 18902, USA; Department of Microbiology and Immunology, Indiana University School of Medicine, Indianapolis, IN, 46202, USA
| | - Richeng Mao
- Institute for Biotechnology and Virology Research, Drexel University College of Medicine, Doylestown, PA, 18902, USA; Department of Microbiology and Immunology, Indiana University School of Medicine, Indianapolis, IN, 46202, USA
| | - Yuanjie Liu
- Department of Microbiology and Immunology, Indiana University School of Medicine, Indianapolis, IN, 46202, USA
| | - Changhua Ji
- Virology Discovery and Translational Area, Roche Pharma Research and Early Development, Nutley, NJ, 07110, USA.
| | - Andrea Cuconati
- Baruch S. Blumberg Institute, Hepatitis B Foundation, Doylestown, PA, 18902, USA.
| | - Haitao Guo
- Institute for Biotechnology and Virology Research, Drexel University College of Medicine, Doylestown, PA, 18902, USA; Department of Microbiology and Immunology, Indiana University School of Medicine, Indianapolis, IN, 46202, USA.
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Bai W, Cui X, Chen R, Tao S, Hong R, Zhang J, Zhang J, Wang Y, Xie Y, Liu J. Re-Designed Recombinant Hepatitis B Virus Vectors Enable Efficient Delivery of Versatile Cargo Genes to Hepatocytes with Improved Safety. Viruses 2016; 8:v8050129. [PMID: 27171107 PMCID: PMC4885084 DOI: 10.3390/v8050129] [Citation(s) in RCA: 9] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/23/2016] [Revised: 04/22/2016] [Accepted: 05/04/2016] [Indexed: 12/18/2022] Open
Abstract
Hepatitis B virus (HBV) takes humans as its sole natural host, and productive infection in vivo is restricted exclusively to hepatocytes in the liver. Consequently, HBV-derived viral vectors are attractive candidates for liver-targeting gene therapies. Previously, we developed a novel recombinant HBV vector, designated 5c3c, from a highly replicative clinical isolate. 5c3c was demonstrated to be capable of efficiently delivering protein or RNA expression into infected primary tupaia hepatocytes (PTH), but the design of 5c3c imposes stringent restrictions on inserted sequences, which have limited its wider adoption. In this work, we addressed issues with 5c3c by re-designing the insertion strategy. The resultant vector, designated 5dCG, was more replicative than parental 5c3c, imposed no specific restrictions on inserted sequences, and allowed insertion of a variety of cargo genes without significant loss of replication efficiency. 5dCG-based recombinant HBV effectively delivered protein and RNA expression into infected PTH. Furthermore, due to the loss of functional core ORF, 5dCG vectors depend on co-infecting wild type HBV for replication and efficient expression of cargo genes. Development of the improved 5dCG vector makes wider applications of recombinant HBV possible, while dependence on co-infecting wild type HBV results in improved safety for certain in vivo applications.
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Affiliation(s)
- Weiya Bai
- Key Laboratory of Medical Molecular Virology (MOH & MOE) and Institutes of Biomedical Sciences, School of Basic Medical Sciences, Shanghai Medical College, Fudan University, Shanghai 200032, China.
| | - Xiaoxian Cui
- Key Laboratory of Medical Molecular Virology (MOH & MOE) and Institutes of Biomedical Sciences, School of Basic Medical Sciences, Shanghai Medical College, Fudan University, Shanghai 200032, China.
| | - Ruidong Chen
- Key Laboratory of Medical Molecular Virology (MOH & MOE) and Institutes of Biomedical Sciences, School of Basic Medical Sciences, Shanghai Medical College, Fudan University, Shanghai 200032, China.
| | - Shuai Tao
- Key Laboratory of Medical Molecular Virology (MOH & MOE) and Institutes of Biomedical Sciences, School of Basic Medical Sciences, Shanghai Medical College, Fudan University, Shanghai 200032, China.
| | - Ran Hong
- Key Laboratory of Medical Molecular Virology (MOH & MOE) and Institutes of Biomedical Sciences, School of Basic Medical Sciences, Shanghai Medical College, Fudan University, Shanghai 200032, China.
| | - Jiming Zhang
- Department of Infectious Diseases, Huashan Hospital, Fudan University, Shanghai 200040, China.
| | - Junqi Zhang
- Key Laboratory of Medical Molecular Virology (MOH & MOE) and Institutes of Biomedical Sciences, School of Basic Medical Sciences, Shanghai Medical College, Fudan University, Shanghai 200032, China.
| | - Yongxiang Wang
- Key Laboratory of Medical Molecular Virology (MOH & MOE) and Institutes of Biomedical Sciences, School of Basic Medical Sciences, Shanghai Medical College, Fudan University, Shanghai 200032, China.
| | - Youhua Xie
- Key Laboratory of Medical Molecular Virology (MOH & MOE) and Institutes of Biomedical Sciences, School of Basic Medical Sciences, Shanghai Medical College, Fudan University, Shanghai 200032, China.
| | - Jing Liu
- Key Laboratory of Medical Molecular Virology (MOH & MOE) and Institutes of Biomedical Sciences, School of Basic Medical Sciences, Shanghai Medical College, Fudan University, Shanghai 200032, China.
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Blondot ML, Bruss V, Kann M. Intracellular transport and egress of hepatitis B virus. J Hepatol 2016; 64:S49-S59. [PMID: 27084037 DOI: 10.1016/j.jhep.2016.02.008] [Citation(s) in RCA: 71] [Impact Index Per Article: 7.9] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 12/01/2015] [Revised: 01/27/2016] [Accepted: 02/03/2016] [Indexed: 12/23/2022]
Abstract
Hepatitis B virus (HBV) replicates its genomic information in the nucleus via transcription and therefore has to deliver its partially double stranded DNA genome into the nucleus. Like other viruses with a nuclear replication phase, HBV genomes are transported inside the viral capsids first through the cytoplasm towards the nuclear envelope. Following the arrival at the nuclear pore, the capsids are transported through, using classical cellular nuclear import pathways. The arrest of nuclear import at the nucleoplasmic side of the nuclear pore is unique, however, and is where the capsids efficiently disassemble leading to genome release. In the latter phase of the infection, newly formed nucleocapsids in the cytosol have to move to budding sites at intracellular membranes carrying the three viral envelope proteins. Capsids containing single stranded nucleic acid are not enveloped, in contrast to empty and double stranded DNA containing capsids. A small linear domain in the large envelope protein and two areas on the capsid surface have been mapped, where point mutations strongly block nucleocapsid envelopment. It is possible that these domains are involved in the envelope--with capsid interactions driving the budding process. Like other enveloped viruses, HBV also uses the cellular endosomal sorting complexes required for transport (ESCRT) machinery for catalyzing budding through the membrane and away from the cytosol.
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Affiliation(s)
- Marie-Lise Blondot
- Univ. de Bordeaux, Microbiologie Fondamentale et Pathogénicité, UMR 5234, Bordeaux, France; CNRS, Microbiologie Fondamentale et Pathogénicité, UMR 5234, Bordeaux, France
| | - Volker Bruss
- Institute for Virology, Helmholtz Zentrum München, Technische Universität Muenchen, Neuherberg, Germany
| | - Michael Kann
- Univ. de Bordeaux, Microbiologie Fondamentale et Pathogénicité, UMR 5234, Bordeaux, France; CNRS, Microbiologie Fondamentale et Pathogénicité, UMR 5234, Bordeaux, France; CHU de Bordeaux, Bordeaux, France.
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46
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Takata A, Otsuka M, Ohno M, Kishikawa T, Yoshikawa T, Koike K. Mutual antagonism between hepatitis B viral mRNA and host microRNA let-7. Sci Rep 2016; 6:23237. [PMID: 26979389 PMCID: PMC4793232 DOI: 10.1038/srep23237] [Citation(s) in RCA: 24] [Impact Index Per Article: 2.7] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/26/2016] [Accepted: 03/02/2016] [Indexed: 02/06/2023] Open
Abstract
The interplay between viral and host factors plays a major role in viral pathogenesis. Hepatitis B virus (HBV) infection is a global health problem that leads to liver cirrhosis and hepatocellular carcinoma (HCC). Although HBV proteins have been studied extensively about their implication in hepatocarcinogenesis, the molecular mechanisms of oncogenesis are still largely unknown. A recent concept in gene regulation, in which competitive endogenous RNAs compete for common microRNAs (miRNAs), suggests that mRNA targets are key elements in the regulation of miRNA availability. Here, we show that HBV mRNA in the preS2 region can be targeted by host miRNA let-7 g. This leads to the sequestration of let-7 g and inhibition of let-7 g function. The expression of HBV transcripts, including the preS2 region, de-repressed let-7 g targets, which may contribute to long-term oncogenesis. HBV transcript-expressing transgenic mice, but not non-targeted transcript-expressing mice, were more prone to chemically induced hepatoocarcinogenesis. Let-7 target protein expression was upregulated in human HCC tissues derived from HBV-infected patients. On the other hand, let-7 g inhibited HBV preS2 protein expression and viral products. These results suggest that the interplay between viral intermediate transcripts during HBV replication and host miRNAs is crucial to the pathogenesis of chronic viral infection.
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Affiliation(s)
- Akemi Takata
- Department of Gastroenterology, Graduate School of Medicine, The University of Tokyo, Tokyo 113-8655, Japan
| | - Motoyuki Otsuka
- Department of Gastroenterology, Graduate School of Medicine, The University of Tokyo, Tokyo 113-8655, Japan
- Japan Science and Technology Agency, PRESTO, Kawaguchi, Saitama 332-0012, Japan
| | - Motoko Ohno
- Department of Gastroenterology, Graduate School of Medicine, The University of Tokyo, Tokyo 113-8655, Japan
| | - Takahiro Kishikawa
- Department of Gastroenterology, Graduate School of Medicine, The University of Tokyo, Tokyo 113-8655, Japan
| | - Takeshi Yoshikawa
- Department of Gastroenterology, Graduate School of Medicine, The University of Tokyo, Tokyo 113-8655, Japan
| | - Kazuhiko Koike
- Department of Gastroenterology, Graduate School of Medicine, The University of Tokyo, Tokyo 113-8655, Japan
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47
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Zhang X, Lu W, Zheng Y, Wang W, Bai L, Chen L, Feng Y, Zhang Z, Yuan Z. In situ analysis of intrahepatic virological events in chronic hepatitis B virus infection. J Clin Invest 2016; 126:1079-92. [PMID: 26901811 DOI: 10.1172/jci83339] [Citation(s) in RCA: 75] [Impact Index Per Article: 8.3] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/19/2015] [Accepted: 12/23/2015] [Indexed: 12/11/2022] Open
Abstract
Persistent hepatitis B virus (HBV) infection is established by the formation of an intranuclear pool of covalently closed circular DNA (cccDNA) in the liver. Very little is known about the intrahepatic distribution of HBV cccDNA in infected patients, particularly at the single-cell level. Here, we established a highly sensitive and specific ISH assay for the detection of HBV RNA, DNA, and cccDNA. The specificity of our cccDNA probe set was confirmed by its strict intranuclear signal and by a series of Southern blot analyses. Use of our in situ assay in conjunction with IHC or immunofluorescence uncovered a surprisingly mosaic distribution of viral antigens and nucleic acids. Most strikingly, a mutually exclusive pattern was found between HBV surface antigen-positive (HBsA-positive) and HBV DNA- and cccDNA-positive cells. A longitudinal observation of patients over a 1-year period of adeforvir therapy confirmed the persistence of a nuclear reservoir of viral DNA, although cytoplasmic DNA was effectively depleted in these individuals. In conclusion, our method for detecting viral nucleic acids, including cccDNA, with single-cell resolution provides a means for monitoring intrahepatic virological events in chronic HBV infection. More important, our observations unravel the complexity of the HBV life cycle in vivo.
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48
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Zhang YY. Duck Hepatitis B Virus cccDNA Amplification Efficiency in Natural Infection Is Regulated by Virus Secretion Efficiency. PLoS One 2015; 10:e0145465. [PMID: 26713436 PMCID: PMC4694612 DOI: 10.1371/journal.pone.0145465] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.2] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/27/2015] [Accepted: 12/03/2015] [Indexed: 01/01/2023] Open
Abstract
Previous mutation based studies showed that ablating synthesis of viral envelope proteins led to elevated hepadnaviral covalently closed circular DNA (cccDNA) amplification, but it remains unknown how cccDNA amplification is regulated in natural hepadnaviral infection because of a lack of research system. In this study we report a simple procedure to prepare two identical duck hepatitis B virus inocula, but they possess 10-100-fold difference in cccDNA amplification in infected cell culture. We demonstrate that the infected cells with higher cccDNA amplification significantly reduce the virus secretion efficiency that results in higher accumulation of relaxed circular DNA (rcDNA) and DHBsAg in the cells. The infected cells with lower cccDNA amplification significantly increase the virus secretion efficiency that leads to lower intracellular rcDNA and DHBsAg accumulation. In contrast with the findings generated in the mutation based experimental system, the regulation of cccDNA amplification in natural hepadnaviral infection bypasses direct regulation of the cellular envelope proteins concentration, instead it modulates virus secretion efficiency that ultimately impacts the intracellular rcDNA concentration, an important factor determining the destination of the synthesized rcDNA in infected cells.
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Affiliation(s)
- Yong-Yuan Zhang
- HBVtech, Germantown, Maryland, United States of America
- * E-mail:
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49
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Liang TJ, Block TM, McMahon BJ, Ghany MG, Urban S, Guo JT, Locarnini S, Zoulim F, Chang KM, Lok AS. Present and future therapies of hepatitis B: From discovery to cure. Hepatology 2015; 62:1893-908. [PMID: 26239691 PMCID: PMC4681668 DOI: 10.1002/hep.28025] [Citation(s) in RCA: 251] [Impact Index Per Article: 25.1] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 03/29/2015] [Accepted: 07/31/2015] [Indexed: 12/11/2022]
Abstract
UNLABELLED Hepatitis B virus (HBV) is a significant global pathogen, infecting more than 240 million people worldwide. While treatment for HBV has improved, HBV patients often require lifelong therapies and cure is still a challenging goal. Recent advances in technologies and pharmaceutical sciences have heralded a new horizon of innovative therapeutic approaches that are bringing us closer to the possibility of a functional cure of chronic HBV infection. In this article, we review the current state of science in HBV therapy and highlight new and exciting therapeutic strategies spurred by recent scientific advances. Some of these therapies have already entered into clinical phase, and we will likely see more of them moving along the development pipeline. CONCLUSION With growing interest in developing and efforts to develop more effective therapies for HBV, the challenging goal of a cure may be well within reach in the near future.
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Affiliation(s)
- T. Jake Liang
- Liver Diseases Branch, NIDDK, NIH, Bethesda, MD. USA
| | | | - Brian J. McMahon
- National Center for Emerging and Zoonotic Infectious Diseases, Centers for Disease Control and Prevention, Anchorage, AK. USA
| | - Marc G. Ghany
- Liver Diseases Branch, NIDDK, NIH, Bethesda, MD. USA
| | - Stephan Urban
- Dept of Infectious Diseases, Molecular Virology and German Center for Infection Diseases (DZIF), Univ Hospital Heidelberg, Heidelberg, Germany
| | - Ju-Tao Guo
- Baruch S. Blumberg Institute, Doylestown, PA. USA
| | | | - Fabien Zoulim
- Victorian Infectious Diseases Reference Laboratory, Doherty Institute, Melbourne, VIC, Australia
| | - Kyong-Mi Chang
- Dept of Medicine, Philadelphia VAMC & University of Pennsylvania, Philadelphia, PA. USA
| | - Anna S. Lok
- Div of Gastroenterology and Hepatology, Univ of Michigan, Ann Arbor, MI. USA
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50
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Guo JT, Guo H. Metabolism and function of hepatitis B virus cccDNA: Implications for the development of cccDNA-targeting antiviral therapeutics. Antiviral Res 2015; 122:91-100. [PMID: 26272257 DOI: 10.1016/j.antiviral.2015.08.005] [Citation(s) in RCA: 120] [Impact Index Per Article: 12.0] [Reference Citation Analysis] [Abstract] [Key Words] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/16/2015] [Accepted: 08/07/2015] [Indexed: 02/07/2023]
Abstract
Persistent hepatitis B virus (HBV) infection relies on the stable maintenance and proper functioning of a nuclear episomal form of the viral genome called covalently closed circular (ccc) DNA. One of the major reasons for the failure of currently available antiviral therapeutics to achieve a cure of chronic HBV infection is their inability to eradicate or inactivate cccDNA. In this review article, we summarize our current understanding of cccDNA metabolism in hepatocytes and the modulation of cccDNA by host pathophysiological and immunological cues. Perspectives on the future investigation of cccDNA biology, as well as strategies and progress in therapeutic elimination and/or transcriptional silencing of cccDNA through rational design and phenotypic screenings, are also discussed. This article forms part of a symposium in Antiviral Research on "An unfinished story: from the discovery of the Australia antigen to the development of new curative therapies for hepatitis B."
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Affiliation(s)
- Ju-Tao Guo
- Baruch S. Blumberg Institute, Hepatitis B Foundation, Doylestown, PA 18902, USA.
| | - Haitao Guo
- Department of Microbiology and Immunology, Indiana University School of Medicine, 635 Barnhill Drive, Indianapolis, IN 46202, USA.
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