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Choi YM, Kim DH, Cho EJ, Kim Z, Jang J, Kim H, Yu SJ, Kim BJ. The sV184A Variant in HBsAg Specific to HBV Subgenotype C2 Leads to Enhanced Viral Replication and Apoptotic Cell Death Induced by PERK-eIF2α-CHOP-Mediated ER Stress. J Med Virol 2025; 97:e70253. [PMID: 39977392 DOI: 10.1002/jmv.70253] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/03/2024] [Revised: 01/15/2025] [Accepted: 02/05/2025] [Indexed: 02/22/2025]
Abstract
HBV genotype C, particularly subgenotype C2, is associated with an elevated risk of HCC and aggressive disease activity. We previously identified a nonsynonymous sV184A variant in the HBsAg region, predominantly in HBV subgenotype C2. This study investigates the mechanistic role of the sV184A variant in promoting liver disease progression. Analysis of 109 chronically HBV-infected patients revealed that the sV184A variant correlates with significantly elevated HBV DNA. Both patient data and public database indicated that sV184A is associated with high frequency of BCP mutations, however, the high HBV DNA in the sV184A group are independent of the presence of BCP mutations. In vitro and in vivo studies demonstrated that the sV184A variant enhances HBV replication and induces ER stress via the PERK-eIF2α-CHOP pathway, leading to apoptosis. HBV large surface (LHB)(LHB) protein was found to be a key factor, responsible for the strong ER stress, as the sV184A variant increases LHB protein stability. Pharmacological inhibition of PERK signaling or mutation of the LHB mitigated HBV proliferation and apoptosis induced by the sV184A variant. The sV184A variant specific to HBV subgenotype C2 significantly promotes HBV replication and apoptosis, serving as a driver of advanced liver disease and potentially increasing mutation rates in affected patients.
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Affiliation(s)
- Yu-Min Choi
- Department of Microbiology and Immunology, College of Medicine, Seoul National University, Seoul, Republic of Korea
- Seoul National University Medical Research Center (SNUMRC), Seoul, Republic of Korea
| | - Dong Hyun Kim
- Department of Microbiology and Immunology, College of Medicine, Seoul National University, Seoul, Republic of Korea
| | - Eun Ju Cho
- Department of Internal Medicine and Liver Research Institute, Seoul National University College of Medicine, Seoul, Republic of Korea
| | - Ziyun Kim
- Department of Microbiology and Immunology, College of Medicine, Seoul National University, Seoul, Republic of Korea
| | - Junghwa Jang
- Department of Microbiology and Immunology, College of Medicine, Seoul National University, Seoul, Republic of Korea
| | - Hyunsoo Kim
- Department of Convergent Bioscience and Informatics, Chungnam National University, Daejeon, Republic of Korea
| | - Su Jong Yu
- Department of Internal Medicine and Liver Research Institute, Seoul National University College of Medicine, Seoul, Republic of Korea
| | - Bum-Joon Kim
- Department of Microbiology and Immunology, College of Medicine, Seoul National University, Seoul, Republic of Korea
- Seoul National University Medical Research Center (SNUMRC), Seoul, Republic of Korea
- Department of Biomedical Sciences, College of Medicine, Seoul National University, Seoul, Republic of Korea
- Liver Research Institute, College of Medicine, Seoul National University, Seoul, Republic of Korea
- Cancer Research Institute, College of Medicine, Seoul National University, Seoul, Republic of Korea
- Institute of Endemic Disease, Seoul National University Medical Research Center, Seoul, Republic of Korea
- BK21 FOUR Biomedical Science Project, Seoul National University College of Medicine, Seoul, Republic of Korea
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Sedohara A, Takahashi K, Arai K, Arizono K, Tuvshinjargal K, Saito M, Nakahara F, Tsutsumi T, Ikeuchi K, Adachi E, Yotsuyanagi H. Characterization of mutations in hepatitis B virus DNA isolated from Japanese HBsAg-positive blood donors in 2021 and 2022. Arch Virol 2024; 169:103. [PMID: 38632180 PMCID: PMC11023964 DOI: 10.1007/s00705-024-06016-4] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/11/2023] [Accepted: 02/04/2024] [Indexed: 04/19/2024]
Abstract
Missense mutations in certain small envelope proteins diminish the efficacy of antibodies. Consequently, tracking the incidence and types of vaccine-escape mutations (VEMs) was crucial both before and after the introduction of universal hepatitis B vaccination in Japan in 2016. In this study, we isolated hepatitis B virus (HBV) DNA from 58 of 169 hepatitis B surface antigen (HBsAg)-positive blood samples from Japanese blood donors and determined the nucleotide sequence encoding the small envelope protein. DNA from six (10%) of the samples had VEMs, but no missense mutations, such as G145R, were detected. Complete HBV genome sequences were obtained from 29 of the 58 samples; the viral genotype was A1 in one (3%), A2 in three (10%), B1 in nine (31%), B2 in five (17%), B4 in one (3%), and C2 in 10 (34%) samples. Tenofovir-resistance mutations were detected in two (7%) samples. In addition, several core promoter mutations, such as 1762A>T and 1764G>A, and a precore nonsense mutation, 1986G>A, which are risk factors for HBV-related chronic liver disease, were detected. These findings provide a baseline for future research and highlight the importance of ongoing monitoring of VEMs and drug resistance mutations in HBV DNA from HBsAg-positive blood donors without HBV antibodies.
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Affiliation(s)
- Ayako Sedohara
- Division of Infectious Diseases, Advanced Clinical Research Center, Institute of Medical Science, The University of Tokyo, Tokyo, Japan.
| | - Kazuaki Takahashi
- Division of Infectious Diseases, Advanced Clinical Research Center, Institute of Medical Science, The University of Tokyo, Tokyo, Japan
| | - Keiko Arai
- Division of Infectious Diseases, Advanced Clinical Research Center, Institute of Medical Science, The University of Tokyo, Tokyo, Japan
| | - Kotaro Arizono
- Department of Computational Biology and Medical Sciences, Graduate School of Frontier Sciences, The University of Tokyo, Tokyo, Japan
| | - Khulan Tuvshinjargal
- Department of Computational Biology and Medical Sciences, Graduate School of Frontier Sciences, The University of Tokyo, Tokyo, Japan
| | - Makoto Saito
- Department of Infectious Disease and Applied Immunology, IMSUT Hospital of The Institute of Medical Science, The University of Tokyo, Tokyo, Japan
| | - Fumio Nakahara
- Division of Infectious Diseases, Advanced Clinical Research Center, Institute of Medical Science, The University of Tokyo, Tokyo, Japan
- Division of Regenerative Medicine, Center for Molecular Medicine, Jichi Medical University, Tochigi, Japan
| | - Takeya Tsutsumi
- Department of Infectious Disease and Applied Immunology, IMSUT Hospital of The Institute of Medical Science, The University of Tokyo, Tokyo, Japan
- Department of Infectious Diseases, Faculty of Medicine, The University of Tokyo, Tokyo, Japan
| | - Kazuhiko Ikeuchi
- Department of Infectious Disease and Applied Immunology, IMSUT Hospital of The Institute of Medical Science, The University of Tokyo, Tokyo, Japan
| | - Eisuke Adachi
- Department of Infectious Disease and Applied Immunology, IMSUT Hospital of The Institute of Medical Science, The University of Tokyo, Tokyo, Japan
| | - Hiroshi Yotsuyanagi
- Division of Infectious Diseases, Advanced Clinical Research Center, Institute of Medical Science, The University of Tokyo, Tokyo, Japan.
- Department of Infectious Disease and Applied Immunology, IMSUT Hospital of The Institute of Medical Science, The University of Tokyo, Tokyo, Japan.
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Hou L, Zhao J, Cai L, Jin L, Liu B, Li S, Yang J, Ji T, Li S, Shi L, Shen B, Yu H, Wang Y, Cai X. HBV PreC interacts with SUV39H1 to induce viral replication by blocking the proteasomal degradation of viral polymerase. J Med Virol 2024; 96:e29607. [PMID: 38628076 DOI: 10.1002/jmv.29607] [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: 01/11/2024] [Revised: 03/13/2024] [Accepted: 04/05/2024] [Indexed: 04/19/2024]
Abstract
Hepatitis B e antigen (HBeAg) seropositivity during the natural history of chronic hepatitis B (CHB) is known to coincide with significant increases in serum and intrahepatic HBV DNA levels. However, the precise underlying mechanism remains unclear. In this study, we found that PreC (HBeAg precursor) genetic ablation leads to reduced viral replication both in vitro and in vivo. Furthermore, PreC impedes the proteasomal degradation of HBV polymerase, promoting viral replication. We discovered that PreC interacts with SUV39H1, a histone methyltransferase, resulting in a reduction in the expression of Cdt2, an adaptor protein of CRL4 E3 ligase targeting HBV polymerase. SUV39H1 induces H3K9 trimethylation of the Cdt2 promoter in a PreC-induced manner. CRISPR-mediated knockout of endogenous SUV39H1 or pharmaceutical inhibition of SUV39H1 decreases HBV loads in the mouse liver. Additionally, genetic depletion of Cdt2 in the mouse liver abrogates PreC-related HBV replication. Interestingly, a negative correlation of intrahepatic Cdt2 with serum HBeAg and HBV DNA load was observed in CHB patient samples. Our study thus sheds light on the mechanistic role of PreC in inducing HBV replication and identifies potential therapeutic targets for HBV treatment.
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Affiliation(s)
- Lidan Hou
- Department of General Surgery, Sir Run Run Shaw Hospital, Zhejiang University School of Medicine, Hangzhou, China
- Zhejiang Province Medical Research Center of Minimally Invasive Diagnosis and Treatment of Abdominal Diseases, Hangzhou, China
- National Engineering Research Center of Innovation and Application of Minimally Invasive Instruments, Hangzhou, China
| | - Jie Zhao
- Department of General Surgery, Sir Run Run Shaw Hospital, Zhejiang University School of Medicine, Hangzhou, China
- Zhejiang Province Medical Research Center of Minimally Invasive Diagnosis and Treatment of Abdominal Diseases, Hangzhou, China
- National Engineering Research Center of Innovation and Application of Minimally Invasive Instruments, Hangzhou, China
| | - Liuxin Cai
- Department of General Surgery, Sir Run Run Shaw Hospital, Zhejiang University School of Medicine, Hangzhou, China
| | - Ling Jin
- Department of General Surgery, Sir Run Run Shaw Hospital, Zhejiang University School of Medicine, Hangzhou, China
- Zhejiang Province Medical Research Center of Minimally Invasive Diagnosis and Treatment of Abdominal Diseases, Hangzhou, China
- National Engineering Research Center of Innovation and Application of Minimally Invasive Instruments, Hangzhou, China
| | - Boqiang Liu
- Department of General Surgery, Sir Run Run Shaw Hospital, Zhejiang University School of Medicine, Hangzhou, China
- Zhejiang Province Medical Research Center of Minimally Invasive Diagnosis and Treatment of Abdominal Diseases, Hangzhou, China
- National Engineering Research Center of Innovation and Application of Minimally Invasive Instruments, Hangzhou, China
| | - Shijie Li
- Department of General Surgery, Sir Run Run Shaw Hospital, Zhejiang University School of Medicine, Hangzhou, China
- Zhejiang Province Medical Research Center of Minimally Invasive Diagnosis and Treatment of Abdominal Diseases, Hangzhou, China
- National Engineering Research Center of Innovation and Application of Minimally Invasive Instruments, Hangzhou, China
| | - Jin Yang
- Department of General Surgery, Sir Run Run Shaw Hospital, Zhejiang University School of Medicine, Hangzhou, China
| | - Tong Ji
- Department of General Surgery, Sir Run Run Shaw Hospital, Zhejiang University School of Medicine, Hangzhou, China
- Zhejiang Province Medical Research Center of Minimally Invasive Diagnosis and Treatment of Abdominal Diseases, Hangzhou, China
- National Engineering Research Center of Innovation and Application of Minimally Invasive Instruments, Hangzhou, China
| | - Songyi Li
- Animal Center, Sir Run Run Shaw Hospital, Zhejiang University School of Medicine, Hangzhou, China
| | - Liang Shi
- Department of General Surgery, Sir Run Run Shaw Hospital, Zhejiang University School of Medicine, Hangzhou, China
- Zhejiang Province Medical Research Center of Minimally Invasive Diagnosis and Treatment of Abdominal Diseases, Hangzhou, China
- National Engineering Research Center of Innovation and Application of Minimally Invasive Instruments, Hangzhou, China
| | - Bo Shen
- Department of General Surgery, Sir Run Run Shaw Hospital, Zhejiang University School of Medicine, Hangzhou, China
- Zhejiang Province Medical Research Center of Minimally Invasive Diagnosis and Treatment of Abdominal Diseases, Hangzhou, China
| | - Hong Yu
- Department of General Surgery, Sir Run Run Shaw Hospital, Zhejiang University School of Medicine, Hangzhou, China
- Zhejiang Province Medical Research Center of Minimally Invasive Diagnosis and Treatment of Abdominal Diseases, Hangzhou, China
| | - Yifan Wang
- Department of General Surgery, Sir Run Run Shaw Hospital, Zhejiang University School of Medicine, Hangzhou, China
- Zhejiang Province Medical Research Center of Minimally Invasive Diagnosis and Treatment of Abdominal Diseases, Hangzhou, China
- National Engineering Research Center of Innovation and Application of Minimally Invasive Instruments, Hangzhou, China
| | - Xiujun Cai
- Department of General Surgery, Sir Run Run Shaw Hospital, Zhejiang University School of Medicine, Hangzhou, China
- Zhejiang Province Medical Research Center of Minimally Invasive Diagnosis and Treatment of Abdominal Diseases, Hangzhou, China
- National Engineering Research Center of Innovation and Application of Minimally Invasive Instruments, Hangzhou, China
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Langat BK, Ochwedo KO, Borlang J, Osiowy C, Mutai A, Okoth F, Muge E, Andonov A, Maritim ES. Genetic diversity, haplotype analysis, and prevalence of Hepatitis B virus MHR mutations among isolates from Kenyan blood donors. PLoS One 2023; 18:e0291378. [PMID: 37963165 PMCID: PMC10645356 DOI: 10.1371/journal.pone.0291378] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/06/2023] [Accepted: 08/28/2023] [Indexed: 11/16/2023] Open
Abstract
BACKGROUND The rapid spread of HBV has resulted in the emergence of new variants. These viral genotypes and variants, in addition to carcinogenic risk, can be key predictors of therapy response and outcomes. As a result, a better knowledge of these emerging HBV traits will aid in the development of a treatment for HBV infection. However, many Sub-Saharan African nations, including Kenya, have insufficient molecular data on HBV strains circulating locally. This study conducted a population-genetics analysis to evaluate the genetic diversity of HBV among Kenyan blood donors. In addition, within the same cohort, the incidence and features of immune-associated escape mutations and stop-codons in Hepatitis B surface antigen (HBsAg) were determined. METHODS In September 2015 to October 2016, 194 serum samples were obtained from HBsAg-positive blood donors residing in eleven different Kenyan counties: Kisumu, Machakos, Uasin Gishu, Nairobi, Nakuru, Embu, Garissa, Kisii, Mombasa, Nyeri, and Turkana. For the HBV surface (S) gene, HBV DNA was isolated, amplified, and sequenced. The sequences obtained were utilized to investigate the genetic and haplotype diversity within the S genes. RESULTS Among the blood donors, 74.74% were male, and the overall mean age was 25.36 years. HBV genotype A1 (88.14%) was the most common, followed by genotype D (10.82%), genotype C (0.52%), and HBV genotype E (0.52%). The phylogenetic analysis revealed twelve major clades, with cluster III comprising solely of 68 blood donor isolates (68/194-35.05%). A high haplotype diversity (Hd = 0.94) and low nucleotide diversity (π = 0.02) were observed. Kisumu county had high number of haplotypes (22), but low haplotype (gene) diversity (Hd = 0.90). Generally, a total of 90 haplotypes with some consisting of more than one sequence were observed. The gene exhibited negative values for Tajima's D (-2.04, p<0.05) and Fu's Fs (-88.84). Several mutations were found in 139 isolates, either within or outside the Major Hydrophilic Area (MHR). There were 29 mutations found, with 37.9% of them situated inside the "a" determinant. The most common mutations in this research were T143M and K122R. Escape mutations linked to diagnostic failure, vaccination and immunoglobulin treatment evasion were also discovered. Also, one stop-codon, W163STP, inside the MHR, was found in one sample from genotype A. CONCLUSION In Kenya, HBV/A1 is still the most common genotype. Despite limited genetic and nucleotide diversity, haplotype network analysis revealed haplotype variance among HBV genotypes from Kenyan blood donors. The virological properties of immune escape, which may be the source of viral replication endurance, were discovered in the viral strains studied and included immune-escape mutations and stop-codon. The discovery of HBsAg mutations in MHR in all isolates highlighted the need of monitoring MHR mutations in Kenya.
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Affiliation(s)
| | - Kevin Omondi Ochwedo
- Department of Biology, Faculty of Science and Technology, University of Nairobi, Nairobi, Kenya
| | | | - Carla Osiowy
- National Microbiology Laboratory, Winnipeg, Canada
| | - Alex Mutai
- Kenya National Blood Transfusion Services, Nairobi, Kenya
| | - Fredrick Okoth
- Centre for Virus Research, Kenya Medical Research Institute, Nairobi, Kenya
| | - Edward Muge
- Department of Medical Biochemistry, University of Nairobi, Nairobi, Kenya
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Establishment and Characterization of an HBV Viral Spread and Infectious System following Long-Term Passage of an HBV Clinical Isolate in the Primary Human Hepatocyte and Fibroblast Coculture System. J Virol 2022; 96:e0084922. [PMID: 36037476 PMCID: PMC9517703 DOI: 10.1128/jvi.00849-22] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/20/2022] Open
Abstract
The existing cell culture-based methods to study hepatitis B virus (HBV) have limitations and do not allow for viral long-term passage. The aim of this study was to develop a robust in vitro long-term viral passage system with optimized cell culture conditions and a viral isolate with the ability to spread and passage. An HBV genotype A clinical isolate was subjected to multiple rounds of UV treatment and passaged in an optimized primary human hepatocyte (PHH)/human fibroblast coculture system. The passaged UV-treated virus was sequenced and further characterized. In addition, a panel of mutant viruses containing different combinations of mutations observed in this virus was investigated. The clinical isolate was passaged for 20 rounds with 21 days per round in an optimized PHH/human fibroblast coculture system while subject to UV mutagenesis. This passaged UV-mutated isolate harbored four mutations: G225A (sR24K) in the S gene, A2062T in the core gene, and two mutations G1764A and C1766T (xV131I) in the basal core promoter (BCP) region. In vitro characterization of the four mutations suggested that the two BCP mutations G1764A and C1766T contributed to the increased viral replication and viral infectivity. A robust in vitro long-term HBV viral passage system has been established by passaging a UV-treated clinical isolate in an optimized PHH/fibroblast coculture system. The two BCP mutations played a key role in the virus's ability to passage. This passage system can be used for studying the entire life cycle of HBV and has the potential for in vitro drug-resistance selection upon further optimization. IMPORTANCE The existing cell culture-based methods to study HBV have limitations and do not allow for viral long-term passage. In this study, an HBV genotype A clinical isolate was subjected to multiple rounds of UV treatment and passaged in an optimized PHH/human fibroblast coculture system. This passaged UV-mutated isolate carried four mutations across the HBV genome, and in vitro characterization of the four mutations suggested that the two basal core promoter (BCP) mutations G1764A and C1766T played a key role in the virus's ability to passage. In summary, we have developed a robust in vitro long-term HBV viral passage system by passaging an UV-treated HBV genotype A clinical isolate in an optimized PHH/human fibroblast coculture system. This passage system can be used for studying the entire life cycle of HBV and has the potential for in vitro drug-resistance selection upon further optimization.
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Wei S, Hu M, Chen H, Xie Q, Wang P, Li H, Peng J. Effectiveness of antiviral treatment in HBeAg-negative chronic hepatitis B patients with normal or mildly elevated alanine aminotransferase: a retrospective study. BMC Gastroenterol 2022; 22:387. [PMID: 35978283 PMCID: PMC9387004 DOI: 10.1186/s12876-022-02471-y] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.7] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 04/01/2022] [Accepted: 08/09/2022] [Indexed: 11/29/2022] Open
Abstract
Background There are inadequate data and no histological evidence regarding the effects of antiviral treatment for hepatitis B e-antigen (HBeAg)-negative chronic hepatitis B (CHB) patients with normal or mildly elevated alanine aminotransferase (ALT). This study investigated the effects of antiviral treatment on these patients. Methods We retrospectively analysed the outcomes of antiviral treatment for HBeAg-negative CHB patients with normal or mildly elevated ALT who were treated with nucleoside/nucleotide analogues (NAs) for up to 96 weeks. Results A total of 128 patients were enrolled; 74 patients had normal ALT and 54 patients had mildly elevated ALT. The total cumulative rates of viral suppression were 64.06%, 81.97%, and 96.39%, at weeks 24, 48, and 96, respectively. The cumulative rates of viral suppression for the normal and mildly elevated ALT groups were 67.85% and 58.97%, 86.39% and 76.31%, and 93.13% and 97.04% at weeks 24, 48, and 96, respectively. The serum HBV DNA levels at week 12 and hepatitis B surface antigen (HBsAg) levels at week 24 were significant predictors of the 96-week virological response. Of the 128 patients, 54 with normal ALT and 33 with mildly elevated ALT underwent FibroScan at baseline. Significant fibrosis (F ≥ 2) was found in 44.4% (n = 24) and 51.5% (n = 17) of the patients in the normal ALT group and mildly elevated ALT group, respectively. Compared with the values at baseline, liver stiffness values significantly decreased at week 48 (8.12 kPa vs. 6.57 kPa; p < 0.001) and week 96 (8.87 kPa vs. 6.43 kPa; p < 0.001), respectively. Conclusions HBeAg-negative CHB patients with normal ALT could benefit from antiviral therapy with NAs, similar to patients with mildly elevated ALT. Antiviral treatment is strongly recommended for HBeAg-negative CHB patients with normal ALT. Additionally, significant liver fibrosis is not rare in HBeAg-negative CHB patients with ALT less than two-times the upper limit of normal, and FibroScan should be performed regularly for these patients. Supplementary Information The online version contains supplementary material available at 10.1186/s12876-022-02471-y.
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Affiliation(s)
- Sufang Wei
- State Key Laboratory of Organ Failure Research, Guangdong Provincial Key Laboratory of Viral Hepatitis Research, Department of Infectious Diseases, Nanfang Hospital, Southern Medical University, Guangzhou, 510080, China
| | - Meixin Hu
- State Key Laboratory of Organ Failure Research, Guangdong Provincial Key Laboratory of Viral Hepatitis Research, Department of Infectious Diseases, Nanfang Hospital, Southern Medical University, Guangzhou, 510080, China
| | - Hongjie Chen
- State Key Laboratory of Organ Failure Research, Guangdong Provincial Key Laboratory of Viral Hepatitis Research, Department of Infectious Diseases, Nanfang Hospital, Southern Medical University, Guangzhou, 510080, China
| | - Qiuli Xie
- Department of Infectious Diseases, Shunde Hospital, Southern Medical University, Foshan, 528300, China
| | - Peng Wang
- Department of Infectious Diseases, Shunde Hospital, Southern Medical University, Foshan, 528300, China
| | - Hong Li
- Department of Infectious Diseases, Shunde Hospital, Southern Medical University, Foshan, 528300, China
| | - Jie Peng
- State Key Laboratory of Organ Failure Research, Guangdong Provincial Key Laboratory of Viral Hepatitis Research, Department of Infectious Diseases, Nanfang Hospital, Southern Medical University, Guangzhou, 510080, China.
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Farshadpour F, Taherkhani R, Saberi F. Molecular evaluation of hepatitis B virus infection and predominant mutations of pre-core, basal core promoter and S regions in an Iranian population with type 2 diabetes mellitus: a case-control study. BMC Infect Dis 2022; 22:553. [PMID: 35715756 PMCID: PMC9206294 DOI: 10.1186/s12879-022-07528-7] [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: 02/22/2022] [Accepted: 06/06/2022] [Indexed: 12/01/2022] Open
Abstract
BACKGROUND This study was designed to evaluate the prevalence, genotypic patterns, and predominant mutations of hepatitis B virus (HBV) infection among diabetic patients. METHODS Serum samples were obtained from 733 patients with type 2 diabetes mellitus and 782 non-diabetic controls. The presence of HBsAg and HBcAb was determined by ELISA. Nested PCR, targeting S and pre-core regions of the HBV genome, followed by sequencing was carried out to determine HBV genotypes and predominant mutations in the S, basal core promoter (BCP), and pre-core regions of the HBV genome. RESULTS Of 733 diabetic patients, 94 cases (12.82%) were positive for HBcAb, 28 cases (3.82%) were positive for HBsAg, and 19 cases (2.59%) had HBV-DNA with genotype D, sub-genotype D1/D3 and subtype ayw2. An occult HBV infection was found in one of the HBV DNA-positive samples, which was positive for HBcAb but negative for HBsAg. P120T/G145R, G1896A/G1899A, and A1762T/G1764T were the most frequent point substitution mutations detected in the S, pre-core, and BCP regions of the HBV genome, respectively. P120T and G145R mutations were associated with low levels or undetectable levels of HBsAg in serum. Therefore, routine tests based on HBsAg detection cannot detect HBsAg-negative infected patients. CONCLUSIONS Relatively high prevalence of HBV infection was found in diabetic patients, while all of the HBV-infected patients were unaware of their infection. Therefore, screening for HBV infection should be included in the management program of diabetes for timely diagnosis and treatment of infected but asymptomatic patients.
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Affiliation(s)
- Fatemeh Farshadpour
- Department of Virology, School of Medicine, Bushehr University of Medical Sciences, Moallem Street, 751463334, Bushehr, Iran
| | - Reza Taherkhani
- Department of Virology, School of Medicine, Bushehr University of Medical Sciences, Moallem Street, 751463334, Bushehr, Iran.
| | - Fatemeh Saberi
- Department of Virology, School of Medicine, Bushehr University of Medical Sciences, Moallem Street, 751463334, Bushehr, Iran
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Liu T, Sun Q, Gu J, Cen S, Zhang Q. Characterization of the tenofovir resistance-associated mutations in the hepatitis B virus isolates across genotypes A to D. Antiviral Res 2022; 203:105348. [PMID: 35644506 DOI: 10.1016/j.antiviral.2022.105348] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/28/2022] [Revised: 05/09/2022] [Accepted: 05/23/2022] [Indexed: 11/24/2022]
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The Korean Association for the Study of the Liver (KASL). KASL clinical practice guidelines for management of chronic hepatitis B. Clin Mol Hepatol 2022; 28:276-331. [PMID: 35430783 PMCID: PMC9013624 DOI: 10.3350/cmh.2022.0084] [Citation(s) in RCA: 69] [Impact Index Per Article: 23.0] [Reference Citation Analysis] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 03/31/2022] [Accepted: 04/01/2022] [Indexed: 01/10/2023] Open
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Taherkhani R, Farshadpour F. Prevalence, genotype distribution and mutations of hepatitis B virus and the associated risk factors among pregnant women residing in the northern shores of Persian Gulf, Iran. PLoS One 2022; 17:e0265063. [PMID: 35271684 PMCID: PMC8912131 DOI: 10.1371/journal.pone.0265063] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/12/2021] [Accepted: 02/22/2022] [Indexed: 12/18/2022] Open
Abstract
BACKGROUND Considering perinatal transmission and the high rate of chronic hepatitis B virus (HBV) infection in infants, diagnosis of HBV infection during pregnancy and timely interventions are of great importance. Therefore, this study was performed to investigate the prevalence and genotype distribution of HBV infection and the associated risk factors among pregnant women in the northern shores of the Persian Gulf, South of Iran. METHODS Serum samples of 1425 pregnant women were tested for the presence of HBsAg and HBcAb by ELISA (HBsAg one-Version ULTRA and HBc Ab ELISA kits, DIA.PRO, Milan, Italy). The seropositive samples were tested for the presence of HBV DNA by nested PCR, targeting S, X, pre-core (pre-C), and basal core promoter (BCP) regions of the HBV genome. The amplified fragments were sequenced by Sanger dideoxy sequencing technology to evaluate the genotype distribution and mutations of HBV infection by using the MEGA 7 software. The HBV seropositive pregnant women were tested for HCV and HIV coinfections by ELISA (HCV Ab and HIV Ab/Ag ELISA kits, DIA.PRO, Milan, Italy). RESULTS Of 1425 participants, 15 pregnant women (1.05%, 95% CI: 0.64%-1.73%) were positive for HBsAg, 41 women (2.88%, 95% CI: 2.10%-3.88%) were positive for HBcAb, and 5 women (0.35%, 95% CI: 0.15% -0.82%) had HBV viremia with genotype D, sub-genotype D3 and subtype ayw2. One of the viremic samples was positive for HBcAb but negative for HBsAg, which is indicative of an occult HBV infection. HBsAg seroprevalence was higher among pregnant women aged 20 to 29 years, women in the third trimester of pregnancy, residents of Khormuj city, Afghan immigrants, illiterate women, and pregnant women with a history of tattoo and HBV vaccination. The highest rate of HBcAb seroprevalence was observed in residents of Borazjan city, Turk ethnicity, the age group >39 years, and those women with more parities and a history of abortion. Nevertheless, HBV seroprevalence among pregnant women was not statistically associated with these variables. In contrast, HBcAb seropositivity was significantly associated with the history of tattoo (P = 0.018). According to mutations analyses, seven amino acid substitutions in the HBsAg, one point mutation in the pre-C region, and five points mutations in the BCP region were detected. Besides, the BCP mutations caused amino acid substitutions in the X protein. Of note, the conversion of Ala → Val at amino acid 168 (A168V) and Thr → Pro at amino acid 127 (T127P) were detected in HBsAg of the occult HBV strain. CONCLUSION These results indicate a relatively low prevalence of HBV infection among pregnant women in the South of Iran, while tattooing is a risk factor for exposure to HBV infection. Moreover, all of the HBV-positive pregnant women were asymptomatic and unaware of their infection. Therefore, routine screening for HBV markers during pregnancy, appropriate treatment of HBV-infected women, and HBV vaccination are recommended to decrease mother-to-child transmission of HBV.
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Affiliation(s)
- Reza Taherkhani
- Department of Virology, School of Medicine, Bushehr University of Medical Sciences, Bushehr, Iran
| | - Fatemeh Farshadpour
- Department of Virology, School of Medicine, Bushehr University of Medical Sciences, Bushehr, Iran
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11
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Kar P, Goswami B, Mahanta J, Bhimo T, Das AK, Deka M, Lynrah KG, Kotwal MR, Bhaumik P, Jini M, Karna R, Karra VK, Kaur H. Epidemiology, Genotyping, Mutational and Phylogenetic Analysis of Hepatitis B Virus Infection in North-east India. J Clin Exp Hepatol 2022; 12:43-51. [PMID: 35068784 PMCID: PMC8766538 DOI: 10.1016/j.jceh.2021.04.002] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 05/13/2020] [Accepted: 04/01/2021] [Indexed: 01/03/2023] Open
Abstract
BACKGROUND/OBJECTIVE Hepatitis B virus (HBV) infection is a major public health problem globally. Northeast India is home to indigenous tribes with different ethnicity and high rates of drug abuse and HIV infection. The study was designed to estimate the burden of HBV infection across various spectrums of liver diseases from this region. HBV genotypes and subgenotypes play a role in the chronicity of disease, response to treatment and its progression. As very limited data are available from this region, we tried to elucidate the role of HBV genotypes, HBV mutants and their phylogenetic analysis. METHOD We designed a prospective multicentric study, and included 7464 liver disease cases, 7432 blood donors and 650 health care workers, who were screened for HBV infection. HBV DNA positive patients were genotyped and subjected to surface protein, precore and core mutation and phylogenetic analysis. RESULTS The prevalence of HBV infection with respect to different types of liver diseases, blood donors and health care workers was 9.9% (1550/15,546). 49.5% (768/1550) cases were found to be HBV DNA positive. The most common genotype was found to be genotype D 74.2% (570/768), followed by genotype C 6.5% (50/768), A 4.4% (34/768) and I 0.9% (7/768). CONCLUSION This study highlights the high hepatitis B burden in Northeast India, reflecting lacunae in health care needs of the region. Also, the different genotype distribution and presence of mutations may translate into different rates of liver disease progression, prognosis and ultimately, clinical significance. However, further prospective cohort study from Northeast India is warranted, to elucidate the clinical significance of multiple genotypes and mutation in this unique population.
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Key Words
- AFP, alpha fetoprotein
- ALT, alanine transaminase
- AVH, acute viral hepatitis
- BCP, basal core promoter mutations
- CAH: chronic active hepatitis, CHB: chronic hepatitis B
- CLD, chronic liver disease
- DNA, deoxyribose nucleic acid
- EASL, European Association for the study of the liver
- FHF, fulminant hepatic failure
- FNAC, fine needle aspiration cytology
- HBV
- HBV, hepatitis B virus
- HBcAg, icosahedral core
- HBsAg, surface proteins
- HCC, hepatocellular carcinoma
- PCR, polymerase chain reaction
- RT, reverse transcriptase
- SGOT, serum glutamic oxaloacetic transaminase
- SGPT, serum glutamic pyruvic transaminase
- SHB, small hepatitis B surface antigen
- ULN, upper limit of normal
- epidemiology
- evolution
- genotype
- mutation
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Affiliation(s)
- Premashis Kar
- Maulana Azad Medical College, University of Delhi, New Delhi, India,Address for correspondence. Premashis Kar, Director Professor of Medicine Maulana Azad Medical College, University of Delhi, New Delhi, India.
| | - Bhabadev Goswami
- Department of Gastroenterology, Gauhati Medical College, Guwahati, Assam, India
| | | | - Thngam Bhimo
- Department of Medicine, Regional Institute of Medical Sciences, Regional Medical College, Imphal, Manipur, India
| | - Anup K. Das
- Department of Medicine, Assam Medical College, Dibrugarh, Assam, India
| | - Manab Deka
- Department of Biotechnology, Gauhati University, Assam, India
| | | | | | - Pradip Bhaumik
- Department of Medicine, Agartala Govt. Medical College, Agartala, Tripura, India
| | - Moji Jini
- General Hospital, Naharlagun, Arunachal Pradesh, India
| | - Rahul Karna
- Maulana Azad Medical College, University of Delhi, New Delhi, India
| | - Vijay K. Karra
- Division of Epidemiology and Communicable Diseases, Indian Council of Medical Research, Department of Health Research, New Delhi, India
| | - Harpreet Kaur
- Division of Epidemiology and Communicable Diseases, Indian Council of Medical Research, Department of Health Research, New Delhi, India
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12
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Wang ML, Liao J, Ye F, Tao YC, Wu DB, He M, Tang H, Chen EQ. Distribution and factors associated with serum HBV pregenomic RNA levels in Chinese chronic hepatitis B patients. J Med Virol 2021; 93:3688-3696. [PMID: 32949174 DOI: 10.1002/jmv.26529] [Citation(s) in RCA: 7] [Impact Index Per Article: 1.8] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/11/2020] [Revised: 09/04/2020] [Accepted: 09/17/2020] [Indexed: 02/05/2023]
Abstract
Correlations between serum hepatitus B virus (HBV) pregenomic RNA (pgRNA), hepatitus B surface antigen (HBsAg), and hepatitus B core-related antigen (HBcrAg) levels, and influencing factors of serum HBV pgRNA levels in Chinese chronic hepatitis B (CHB) patients are rarely reported. This was a retrospective cohort study consisting of 204 outpatients with CHB. Serum levels of HBV pgRNA, HBsAg, and HBcrAg were quantitative measured in frozen blood samples. The linear regression and multivariate logistic regression analysis were performed to determine associated factors of serum HBV pgRNA levels. In this cohort, the median serum HBV pgRNA level was 4.12 log10 copies/ml and 33.33% (68/204) of them had serum HBV pgRNA under low limit of detection (LLD) (<500 copies/ml); and the percentage of patients with serum HBV pgRNA under LLD in hepatitis B e antigen (HBeAg)-positive patients was significantly lower than that in HBeAg-negative patients (15.75% [23/46] vs. 77.59% [45/58], p < .001). Overall, serum HBV pgRNA strongly correlated with HBcrAg (r = 0.760, p < .001), and moderately correlated with HBV DNA (r = 0.663, p < .001) and HBsAg (r = 0.670, p < .001). As compared with HBsAg and HBV DNA, only HBcrAg showed stable correlation with serum HBV pgRNA both in HBeAg-positive and HBeAg-negative patients. Serum HBV pgRNA level differed between HBeAg-positive and HBeAg-negative patients; and it had better and more stable correlation with serum HBcrAg than serum HBV DNA and HBsAg, irrespective of HBeAg status.
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Affiliation(s)
- Meng-Lan Wang
- Center of Infectious Diseases, West China Hospital of Sichuan University, Chengdu, China
| | - Juan Liao
- Center of Infectious Diseases, West China Hospital of Sichuan University, Chengdu, China
| | - Feng Ye
- Beijing GenomePrecision Technology Co., Ltd., Beijing, China
| | - Ya-Chao Tao
- Center of Infectious Diseases, West China Hospital of Sichuan University, Chengdu, China
| | - Dong-Bo Wu
- Center of Infectious Diseases, West China Hospital of Sichuan University, Chengdu, China
| | - Min He
- Center of Infectious Diseases, West China Hospital of Sichuan University, Chengdu, China
| | - Hong Tang
- Center of Infectious Diseases, West China Hospital of Sichuan University, Chengdu, China
| | - En-Qiang Chen
- Center of Infectious Diseases, West China Hospital of Sichuan University, Chengdu, China
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13
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Liu H, Wan Z, She L, Zhu Y, Cai Z, Wu B, Zhuang Q, Ke P, Wu X, Li Z, Huang X. Inflammation Pharmacological Reaction and YMDD Mutational Patterns in Lamivudine Therapeutics Hepatitis B Virus. Front Pharmacol 2021; 12:648170. [PMID: 33935748 PMCID: PMC8081950 DOI: 10.3389/fphar.2021.648170] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/31/2020] [Accepted: 02/19/2021] [Indexed: 02/05/2023] Open
Abstract
Background/Aims: Emergence of tyrosine-methionine-aspartate-aspartate (YMDD) motif in reverse transcriptase is a serious problem in chronic hepatitis B(CHB) patients after Lamivudine (LAM) therapy. However, the relationship between inflammation pharmacological reaction and YMDD mutational patterns of CHB has not been well-characterized. The aim of this study was to investigate the inflammation pharmacological reaction and different YMDD mutants patterns of CHB patients. Methods: We investigated the inflammation pharmacological reaction and YMDD mutational patterns through biochemical, serological and virological detection among 83 CHB patients, including 25 YMDD mutants, 25 under detection, and 33 control patients without YMDD mutants. Results: Prevalence of YMDD mutation patterns is different. Among 25 YMDD mutants patients, YIDD was the dominant mutation (72%), followed YVDD (16%) and the hybrid YIDD + YVDD (12%). The time course during the YMDD mutations was also different. 52.4% patients developed the mutation less than 12 months after the LAM therapy. Serum hepatitis B virus (HBV) DNA level in patients with YMDD mutants were significantly higher than that in control and negative groups. Serum HbsAg and HbeAg in patients with YMDD mutants were also higher than those in control and negative groups, despite no significant difference was found forserum HbeAb. ALT and AST levels were also significantly higher in mutants group. Conclusions: Illuminating inflammation pharmacological reaction and YMDD mutational patterns of CHB during pathological process may have implications for future therapy in YMDD mutation patients. This may have impact on the choice of treatment strategies for lamivudine-resistant HBV.
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Affiliation(s)
- Hongcan Liu
- Department of Laboratory Medicine, The Second Affiliated Hospital of Guangzhou University of Chinese Medicine, Guangzhou, China
- *Correspondence: Hongcan Liu, ; Zhuo Li, ; Xianzhang Huang,
| | - Zemin Wan
- Department of Laboratory Medicine, The Second Affiliated Hospital of Guangzhou University of Chinese Medicine, Guangzhou, China
| | - Lanhui She
- Department of Infectious Diseases, Guangzhou Women and Children’s Medical Center, Guangzhou, China
| | - Yajuan Zhu
- Department of Ultrasound, The First Affiliated Hospital of USTC, Division of Life Sciences and Medicine, University of Science and Technology of China, Hefei, China
| | - Zhiliang Cai
- Department of Laboratory Medicine, The Second Affiliated Hospital of Guangzhou University of Chinese Medicine, Guangzhou, China
| | - Bin Wu
- Genetic Testing Lab, The Second Affiliated Hospital of Guangzhou University of Chinese Medicine, Guangzhou, China
| | - Qizhen Zhuang
- Department of Laboratory Medicine, The Second Affiliated Hospital of Guangzhou University of Chinese Medicine, Guangzhou, China
| | - Peifeng Ke
- Department of Laboratory Medicine, The Second Affiliated Hospital of Guangzhou University of Chinese Medicine, Guangzhou, China
| | - Xinzhong Wu
- Department of Laboratory Medicine, The Second Affiliated Hospital of Guangzhou University of Chinese Medicine, Guangzhou, China
| | - Zhuo Li
- Genetic Testing Lab, The Second Affiliated Hospital of Guangzhou University of Chinese Medicine, Guangzhou, China
- *Correspondence: Hongcan Liu, ; Zhuo Li, ; Xianzhang Huang,
| | - Xianzhang Huang
- Department of Laboratory Medicine, The Second Affiliated Hospital of Guangzhou University of Chinese Medicine, Guangzhou, China
- *Correspondence: Hongcan Liu, ; Zhuo Li, ; Xianzhang Huang,
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14
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de Almeida Pondé RA. Dynamic profile of the HBeAg-anti-HBe system in acute and chronic hepatitis B virus infection: A clinical-laboratory approach. Mol Biol Rep 2020; 48:843-854. [PMID: 33296069 DOI: 10.1007/s11033-020-06056-4] [Citation(s) in RCA: 9] [Impact Index Per Article: 1.8] [Reference Citation Analysis] [Abstract] [Key Words] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/17/2020] [Accepted: 12/01/2020] [Indexed: 02/08/2023]
Abstract
Wild-type HBV infection is followed by the blood expression of its widely known serological markers of infection, and designated as, hepatitis B virus surface antigen (HBsAg) and its antibody (anti-HBs), anti-HBc antibodies (IgM/IgG), and hepatitis B virus 'e' antigen (HBeAg) and its antibody (anti-HBe). These markers are detected as the infection develops and its kinetic behavior serves as a basis for monitoring the disorder and for diagnosing the clinical form or infection phase. Among these, the HBeAg-anti-HBe system markers demonstrate a dynamic profile whose interpretation, both in the acute or chronic HBV infection context, can offer greater difficulty to the health professionals, due to its particularities. This review offers a revisit to the markers dynamics of this system in the acute and chronic HBV infection and to the clinical and laboratory significance of its expression in these two clinical contexts.
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Affiliation(s)
- Robério Amorim de Almeida Pondé
- Laboratory of Human Virology, Institute of Tropical Pathology and Public Health, Federal University of Goiás, Goiânia, Goiás, Brazil. .,Secretaria de Estado da Saúde -SES/Superintendência de Vigilância em Saúde-SUVISA/GO, Gerência de Vigilância Epidemiológica-GVE/Coordenação de Análises e Pesquisas-CAP, Goiânia, Goiás, Brazil.
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15
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Li MR, Xu ZG, Lu JH, Zheng HW, Ye LH, Liu YY, Liu ZQ, Zhang HC, Huang Y, Dai EH, Pan CQ. Clinical features of hepatitis B patients at immune-tolerance phase with basal core promoter and/or precore mutations. J Viral Hepat 2020; 27:1044-1051. [PMID: 32384194 DOI: 10.1111/jvh.13315] [Citation(s) in RCA: 4] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 12/16/2019] [Revised: 03/27/2020] [Accepted: 04/18/2020] [Indexed: 12/09/2022]
Abstract
Little data exist on basal core promoter/precore (BCP/PC) mutations in chronic hepatitis B (CHB) patients at the immune-tolerance (IT) phase. We studied consecutive treatment-naïve, CHBe-antigen (HBeAg)-positive patients who had undergone liver biopsy and genotyping. Those in the IT phase or immune-clearance (IC) phase were enrolled for comparison of the frequency of BCP/PC mutations and their clinical presentations. Subgroup analyses for the IT group were also performed between patients with and without mutations, and IC patients between fibrosis stages ≤2 vs fibrosis >2. Among 301 patients enrolled, 88/301 (29.24%) and 213/301 (70.76%) were at the IT and IC phase, respectively. The frequency of BCP/PC mutations in IT phase was significantly lower than those in IC phase (15.91% vs 64.79%, P < .001). The BCP mutation only was significantly more frequent than the PC mutation in both groups and also in all IC subgroups. IT patients with BCP/PC mutations had significantly higher quantitative anti-HBc levels compared with those of patients with wild-type virus (P < .05). They also had significantly lower mean levels of alanine transaminase, aspartate transaminase, total bilirubin and qAnti-HBc compared with those of IC patients (all P < .05). Additionally, they were significantly younger in mean age, had higher platelet count, higher levels of HBV DNA and surface antigen, as well as higher frequency of genotype B than those of IC patients with fibrosis >2 (all P < .05). BCP/PC mutations were found in IT patients with CHB. They had distinct clinical characteristics when compared with patients with wild-type or at IC phase. Further studies are needed to understand their natural history and treatment outcomes.
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Affiliation(s)
- Min-Ran Li
- Division of Infectious Disease, The First Affiliated Hospital of Jinan University, Guangzhou, China
- Division of Liver Disease, The Fifth Hospital of Shijiazhuang, Hebei Medical University, Shijiazhuang, China
| | - Zun-Gui Xu
- Division of Liver Disease, The Fifth Hospital of Shijiazhuang, Hebei Medical University, Shijiazhuang, China
| | - Jian-Hua Lu
- Division of Liver Disease, The Fifth Hospital of Shijiazhuang, Hebei Medical University, Shijiazhuang, China
| | - Huan-Wei Zheng
- Division of Liver Disease, The Fifth Hospital of Shijiazhuang, Hebei Medical University, Shijiazhuang, China
| | - Li-Hong Ye
- Division of Liver Disease, The Fifth Hospital of Shijiazhuang, Hebei Medical University, Shijiazhuang, China
| | - Yun-Yan Liu
- Division of Liver Disease, The Fifth Hospital of Shijiazhuang, Hebei Medical University, Shijiazhuang, China
| | - Zhi-Quan Liu
- Division of Liver Disease, The Fifth Hospital of Shijiazhuang, Hebei Medical University, Shijiazhuang, China
| | - Hai-Cong Zhang
- Division of Liver Disease, The Fifth Hospital of Shijiazhuang, Hebei Medical University, Shijiazhuang, China
| | - Yan Huang
- Division of Liver Disease, The Fifth Hospital of Shijiazhuang, Hebei Medical University, Shijiazhuang, China
| | - Er-Hei Dai
- Division of Liver Disease, The Fifth Hospital of Shijiazhuang, Hebei Medical University, Shijiazhuang, China
| | - Calvin Q Pan
- Center of Liver Diseases, Beijing Ditan Hospital, Capital Medical Univerisity, Beijing, China
- Division of Gastroenterology and Hepatology, Department of Medicine, NYU Langone Health, New York University School of Medicine, New York, NY, USA
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16
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Verbinnen T, Tan Y, Wang G, Dehertogh P, Vergauwen K, Neefs JM, Jacoby E, Lenz O, Berke JM. Anti-HBV activity of the HBV capsid assembly modulator JNJ-56136379 across full-length genotype A–H clinical isolates and core site-directed mutants in vitro. J Antimicrob Chemother 2020; 75:2526-2534. [DOI: 10.1093/jac/dkaa179] [Citation(s) in RCA: 10] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/27/2019] [Revised: 04/03/2020] [Accepted: 04/09/2020] [Indexed: 12/16/2022] Open
Abstract
Abstract
Objectives
To characterize antiviral activity of the capsid assembly modulator (CAM-N) JNJ-56136379 against HBV genotypes and variants carrying amino acid substitutions in the core protein.
Methods
Anti-HBV activity of JNJ-56136379 was investigated against a diverse panel of 53 HBV clinical isolates (genotypes A–H). The impact of core amino acid substitutions using site-directed mutants (SDMs) was assessed in a transient replication assay.
Results
JNJ-56136379 median 50% effective concentration (EC50) values across all genotypes were 10–33 nM versus 17 nM (genotype D reference). JNJ-56136379 remained active against isolates carrying nucleos(t)ide analogue resistance mutations (median EC50 2–25 nM) or basal core promoter (BCP) ± precore (PC) mutations (median EC50 13–20 nM) or PC mutations (median EC50 11 nM), representing activity against isolates from HBeAg-positive and -negative hepatitis B patients. Core amino acid substitutions in the CAM-binding pocket, when tested as SDMs at positions 23, 25, 30, 33, 37, 106, 110, 118, 124, 127 and 128, reduced JNJ-56136379 anti-HBV activity; EC50 fold increases ranged from 3.0 (S106T) to 85 (T33N). All substitutions were rare in a public database of >7600 HBV core sequences (frequencies 0.01%–0.3%). Nucleos(t)ide analogues retained full activity against these core SDMs.
Conclusions
JNJ-56136379, a potent HBV CAM-N, currently in Phase 2 clinical development, was generally fully active against an extensive panel of genotype A–H clinical isolates, regardless of the presence of nucleos(t)ide analogue resistance or BCP/PC mutations. JNJ-56136379 activity was reduced by some core amino acid substitutions in the CAM-binding pocket.
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Affiliation(s)
- Thierry Verbinnen
- Janssen Research and Development, Turnhoutseweg 30, 2340 Beerse, Belgium
| | - Ying Tan
- Janssen China Research & Development Center, 5F North Building #1 Jinchuang Mansion, 4560 Jinke Road, Shanghai 201210, China
| | - Gengyan Wang
- Janssen China Research & Development Center, 5F North Building #1 Jinchuang Mansion, 4560 Jinke Road, Shanghai 201210, China
| | - Pascale Dehertogh
- Janssen Research and Development, Turnhoutseweg 30, 2340 Beerse, Belgium
| | - Karen Vergauwen
- Janssen Research and Development, Turnhoutseweg 30, 2340 Beerse, Belgium
| | - Jean-Marc Neefs
- Janssen Research and Development, Turnhoutseweg 30, 2340 Beerse, Belgium
| | - Edgar Jacoby
- Janssen Research and Development, Turnhoutseweg 30, 2340 Beerse, Belgium
| | - Oliver Lenz
- Janssen Research and Development, Turnhoutseweg 30, 2340 Beerse, Belgium
| | - Jan Martin Berke
- Janssen Research and Development, Turnhoutseweg 30, 2340 Beerse, Belgium
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17
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Di Bisceglie AM, King WC, Lisker-Melman M, Khalili M, Belle SH, Feld JJ, Ghany MG, Janssen HL, Lau D, Lee WM, Ling SC, Cooper S, Rosenthal P, Schwarz KB, Sterling RK, Teckman JH, Terrault N, Hepatitis B Research Network (HBRN). Age, race and viral genotype are associated with the prevalence of hepatitis B e antigen in children and adults with chronic hepatitis B. J Viral Hepat 2019; 26:856-865. [PMID: 30974509 PMCID: PMC6592737 DOI: 10.1111/jvh.13104] [Citation(s) in RCA: 14] [Impact Index Per Article: 2.3] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 01/28/2019] [Revised: 03/07/2019] [Accepted: 03/15/2019] [Indexed: 12/23/2022]
Abstract
Hepatitis B e antigen (HBeAg) is an important serological marker of hepatitis B virus (HBV) infection and is associated with higher levels of viraemia, increased risk of infectivity to others and increased risk of hepatocellular carcinoma. We analysed HBeAg status in a large cohort of adults and children enrolled in Cohort Studies of the Hepatitis B Research Network, long-term natural history studies of chronic HBV infection. A cross-sectional analysis examined factors associated with HBeAg positivity, including demographic and virologic data, across the age spectrum. Among 2241 enrolled participants who met criteria for this analysis, 825 (37%) were seropositive for HBeAg. The prevalence of HBeAg was lower in those with older age, ranging from 85% among those up to 10 years of age to only 12% among those older than 50 years. In addition to age, both race and HBV genotype were independently associated with HBeAg positivity. There was a significant interaction between age and race; the prevalence of HBeAg was significantly higher among Asians > 10-30 years old vs Whites or Blacks who were >10 to 30 years old and those infected with HBV genotype C. Conversely, the presence of the basal core promoter and precore variants was associated with significantly lower prevalence of HBeAg, even when adjusted for age, race and genotype. These data will provide a better understanding of factors associated with seropositivity for HBeAg and may lead to better strategies for preventing HBV infection and broader indications for antiviral therapy.
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Affiliation(s)
| | - Wendy C. King
- Department of Epidemiology, Graduate School of Public Health, University of Pittsburgh
| | | | - Mandana Khalili
- University of California at San Francisco, Toronto General Hospital, University of Toronto
| | - Steven H. Belle
- Department of Epidemiology, Graduate School of Public Health, University of Pittsburgh
| | - Jordan J. Feld
- Toronto Centre for Liver Disease, Toronto General Hospital, University of Toronto
| | - Marc G. Ghany
- Liver Diseases Branch, National Institute of Diabetes and Digestive and Kidney Diseases
| | - Harry L.A. Janssen
- Toronto Centre for Liver Disease, Toronto General Hospital, University of Toronto
| | - Daryl Lau
- Beth Israel Deaconess Medical Center
| | | | - Simon C. Ling
- Toronto Centre for Liver Disease, Toronto General Hospital, University of Toronto
| | | | - Philip Rosenthal
- University of California at San Francisco, Toronto General Hospital, University of Toronto
| | | | | | | | - Norah Terrault
- University of California at San Francisco, Toronto General Hospital, University of Toronto
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18
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Torres MC, Civetta E, D'amico C, Barbini L. Hepatitis B virus in Mar del Plata, Argentina: Genomic characterization and evolutionary analysis of subgenotype F1b. J Med Virol 2019; 91:791-802. [PMID: 30570771 DOI: 10.1002/jmv.25383] [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: 07/13/2018] [Accepted: 12/14/2018] [Indexed: 11/07/2022]
Abstract
The aim is to describe the molecular epidemiology and perform a genomic characterization of hepatitis B virus (HBV) circulating in Mar del Plata and to identify the origin and diversification patterns of the most prevalent genotype. The S gene and the region encompassing the X gene, basal core promoter (BCP), and precore (preC) was analyzed in 56 samples. They were genotyped as: 80% F1b, 9% A2, 7% D3, and 2% D1. A recombinant F4/D2 genome was detected. The double substitution G1764A/A1762T at the BCP (reduced HBeAg expression) was found in 20% F1b, 2% A2, 2% D1, and 2% D3 samples. A unique D3 presented the G1896A substitution at the preC (HBeAg negative phenotype). A 13% of the samples showed mutations at the HBsAg "a" immunodeterminant (escape from neutralizing antibodies). Mutations at the polymerase (antiviral resistance) were found in 52% of the samples. Coalescent analysis of subgenotype F1b, the most prevalent in the city, showed that viral diversification in Mar del Plata started by year 2000. F1b was the most prevalent genotype detected, being a characteristic of actual HBV infections in Mar del Plata. Local HBV exhibit clinically relevant mutations, but a minority of them was shown to be associated to potential vaccination escape or antiviral resistance. Nevertheless, further studies are needed to determine whether any of these mutants could pose a threat to prevention, diagnosis, or treatment.
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Affiliation(s)
| | - Elida Civetta
- Unidad de Hepatología y Alcoholismo, HIGA Dr. O. Alende, Mar del Plata, Argentina
| | - Claudia D'amico
- Centro de Especialidades Médicas Ambulatorias, Unidad de Hepatología, Mar del Plata, Argentina
| | - Luciana Barbini
- Departamento de Química, FCEyN, UNMdP, Buenos Aires, Argentina
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Yeh CSH, Hsu CW, Liang KH, Chen YC, Lin CL, Chien RN, Hu TH, Lin WR, Lai MW, Chu YD, Yeh CT. Development of a fibrosis index including hepatitis B virus basal core promoter A1762T mutation for pretherapeutic evaluation. J Gastroenterol Hepatol 2018; 33:1530-1537. [PMID: 29424069 DOI: 10.1111/jgh.14120] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 12/13/2017] [Revised: 01/22/2018] [Accepted: 01/24/2018] [Indexed: 12/15/2022]
Abstract
BACKGROUND AND AIM Commonly used non-invasive fibrosis scores usually included serum transaminase levels in the equations, including Aspartate transaminase to Platelet Ratio Index (APRI) and fibrosis-4 (FIB-4). Transaminases fluctuated significantly in chronic hepatitis B patients with exacerbations, leading to unsteady score values. As such, here, we aim to develop a transaminase-free score suitable for pretherapeutic evaluation of fibrosis stages. METHODS Firstly, 1082 treatment-naïve chronic hepatitis B patients were enrolled and divided into modeling (n = 541) and verification (n = 541) cohorts. Secondly, 265 patients having received liver biopsy, with known Ishak fibrosis stages, were included for independent correlation. RESULTS Cross-sectional analysis of 1082 patients revealed age-dependent variation of association between virological factors and cirrhosis. A fibrosis score including Anti-hepatitis B e antibody, Basal core promoter (BCP) A1762T mutation, and Platelet count Index (named ABPI) was derived from the modeling cohort. ABPI performed better than APRI and FIB-4 in the verification cohort for identifying cirrhotic patients (comparison of area under the receiver operating characteristic curves: ABPI vs APRI and FIB-4 = 0.785 vs 0.563 [P < 0.001] and 0.700 [P = 0.026], respectively). The performance of ABPI was even better in young (< 40 years old) hepatitis B patients (area under the receiver operating characteristic curves: 0.856 vs 0.402 [P < 0.001] and 0.599 [P = 0.009], respectively). Finally, in the independent cohort of 265 patients with known Ishak fibrosis stages, it was found that ABPI effectively distinguished between Ishak fibrosis stages 3 and > 3 and between 4 and > 4 (P < 0.001 for each). CONCLUSIONS We developed a transaminase-free fibrosis score (ABPI) utilizing basal core promoter A1762T data, which outperformed APRI and FIB-4.
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Affiliation(s)
- Christopher Sung-Huan Yeh
- Liver Research Center, Chang Gung Memorial Hospital, Taoyuan, Taiwan
- Department of Cognitive Science, College of Letters and Science, University of California, California, Los Angeles, USA
| | - Chao-Wei Hsu
- Liver Research Center, Chang Gung Memorial Hospital, Taoyuan, Taiwan
- Department of Internal Medicine, Chang Gung University College of Medicine, Taoyuan, Taiwan
| | - Kung-Hao Liang
- Medical Research Department, Taipei Veterans General Hospital, Taipei, Taiwan
| | - Yi-Cheng Chen
- Liver Research Center, Chang Gung Memorial Hospital, Taoyuan, Taiwan
| | - Chih-Lang Lin
- Liver Research Unit, Chang Gung Memorial Hospital, Keelung, Taiwan
| | - Rong-Nan Chien
- Liver Research Unit, Chang Gung Memorial Hospital, Keelung, Taiwan
| | - Tsung-Hui Hu
- Division of Hepato-gastroenterology, Department of Internal Medicine, Kaohsiung Chang Gung Memorial Hospital, Kaohsiung, Taiwan
| | - Wey-Ran Lin
- Liver Research Center, Chang Gung Memorial Hospital, Taoyuan, Taiwan
| | - Ming-Wei Lai
- Liver Research Center, Chang Gung Memorial Hospital, Taoyuan, Taiwan
| | - Yu-De Chu
- Liver Research Center, Chang Gung Memorial Hospital, Taoyuan, Taiwan
| | - Chau-Ting Yeh
- Liver Research Center, Chang Gung Memorial Hospital, Taoyuan, Taiwan
- Molecular Medicine Research Center, Chang Gung University, Taoyuan, Taiwan
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Analysis of HBV basal core promoter/precore gene variability in patients with HBV drug resistance and HIV co-infection in Northwest Ethiopia. PLoS One 2018; 13:e0191970. [PMID: 29408943 PMCID: PMC5800642 DOI: 10.1371/journal.pone.0191970] [Citation(s) in RCA: 7] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/18/2017] [Accepted: 01/15/2018] [Indexed: 12/13/2022] Open
Abstract
Background We recently reported complex hepatitis B virus (HBV) drug resistant and concomitant vaccine escape hepatitis B surface antigen (HBsAg) variants during human immunodeficiency virus (HIV) co-infection and antiretroviral therapy (ART) exposure in Ethiopia. As a continuation of this report using the HBV positive sera from the same study participants, the current study further analyzed the HBV basal core promoter (BCP)/precore (PC) genes variability in patients with HBV drug resistance (at tyrosine-methionine-aspartate-aspartate (YMDD) reverse transcriptase (RT) motifs) and HIV co-infection in comparison with HBV mono-infected counterparts with no HBV drug resistant gene variants. Materials and methods A total of 143 participants of HBV-HIV co-infected (n = 48), HBV mono-infected blood donors (n = 43) and chronic liver disease (CLD) patients (n = 52) were included in the study. The BCP/PC genome regions responsible for HBeAg expression from the EcoRI site (nucleotides 1653–1959) were sequenced and analyzed for the BCP/PC mutant variants. Results Among the major mutant variants detected, double BCP mutations (A1762T/G1764A) (25.9%), Kozak sequences mutations (nt1809-1812) (51.7%) and the classical PC mutations such as A1814C/C1816T (15.4%), G1896A (25.2%) and G1862T (44.8%) were predominant mutant variants. The prevalence of the double BCP mutations was significantly lower in HIV co-infected patients (8.3%) compared with HBV mono-infected blood donors (32.6%) and CLD patients (36.5%). However, the Kozak sequences BCP mutations and the majority of PC mutations showed no significant differences among the study groups. Moreover, except for the overall BCP/PC mutant variants, co-prevalence rates of each major BCP/PC mutations and YMDDRT motif associated lamivudine (3TC)/entecavir (ETV) resistance mutations showed no significant differences when compared with the rates of BCP/PC mutations without YMDD RT motif drug resistance gene mutations. Unlike HIV co-infected group, no similar comparison made among HBV mono-infected blood donors and CLD patients since none of them developed the YMDD RT motif associated 3TC/ETV resistance mutations. However, HBV mono-infected blood donors and CLD patients who had no any drug resistance gene variants developed comparable G1862T (60.6% vs. 65.1%) and G1896A (24.2% vs. 11.6%) PC gene mutations. Conclusion No correlation observed between the BCP/PC genome variability and the YMDD RT motif associated HBV drug resistance gene variants during HIV co-infection. Nevertheless, irrespective of HIV co-infection status, the higher records of the BCP/PC gene variability in this study setting indicate a high risk of potential HBeAg negative chronic HBV infection in Northwest Ethiopia.
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Chen YL, Mo YQ, Zheng DH, Ma JD, Jing J, Dai L. Patients with Coexistence of Circulating Hepatitis B Surface Antigen and Its Antibody May Have a Strong Predisposition to Virus Reactivation During Immunosuppressive Therapy: A Hypothesis. Med Sci Monit 2017; 23:5980-5985. [PMID: 29248936 PMCID: PMC5742491 DOI: 10.12659/msm.905033] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.1] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/23/2022] Open
Abstract
Hepatitis B virus (HBV) reactivation is a well-recognized complication in patients who undergo immunosuppressive drug therapy. Although the recommendation of antiviral prophylaxis made by the American Gastroenterological Association in 2015 focuses on the risk stratification of different immunosuppressive drugs, risk factors for HBV reactivation are also worth identifying in clinical practice. Recent studies have shown that the uncommon serological pattern of coexistent circulating HBV surface antigen (HBsAg) and its antibody (anti-HBs) was associated with double mutations (A1762T/G1764A) in the basal core promoter (BCP) region of the HBV genome, which is critical for HBV replication. Here, we depicted rheumatoid arthritis (RA) patients with coexistent HBsAg and anti-HBs in our medical center, who developed HBV reactivation during immunosuppressive drug therapy. DNA sequencing analysis of the HBV genome revealed triple mutations (A1762T, G1764A, and T1753V) in the BCP region, which could further enhance the ability of HBV replication. Hence, a novel hypothesis is advanced for the first time that patients with coexistent HBsAg and anti-HBs may have a strong predisposition to HBV reactivation due to specific BCP mutations. This hypothesis would, if correct, justify the concurrent detection of HBsAg and anti-HBs in HBV screening in patients with rheumatic diseases and quickly recognize patients with high risk of HBV reactivation. Further controlled studies are needed to confirm this hypothesis.
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Affiliation(s)
- Yu-Lan Chen
- Department of Rheumatology, Sun Yat-Sen Memorial Hospital, Sun Yat-Sen University, Guangzhou, Guangdong, China (mainland)
| | - Ying-Qian Mo
- Department of Rheumatology, Sun Yat-Sen Memorial Hospital, Sun Yat-Sen University, Guangzhou, Guangdong, China (mainland)
| | - Dong-Hui Zheng
- Department of Rheumatology, Sun Yat-Sen Memorial Hospital, Sun Yat-Sen University, Guangzhou, Guangdong, China (mainland)
| | - Jian-Da Ma
- Department of Rheumatology, Sun Yat-Sen Memorial Hospital, Sun Yat-Sen University, Guangzhou, Guangdong, China (mainland)
| | - Jun Jing
- Department of Rheumatology, Sun Yat-Sen Memorial Hospital, Sun Yat-Sen University, Guangzhou, Guangdong, China (mainland)
| | - Lie Dai
- Department of Rheumatology, Sun Yat-Sen Memorial Hospital, Sun Yat-Sen University, Guangzhou, Guangdong, China (mainland)
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22
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Shirvani-Dastgerdi E, Winer BY, Celià-Terrassa T, Kang Y, Tabernero D, Yagmur E, Rodríguez-Frías F, Gregori J, Luedde T, Trautwein C, Ploss A, Tacke F. Selection of the highly replicative and partially multidrug resistant rtS78T HBV polymerase mutation during TDF-ETV combination therapy. J Hepatol 2017; 67:246-254. [PMID: 28392234 PMCID: PMC6016549 DOI: 10.1016/j.jhep.2017.03.027] [Citation(s) in RCA: 39] [Impact Index Per Article: 4.9] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 09/07/2016] [Revised: 03/01/2017] [Accepted: 03/16/2017] [Indexed: 02/07/2023]
Abstract
BACKGROUND & AIMS Patients chronically infected with the hepatitis B virus (HBV) and receiving long-term treatment with nucleoside or nucleotide analogues are at risk of selecting HBV strains with complex mutational patterns. We herein report two cases of HBV-infected patients with insufficient viral suppression, despite dual antiviral therapy with entecavir (ETV) and tenofovir (TDF). One patient died from aggressive hepatocellular carcinoma (HCC). METHODS Serum samples from the two patients at different time points were analyzed using ultra-deep pyrosequencing analysis. HBV mutations were identified and transiently transfected into hepatoma cells in vitro using replication-competent HBV vectors, and functionally analyzed. We assessed replication efficacy, resistance to antivirals and potential impact on HBV secretion (viral particles, exosomes). RESULTS Sequencing analyses revealed the selection of the rtS78T HBV polymerase mutation in both cases that simultaneously creates a premature stop codon at sC69 and thereby deletes almost the entire small HBV surface protein. One of the patients had an additional 261bp deletion in the preS1/S2 region. Functional analyses of the mutations in vitro revealed that the rtS78T/sC69∗ mutation, but not the preS1/S2 deletion, significantly enhanced viral replication and conferred reduced susceptibility to ETV and TDF. The sC69∗ mutation caused truncation of HBs protein, leading to impaired detection by commercial HBsAg assay, without causing intracellular HBsAg retention or affecting HBV secretion. CONCLUSIONS The rtS78T/sC69∗ HBV mutation, associated with enhanced replication and insufficient response to antiviral treatment, may favor long-term persistence of these isolates. In addition to the increased production of HBV transcripts and the sustained secretion of viral particles in the absence of antigenic domains of S protein, this HBV mutation may predispose patients to carcinogenic effects. LAY SUMMARY Long-term treatment with antiviral drugs carries the risk of selecting mutations in the hepatitis B virus (HBV). We herein report two cases of patients with insufficient response to dual tenofovir and entecavir therapy. Molecular analyses identified a distinct mutation, rtS78T/sC69∗, that abolishes HBsAg detection, enhances replication, sustains exosome-mediated virion secretion and decreases susceptibility to antivirals, thereby representing a potentially high-risk mutation for HBV-infected individuals.
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Affiliation(s)
- Elham Shirvani-Dastgerdi
- Department of Medicine III, RWTH-University Hospital Aachen, Aachen, Germany; Department of Molecular Biology, Princeton University, Princeton, NJ, USA
| | - Benjamin Y Winer
- Department of Molecular Biology, Princeton University, Princeton, NJ, USA
| | | | - Yibin Kang
- Department of Molecular Biology, Princeton University, Princeton, NJ, USA
| | - David Tabernero
- Centro de Investigación Biomédica en Red de Enfermedades Hepáticas y Digestivas (CIBERehd), Instituto de Salud Carlos III, Madrid, Spain; Liver Unit, Vall d'Hebron Institute of Research (VHIR), Universitat Autònoma de Barcelona (UAB), Barcelona, Spain
| | - Eray Yagmur
- Medical Care Centre, Dr Stein and Colleagues, Mönchengladbach, Germany
| | - Francisco Rodríguez-Frías
- Centro de Investigación Biomédica en Red de Enfermedades Hepáticas y Digestivas (CIBERehd), Instituto de Salud Carlos III, Madrid, Spain; Liver Unit, Vall d'Hebron Institute of Research (VHIR), Universitat Autònoma de Barcelona (UAB), Barcelona, Spain
| | | | - Tom Luedde
- Department of Medicine III, RWTH-University Hospital Aachen, Aachen, Germany
| | - Christian Trautwein
- Department of Medicine III, RWTH-University Hospital Aachen, Aachen, Germany
| | - Alexander Ploss
- Department of Molecular Biology, Princeton University, Princeton, NJ, USA
| | - Frank Tacke
- Department of Medicine III, RWTH-University Hospital Aachen, Aachen, Germany.
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23
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Hundie GB, Stalin Raj V, Gebre Michael D, Pas SD, Koopmans MP, Osterhaus ADME, Smits SL, Haagmans BL. A novel hepatitis B virus subgenotype D10 circulating in Ethiopia. J Viral Hepat 2017; 24:163-173. [PMID: 27808472 DOI: 10.1111/jvh.12631] [Citation(s) in RCA: 12] [Impact Index Per Article: 1.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 07/21/2016] [Accepted: 10/06/2016] [Indexed: 12/13/2022]
Abstract
Hepatitis B virus (HBV) is genetically highly divergent and classified in ten genotypes and forty subgenotypes in distinct ethno-geographic populations worldwide. Ethiopia is a country with high HBV prevalence; however, little is known about the genetic variability of HBV strains that circulate. Here, we characterize the complete genome of 29 HBV strains originating from five Ethiopian regions, by 454 deep sequencing and Sanger sequencing. Phylogenetically, ten strains were classified as genotype A1 and nineteen as genotype D. Fifteen genotype D strains, provisionally named subgenotype D10, showed a novel distinct cluster supported by high bootstrap value and >4% nucleotide divergence from other known subgenotypes. In addition, the novel D10 strains harboured nine unique amino acid signatures in the surface, polymerase and X genes. Seventy-two per cent of the genotype D strains had the precore premature stop codon G1896A. In addition, 63% genotype A and 33% genotype D strains had the basal core promoter mutations, A1762T/G1764A. Furthermore, four pre-S deletion variants and two recombinants were identified in this study. In conclusion, we identified a novel HBV subgenotype D10 circulating in Ethiopia, underlining the high genetic variability of HBV strains in Africa.
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Affiliation(s)
- G B Hundie
- Department of Viroscience, Erasmus Medical Center, Rotterdam, The Netherlands
| | - V Stalin Raj
- Department of Viroscience, Erasmus Medical Center, Rotterdam, The Netherlands
| | - D Gebre Michael
- National blood bank services, Ministry of Health, Addis Ababa, Ethiopia
| | - S D Pas
- Department of Viroscience, Erasmus Medical Center, Rotterdam, The Netherlands
| | - M P Koopmans
- Department of Viroscience, Erasmus Medical Center, Rotterdam, The Netherlands
| | - A D M E Osterhaus
- Artemis One health, Utrecht, The Netherlands.,Center for Infection Medicine and Zoonoses Research, University of Veterinary Medicine, Hannover, Germany
| | - S L Smits
- ViroClinics BioScience BV, Rotterdam, The Netherlands
| | - B L Haagmans
- Department of Viroscience, Erasmus Medical Center, Rotterdam, The Netherlands
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Peng Y, Li Y, Hou J, Sun J, Su M, Li Y, Xiang K, Yan L, Zhuang H, Li T. The nucleotide changes within HBV core promoter/precore during the first 12weeks of nucleos(t)ide treatment might be associated with a better virological response. INFECTION GENETICS AND EVOLUTION 2017; 49:116-121. [PMID: 28088502 DOI: 10.1016/j.meegid.2017.01.009] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.1] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Subscribe] [Scholar Register] [Received: 08/28/2016] [Revised: 01/05/2017] [Accepted: 01/06/2017] [Indexed: 02/08/2023]
Abstract
OBJECTIVES We aimed to study the dynamic changes of hepatitis B virus (HBV) core promoter/precore (CP/preC) sequences during antiviral treatment and their associations with virological responses. MATERIALS AND METHODS The baseline and 12-week CP/preC sequences (nts 1655-2014) were obtained from 52 chronic hepatitis B patients with positive hepatitis B e antigen (HBeAg), who received a 104-week lamivudine and adefovir dipivoxil combination therapy. The mutations within the CP/preC were analyzed against genotype specific reference sequences. The nucleotide change rates in individuals during therapy were analyzed in a pairwise comparison manner. RESULTS There was no significant difference of the mutation rate at each nucleotide site between baseline and week 12 of treatment (P>0.05). The mutation rates of A1762T/G1764A and G1896A were found to decrease from 46.2% (24/52) at baseline to 36.5% (19/52) at week 12 (P=0.426) and from 28.8% (15/52) to 21.2% (11/52) (P=0.497), respectively. The nucleotide change rates varied from 0.0% - 7.8% in individuals [0.0% in Group 1 (N=26); 0.3% - 7.8% in Group 2 (N=26)] during the first 12-week treatment. HBV DNA levels in Group 2 were significantly lower than those in Group 1 throughout therapy (P<0.01) (e.g., 1.5±1.3log10 IU/ml vs. 2.6±1.0log10 IU/ml at week 104, P=0.001). At week 104 the rates of HBV DNA undetectable and HBeAg loss in Group 2 were significantly higher than those in Group 1 (P<0.05). Along with the increased nucleotide change rates, the rate of HBV DNA undetectable at week 104 tended to increase (odds ratio=0.323, 95% confidence interval=0.138-0.758, P<0.001). CONCLUSION Our findings suggested that the nucleotide changes within HBV CP/preC region during the first 12-week treatment might be associated with a better virological response.
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Affiliation(s)
- Yaqin Peng
- Department of Microbiology and Infectious Disease Center, School of Basic Medical Sciences, Peking University Health Science Center, Beijing 100191, China; Department of Clinical Laboratory, The First Affiliated Hospital, Sun Yat-sen University, Guangzhou 510080, China
| | - Yutang Li
- Department of Microbiology and Infectious Disease Center, School of Basic Medical Sciences, Peking University Health Science Center, Beijing 100191, China
| | - Jinlin Hou
- Department of Infectious Diseases and Hepatology Unit, Nanfang Hospital, Southern Medical University, Guangzhou 510515, China
| | - Jian Sun
- Department of Infectious Diseases and Hepatology Unit, Nanfang Hospital, Southern Medical University, Guangzhou 510515, China
| | - Mingze Su
- Department of Microbiology and Infectious Disease Center, School of Basic Medical Sciences, Peking University Health Science Center, Beijing 100191, China
| | - Yao Li
- Department of Microbiology and Infectious Disease Center, School of Basic Medical Sciences, Peking University Health Science Center, Beijing 100191, China
| | - Kuanhui Xiang
- Department of Microbiology and Infectious Disease Center, School of Basic Medical Sciences, Peking University Health Science Center, Beijing 100191, China
| | - Ling Yan
- Department of Microbiology and Infectious Disease Center, School of Basic Medical Sciences, Peking University Health Science Center, Beijing 100191, China
| | - Hui Zhuang
- Department of Microbiology and Infectious Disease Center, School of Basic Medical Sciences, Peking University Health Science Center, Beijing 100191, China
| | - Tong Li
- Department of Microbiology and Infectious Disease Center, School of Basic Medical Sciences, Peking University Health Science Center, Beijing 100191, China.
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Li Y, Ito M, Sun S, Chida T, Nakashima K, Suzuki T. LUC7L3/CROP inhibits replication of hepatitis B virus via suppressing enhancer II/basal core promoter activity. Sci Rep 2016; 6:36741. [PMID: 27857158 PMCID: PMC5114668 DOI: 10.1038/srep36741] [Citation(s) in RCA: 14] [Impact Index Per Article: 1.6] [Reference Citation Analysis] [Abstract] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/27/2016] [Accepted: 10/20/2016] [Indexed: 02/08/2023] Open
Abstract
The core promoter of hepatitis B virus (HBV) genome is a critical region for transcriptional initiation of 3.5 kb, pregenome and precore RNAs and for the viral replication. Although a number of host-cell factors that potentially regulate the viral promoter activities have been identified, the molecular mechanisms of the viral gene expression, in particular, regulatory mechanisms of the transcriptional repression remain elusive. In this study, we identified LUC7 like 3 pre-mRNA splicing factor (LUC7L3, also known as hLuc7A or CROP) as a novel interacting partner of HBV enhancer II and basal core promoter (ENII/BCP), key elements within the core promoter, through the proteomic screening and found that LUC7L3 functions as a negative regulator of ENII/BCP. Gene silencing of LUC7L3 significantly increased expression of the viral genes and antigens as well as the activities of ENII/BCP and core promoter. In contrast, overexpression of LUC7L3 inhibited their activities and HBV replication. In addition, LUC7L3 possibly contributes to promotion of the splicing of 3.5 kb RNA, which may also be involved in negative regulation of the pregenome RNA level. This is the first to demonstrate the involvement of LUC7L3 in regulation of gene transcription and in viral replication.
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Affiliation(s)
- Yuan Li
- Department of Virology and Parasitology, Hamamatsu University School of Medicine, Shizuoka 431-3192, Japan
| | - Masahiko Ito
- Department of Virology and Parasitology, Hamamatsu University School of Medicine, Shizuoka 431-3192, Japan
| | - Suofeng Sun
- Department of Virology and Parasitology, Hamamatsu University School of Medicine, Shizuoka 431-3192, Japan
| | - Takeshi Chida
- Department of Virology and Parasitology, Hamamatsu University School of Medicine, Shizuoka 431-3192, Japan
| | - Kenji Nakashima
- Department of Virology and Parasitology, Hamamatsu University School of Medicine, Shizuoka 431-3192, Japan
| | - Tetsuro Suzuki
- Department of Virology and Parasitology, Hamamatsu University School of Medicine, Shizuoka 431-3192, Japan
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Tacke F, Kroy DC. Treatment for hepatitis B in patients with drug resistance. ANNALS OF TRANSLATIONAL MEDICINE 2016; 4:334. [PMID: 27761438 DOI: 10.21037/atm.2016.09.19] [Citation(s) in RCA: 20] [Impact Index Per Article: 2.2] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 12/19/2022]
Abstract
Persistent hepatitis B virus (HBV) infections affect about 240 million patients worldwide that are at risk of developing liver cirrhosis or hepatocellular carcinoma. HBV is a small, partially double stranded DNA virus with four overlapping genes and a unique life cycle, which involves the generation of an RNA template for replication via reverse transcription. Mutations occur frequently during chronic infection, and particular selection pressures select distinct mutants. Nucleoside and nucleotide analogues like lamivudine (LMV), entecavir (ETV), telbivudine (LdT), adefovir dipivoxil (ADV) and tenofovir (TDF) are used to achieve long-term suppression of viral replication. Importantly, these drugs have different barriers to resistance, explaining the higher incidence of treatment failure in the past due to drug resistant viral strains for the older compounds LMV, LdT and ADV. On a molecular level, drug resistant mutations usually affect the reverse transcriptase domain of the HBV polymerase protein. Secondary compensatory mutations restore the replication fitness of the mutant virus. From a clinical point of view, patients undergoing antiviral therapy require regular testing for HBV DNA (every 3-6 months). In case of insufficient viral suppression or viral breakthrough (>1 log increase in HBV DNA above nadir), strict adherence to therapy needs to be ensured. If drug resistance is suspected or even molecularly confirmed, rescue therapy strategies exist, usually switching to a noncross-resistant antiviral drug. LMV, LdT and ETV resistant HBV can be treated with TDF monotherapy, ADV resistance with ETV or TDF, and insufficient responses to TDF may require ETV either as mono- or combination therapy. Complex treatment histories with many antivirals may sometimes necessitate the combination of highly effective antivirals like ETV and TDF. Novel treatment targets such as core (capsid) inhibitors, siRNA targeting protein translation, entry inhibitors or immune modulators aim at improving the efficacy of antivirals in order to (functionally) cure hepatitis B.
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Affiliation(s)
- Frank Tacke
- Department of Medicine III, RWTH Aachen University Hospital, Aachen, Germany
| | - Daniela C Kroy
- Department of Medicine III, RWTH Aachen University Hospital, Aachen, Germany
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27
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Zhong YW, Di FL, Liu C, Zhang XC, Bi JF, Li YL, Wu SQ, Dong H, Liu LM, He J, Shi YM, Zhang HF, Zhang M. Hepatitis B virus basal core promoter/precore mutants and association with liver cirrhosis in children with chronic hepatitis B virus infection. Clin Microbiol Infect 2015; 22:379.e1-379.e8. [PMID: 26577140 DOI: 10.1016/j.cmi.2015.10.033] [Citation(s) in RCA: 8] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/12/2015] [Revised: 10/29/2015] [Accepted: 10/30/2015] [Indexed: 02/08/2023]
Abstract
We investigated 168 children and analysed the virological characterization and association with disease progression in children with hepatitis B virus (HBV) basal core promoter/precore (BCP/PC) mutants. Among 168 patients with HBV infection (aged 0.5-18 years old, mean 10.1), 86 of them had HBV-related liver cirrhosis (LC) and 82 had HBV-related chronic hepatitis B (CHB). A direct sequencing method was employed to determine the HBV genotypes and the mutations in BCP/PC regions. In all, 133 of them were infected with genotype C viruses (79.17%); only 35 patients (20.83%) were infected with genotype B viruses. Both LC patients and CHB patients had significantly higher ratios of genotype C when compared with the ratios of genotype B (83.7%-16.3% versus 74.4%-25.6%). For patients with CHB, the prevalence of BCP/PC wild-type viruses was 52.4%; but this was only 4.7% in patients with LC. The C1653T, T1753C, A1762T/G1764A and G1896A mutations had a significantly higher prevalence in patients with LC. Among all the patients with genotype B viruses, those with LC had lower HBV DNA levels and higher G1899A mutation frequency than patients with CHB. Among all the patients with genotype C viruses, the patients with LC had higher prevalence of C1653T, A1762T/G1764A and G1896A mutation frequency, higher hepatitis B e antigen (HBeAg) -negative rates, lower viral load, lower elevated alanine aminotransferase and lower anti-HBe positive rates than CHB patients. The HBV BCP/PC variants were more common in HBeAg-negative LC patients than in the CHB group (BCP, 53.4% versus 15.6%; PC, 18.6% versus 3.7%, respectively, p < 0.001). Patients with HBV genotype C viruses, high viral load and C1653T, A1762T/G1764A, G1896A mutant viruses, were more susceptible to developing LC.
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Affiliation(s)
- Y W Zhong
- Institute of Infectious Diseases, Pediatric Liver Disease Therapy and Research Centre, Beijing 302 Hospital, Beijing, China.
| | - F L Di
- Institute of Infectious Diseases, Pediatric Liver Disease Therapy and Research Centre, Beijing 302 Hospital, Beijing, China
| | - C Liu
- Institute of Infectious Diseases, Pediatric Liver Disease Therapy and Research Centre, Beijing 302 Hospital, Beijing, China
| | - X C Zhang
- HeBei North University, Zhangjiakou, China.
| | - J F Bi
- Institute of Infectious Diseases, Pediatric Liver Disease Therapy and Research Centre, Beijing 302 Hospital, Beijing, China
| | - Y L Li
- Institute of Infectious Diseases, Pediatric Liver Disease Therapy and Research Centre, Beijing 302 Hospital, Beijing, China
| | - S Q Wu
- Institute of Infectious Diseases, Pediatric Liver Disease Therapy and Research Centre, Beijing 302 Hospital, Beijing, China
| | - H Dong
- Institute of Infectious Diseases, Pediatric Liver Disease Therapy and Research Centre, Beijing 302 Hospital, Beijing, China
| | - L M Liu
- Institute of Infectious Diseases, Pediatric Liver Disease Therapy and Research Centre, Beijing 302 Hospital, Beijing, China
| | - J He
- Institute of Infectious Diseases, Pediatric Liver Disease Therapy and Research Centre, Beijing 302 Hospital, Beijing, China
| | - Y M Shi
- Institute of Infectious Diseases, Pediatric Liver Disease Therapy and Research Centre, Beijing 302 Hospital, Beijing, China
| | - H F Zhang
- Institute of Infectious Diseases, Pediatric Liver Disease Therapy and Research Centre, Beijing 302 Hospital, Beijing, China.
| | - M Zhang
- Institute of Infectious Diseases, Pediatric Liver Disease Therapy and Research Centre, Beijing 302 Hospital, Beijing, China.
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Shirvani-Dastgerdi E, Pourkarim MR, Herbers U, Amini-Bavil-Olyaee S, Yagmur E, Alavian SM, Trautwein C, Tacke F. Hepatitis delta virus facilitates the selection of hepatitis B virus mutants in vivo and functionally impacts on their replicative capacity in vitro. Clin Microbiol Infect 2015; 22:98.e1-98.e6. [PMID: 26433026 DOI: 10.1016/j.cmi.2015.09.020] [Citation(s) in RCA: 6] [Impact Index Per Article: 0.6] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/04/2015] [Revised: 09/07/2015] [Accepted: 09/19/2015] [Indexed: 02/06/2023]
Abstract
To identify molecular interactions between hepatitis B virus (HBV) and hepatitis delta virus (HDV), HBV sequences were analysed in HBV/HDV-infected patients. Characteristic amino acid substitutions were found in cytosolic domains of hepatitis B surface antigen (HBsAg), in contrast to HBV-mono-infected controls. The functional impact of HDV on the replication of wild-type and mutant HBV was assessed in vitro. HDV co-transfection significantly reduced the replication of HBV strains containing precore or basal core promoter mutations, and HBV polymerase or surface antigen mutants affected HDV replication in vitro. Conclusively, our study revealed distinct HBsAg mutational patterns in HBV/HDV-infected patients and novel functional interactions between HBV and HDV.
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Affiliation(s)
| | - M R Pourkarim
- Department of Microbiology and Immunology, Laboratory of Clinical and Epidemiological Virology, Rega Institute for Medical Research, KU Leuven, Belgium; Blood Transfusion Research Centre, High Institute for Research and Education in Transfusion Medicine, Tehran, Iran
| | - U Herbers
- Department of Medicine III, RWTH-University Hospital Aachen, Aachen, Germany
| | - S Amini-Bavil-Olyaee
- Department of Molecular Microbiology and Immunology, Keck School of Medicine, University of Southern California, Harlyne J. Norris Cancer Research Tower, Los Angeles, CA, USA
| | - E Yagmur
- Medical Care Centre, Dr Stein and Colleagues, Mönchengladbach, Germany
| | - S M Alavian
- Baqiyatallah Research Centre for Gastroenterology and Liver Diseases, Baqiyatallah University of Medical Sciences, Tehran, Iran
| | - C Trautwein
- Department of Medicine III, RWTH-University Hospital Aachen, Aachen, Germany
| | - F Tacke
- Department of Medicine III, RWTH-University Hospital Aachen, Aachen, Germany.
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Leng XH, Chen EQ, Du LY, Bai L, Gong DY, Cheng X, Huang FJ, Tang H. Biological characteristics of the A1762T/G1764A mutant strain of hepatitis B virus in vivo. Mol Med Rep 2015; 12:5141-5148. [PMID: 26165271 DOI: 10.3892/mmr.2015.4072] [Citation(s) in RCA: 12] [Impact Index Per Article: 1.2] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/13/2014] [Accepted: 04/24/2015] [Indexed: 02/05/2023] Open
Abstract
The double nucleotide, A1762T and G1764A exchange (TA mutation), in the hepatitis B virus (HBV) genome basal core promoter (BCP) region is a common viral mutation in patients with chronic HBV infection. This mutation is located in the binding site of hepatocyte nuclear factor 4 (HNF4), and a number of liver‑enriched transcription factors are involved in the regulation of HBV transcription and replication. The aim of the present study was to investigate the biological characteristics of the HBV strain with this mutation, and the effect of HNF4 inhibition on the replication of this strain in vivo. The results indicated that in vivo the HBV strain with the TA mutation supported a higher level of pregenomic RNA transcription and HBV DNA replication, compared with the wild‑type strain. Furthermore, the concentration of serum HBeAg in the TA mutant group was lower than that in the wild‑type strain. Following treatment of the mice with entecavir (ETV) or tenofovir disoproxil fumarate (TDF), the transcription and replication levels of wild‑type and mutant strains were reduced. In the groups treated with TDF, the inhibition effect was more marked. In hepatocytes in which HNF4 expression was specifically inhibited, the level of 3.5 kb mRNA of HBV was reduced compared with that in mouse cells with normal HNF4 expression, and HBV DNA replication levels were also reduced to a greater extent. Furthermore, following liver‑specific knockdown of HNF4, the reduction in variant virus expression was greater than that of the wild‑type virus. In conclusion, the replication capacity of HBV with the TA mutation was increased, and the mutation was associated with a reduction in serum HBeAg levels. This mutant strain remained sensitive to ETV and TDF, and HNF4 supported a higher replication level of TA mutant HBV in vivo.
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Affiliation(s)
- Xiao-Hua Leng
- Center of Infectious Diseases, West China Hospital, Sichuan University, Chengdu, Sichuan 610041, P.R. China
| | - En-Qiang Chen
- Center of Infectious Diseases, West China Hospital, Sichuan University, Chengdu, Sichuan 610041, P.R. China
| | - Ling-Yao Du
- Center of Infectious Diseases, West China Hospital, Sichuan University, Chengdu, Sichuan 610041, P.R. China
| | - Lang Bai
- Center of Infectious Diseases, West China Hospital, Sichuan University, Chengdu, Sichuan 610041, P.R. China
| | - Dao-Ying Gong
- Department of Forensic Pathology, Medical School of Basic and Forensic Sciences, Sichuan University, Chengdu, Sichuan 610041, P.R. China
| | - Xing Cheng
- Center of Infectious Diseases, West China Hospital, Sichuan University, Chengdu, Sichuan 610041, P.R. China
| | - Fei-Jun Huang
- Department of Forensic Pathology, Medical School of Basic and Forensic Sciences, Sichuan University, Chengdu, Sichuan 610041, P.R. China
| | - Hong Tang
- Center of Infectious Diseases, West China Hospital, Sichuan University, Chengdu, Sichuan 610041, P.R. China
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30
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Hermans LE, Svicher V, Pas SD, Salpini R, Alvarez M, Ben Ari Z, Boland G, Bruzzone B, Coppola N, Seguin-Devaux C, Dyda T, Garcia F, Kaiser R, Köse S, Krarup H, Lazarevic I, Lunar MM, Maylin S, Micheli V, Mor O, Paraschiv S, Paraskevis D, Poljak M, Puchhammer-Stöckl E, Simon F, Stanojevic M, Stene-Johansen K, Tihic N, Trimoulet P, Verheyen J, Vince A, Weis N, Yalcinkaya T, Lepej SZ, Perno C, Boucher CAB, Wensing AMJ. Combined Analysis of the Prevalence of Drug-Resistant Hepatitis B Virus in Antiviral Therapy-Experienced Patients in Europe (CAPRE). J Infect Dis 2015; 213:39-48. [PMID: 26136470 DOI: 10.1093/infdis/jiv363] [Citation(s) in RCA: 24] [Impact Index Per Article: 2.4] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/04/2015] [Accepted: 06/23/2015] [Indexed: 01/05/2023] Open
Abstract
BACKGROUND European guidelines recommend treatment of chronic hepatitis B virus infection (CHB) with the nucleos(t)ide analogs (NAs) entecavir or tenofovir. However, many European CHB patients have been exposed to other NAs, which are associated with therapy failure and resistance. The CAPRE study was performed to gain insight in prevalence and characteristics of NA resistance in Europe. METHODS A survey was performed on genotypic resistance testing results acquired during routine monitoring of CHB patients with detectable serum hepatitis B virus DNA in European tertiary referral centers. RESULTS Data from 1568 patients were included. The majority (73.8%) were exposed to lamivudine monotherapy. Drug-resistant strains were detected in 52.7%. The most frequently encountered primary mutation was M204V/I (48.7%), followed by A181T/V (3.8%) and N236T (2.6%). In patients exposed to entecavir (n = 102), full resistance was present in 35.3%. Independent risk factors for resistance were age, viral load, and lamivudine exposure (P < .001). CONCLUSIONS These findings support resistance testing in cases of apparent NA therapy failure. This survey highlights the impact of exposure to lamivudine and adefovir on development of drug resistance and cross-resistance. Continued use of these NAs needs to be reconsidered at a pan-European level.
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Affiliation(s)
- Lucas Etienne Hermans
- Department of Medical Microbiology, University Medical Centre Utrecht Department of Virology, Erasmus Medical Centre, Rotterdam, The Netherlands
| | - Valentina Svicher
- Department of Experimental Medicine and Surgery, University of Rome Tor Vergata, Italy
| | | | - Romina Salpini
- Department of Experimental Medicine and Surgery, University of Rome Tor Vergata, Italy
| | - Marta Alvarez
- Servicio de Microbiología, Hospital San Cecilio, Instituto de Investigación Biosanitaria ibs. GRANADA, Hospitales Universitarios de Granada, Spain
| | - Ziv Ben Ari
- Liver Disease Centre, Sheba Medical Centre, Ramat Gan, Israel
| | - Greet Boland
- Department of Medical Microbiology, University Medical Centre Utrecht
| | | | - Nicola Coppola
- Malattie Infettive, Seconda Università degli studi di Napoli, Naples, Italy
| | | | - Tomasz Dyda
- Molecular Diagnostics Laboratory, Hospital of Infectious Diseases, Warsaw, Poland
| | - Federico Garcia
- Servicio de Microbiología, Hospital San Cecilio, Instituto de Investigación Biosanitaria ibs. GRANADA, Hospitales Universitarios de Granada, Spain
| | - Rolf Kaiser
- Institute of Virology, University of Cologne, Germany
| | - Sukran Köse
- Clinic of Infectious Diseases and Clinical Microbiology, Izmir Tepecik Education and Research Hospital, Turkey
| | - Henrik Krarup
- Section of Molecular Diagnostics, Clinical Biochemistry, Aalborg University Hospital, Denmark
| | - Ivana Lazarevic
- Institute of Microbiology and Immunology, Faculty of Medicine, University of Belgrade, Serbia
| | - Maja M Lunar
- Institute of Microbiology and Immunology, Faculty of Medicine, University of Ljubljana, Slovenia
| | - Sarah Maylin
- Service de Microbiologie, University Paris Diderot, Hôpital Saint Louis, France
| | | | - Orna Mor
- National HIV Reference Laboratory, Central Virology Laboratory, Ministry of Health, Tel Hashomer, Ramat Gan, Israel
| | - Simona Paraschiv
- Molecular Diagnostics Laboratory, National Institute for Infectious Diseases Matei Bals, Bucharest, Romania
| | - Dimitrios Paraskevis
- National Retrovirus Reference Centre, Department of Hygiene, Epidemiology and Medical Statistics, Faculty of Medicine, National and Kapodistrian University of Athens, Greece
| | - Mario Poljak
- Institute of Microbiology and Immunology, Faculty of Medicine, University of Ljubljana, Slovenia
| | | | - François Simon
- Service de Microbiologie, University Paris Diderot, Hôpital Saint Louis, France
| | - Maja Stanojevic
- Institute of Microbiology and Immunology, Faculty of Medicine, University of Belgrade, Serbia
| | | | - Nijaz Tihic
- Institute of Microbiology, Polyclinic for Laboratory Diagnostics, University Clinical Centre Tuzla, Bosnia and Herzegovina
| | - Pascale Trimoulet
- Virology Laboratory, Centre Hospitalier Régional et Université Victor Segalen, Bordeaux, France
| | - Jens Verheyen
- Institute of Virology, University-Hospital, University Duisburg-Essen, Germany
| | - Adriana Vince
- University of Zagreb School of Medicine and University Hospital for Infectious Diseases "Dr Fran Mihaljevic", Croatia
| | - Nina Weis
- Department of Infectious Diseases, Copenhagen University Hospital, Denmark
| | | | - Snjezana Zidovec Lepej
- University of Zagreb School of Medicine and University Hospital for Infectious Diseases "Dr Fran Mihaljevic", Croatia
| | - Carlo Perno
- Department of Experimental Medicine and Surgery, University of Rome Tor Vergata, Italy
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Shirvani-Dastgerdi E, Tacke F. Molecular interactions between hepatitis B virus and delta virus. World J Virol 2015; 4:36-41. [PMID: 25964870 PMCID: PMC4419120 DOI: 10.5501/wjv.v4.i2.36] [Citation(s) in RCA: 24] [Impact Index Per Article: 2.4] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 01/13/2015] [Revised: 02/12/2015] [Accepted: 03/09/2015] [Indexed: 02/05/2023] Open
Abstract
As a deficient virus due to the lack of envelope proteins, hepatitis D virus (HDV) causes chronic or fulminant “delta hepatitis” only in people with simultaneous hepatitis B virus (HBV) infection. HBV encodes three types of surface proteins known as small (S), medium (M) and large (L) envelope proteins. All three types of HBV surface antigens (HBsAgs) are present on HDV virions. The envelopment process of HDV occurs through interactions between the HDV ribonucleoprotein (RNP) complex and HBV HBsAgs. While HBsAg is the only protein required by HDV, the exact interaction sites between the S protein and pre-mature HDV are not well defined yet. In fact, these sites are distributed along the S protein with some hot spots for the envelopment process. Moreover, in most clinically studied samples, HDV infection is associated with a dramatically reduced HBV viral load, temporarily or permanently, while HBsAg resources are available for HDV packaging. Thus, beyond interacting with HBV envelope proteins, controlling mechanisms exist by which HDV inhibits HBV-DNA replication while allowing a selective transcription of HBV proteins. Here we discuss the molecular interaction sites between HBsAg and the HDV-RNP complex and address the proposed indirect mechanisms, which are employed by HBV and HDV to facilitate or inhibit each other’s viral replication. Understanding molecular interactions between HBV and HDV may help to design novel therapeutic strategies for delta hepatitis.
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Croagh CM, Desmond PV, Bell SJ. Genotypes and viral variants in chronic hepatitis B: A review of epidemiology and clinical relevance. World J Hepatol 2015; 7:289-303. [PMID: 25848459 PMCID: PMC4381158 DOI: 10.4254/wjh.v7.i3.289] [Citation(s) in RCA: 98] [Impact Index Per Article: 9.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 09/15/2014] [Revised: 12/04/2014] [Accepted: 12/29/2014] [Indexed: 02/06/2023] Open
Abstract
The Hepatitis B Virus (HBV) has a worldwide distribution and is endemic in many populations. It is constantly evolving and 10 genotypic strains have been identified with varying prevalences in different geographic regions. Numerous stable mutations in the core gene and in the surface gene of the HBV have also been identified in untreated HBV populations. The genotypes and viral variants have been associated with certain clinical features of HBV related liver disease and Hepatocellular carcinoma. For example Genotype C is associated with later hepatitis B e antigen (HBeAg) seroconversion, and more advanced liver disease. Genotype A is associated with a greater risk of progression to chronicity in adult acquired HBV infections. Genotype D is particularly associated with the precore mutation and HBeAg negative chronic hepatitis B (CHB). The genotypes prevalent in parts of West Africa, Central and South America, E, F and H respectively, are less well studied. Viral variants especially the Basal Core Promotor mutation is associated with increased risk of fibrosis and cancer of the liver. Although not currently part of routine clinical care, evaluation of genotype and viral variants may provide useful adjunctive information in predicting risk about liver related morbidity in patients with CHB.
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Affiliation(s)
- Catherine Mn Croagh
- Catherine MN Croagh, Paul V Desmond, Sally J Bell, Department of Gastroenterology, St Vincent's Hospital, Fitzroy, Victoria 3065, Australia
| | - Paul V Desmond
- Catherine MN Croagh, Paul V Desmond, Sally J Bell, Department of Gastroenterology, St Vincent's Hospital, Fitzroy, Victoria 3065, Australia
| | - Sally J Bell
- Catherine MN Croagh, Paul V Desmond, Sally J Bell, Department of Gastroenterology, St Vincent's Hospital, Fitzroy, Victoria 3065, Australia
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Park YM. Clinical utility of complex mutations in the core promoter and proximal precore regions of the hepatitis B virus genome. World J Hepatol 2015; 7:113-120. [PMID: 25625002 PMCID: PMC4295188 DOI: 10.4254/wjh.v7.i1.113] [Citation(s) in RCA: 3] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 08/28/2014] [Revised: 10/12/2014] [Accepted: 10/29/2014] [Indexed: 02/06/2023] Open
Abstract
The core promoter and proximal precore regions are the most complex portions of the hepatitis B virus (HBV) genome. These regions cooperatively regulate viral replication and differentially regulate the synthesis of the viral proteins E, core, and X. Multiple mutations in these regions are associated with the persistency of viral infection and the development of cirrhosis and hepatocellular carcinoma (HCC). In South Korea, nearly all HBVs are classified as HBV genotype C2; the majority of these viruses have the basal core promoter double mutation, a precore stop mutation, or both. These mutations may play a role in the alteration of viral and clinical features, and abundant and complex mutations are particularly prevalent in the core promoter and proximal precore regions. We previously demonstrated that the accumulation of ≥ 6 mutations at eight key nucleotides located in these regions (G1613A, C1653T, T1753V, A1762T, G1764A, A1846T, G1896A, and G1899A) is a useful marker to predict the development of HCC regardless of advanced liver disease. In addition, certain mutation combinations were predominant in cases with ≥ 4 mutations. In cases with ≤ 5 mutations, a low Hepatitis B e antigen titer (< 35 signal to noise ratio) was indicative of HCC risk. Viral mutation data of the single HBV genotype C2 suggest that the combined effect of the number and pattern of mutations in the core promoter and proximal precore regions is helpful in predicting HCC risk.
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Cui XJ, Cho YK, Song BC. Influence of the basal core promoter and precore mutation on replication of hepatitis B virus and antiviral susceptibility of different genotypes. J Med Virol 2015; 87:601-8. [PMID: 25612255 DOI: 10.1002/jmv.24117] [Citation(s) in RCA: 6] [Impact Index Per Article: 0.6] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Accepted: 10/21/2014] [Indexed: 12/22/2022]
Abstract
Mutations in the basal core promoter (BCP) and precore (PC) regions of the hepatitis B virus (HBV) are more common in genotypes B and C than in genotype A, suggesting that these mutations might affect replication competency depending on genotype. The purpose of the study was to investigate the influence of these mutations on the capacity of HBV for replication and antiviral drug susceptibility according to genotype. Genotypes A, B, and C of HBV strains with a BCP mutation, PC mutation, or BCP + PC mutation were made by site-directed mutagenesis. Replication competency of each construct and susceptibility to nucleos(t) ide analogues were tested in an Huh7 cell line. In genotype A, the BCP and BCP + PC mutations increased the viral replication around 6.5 times compared with the wild type, and the PC mutation alone similarly increased the viral replication around three times. In genotypes B and C, all three mutant types increased viral replication to a similar extent, regardless of mutation pattern. Interestingly, the BCP mutation appeared to have a greater effect on viral replication in genotype A than in genotypes B and C. This finding was unexpected because the BCP mutation is more common in HBV genotypes B and C. Moreover, the BCP, PC, and BCP + PC mutations decreased the sensitivity of HBV to antiviral agents to various degrees (2- to 10-fold) regardless of genotype. In conclusion, BCP and PC mutations increased viral replication regardless of HBV genotype and decreased in vitro antiviral susceptibility to the nucleos(t) ide analogues.
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Affiliation(s)
- Xiu-Ji Cui
- Department of Internal Medicine, School of Medicine, Jeju National University, Korea
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Yildiz UH, Inci F, Wang S, Toy M, Tekin HC, Javaid A, Lau DTY, Demirci U. Recent advances in micro/nanotechnologies for global control of hepatitis B infection. Biotechnol Adv 2015; 33:178-190. [PMID: 25450190 PMCID: PMC4433022 DOI: 10.1016/j.biotechadv.2014.11.003] [Citation(s) in RCA: 29] [Impact Index Per Article: 2.9] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/21/2014] [Revised: 11/03/2014] [Accepted: 11/04/2014] [Indexed: 12/18/2022]
Abstract
The control of hepatitis B virus (HBV) infection is a challenging task, specifically in developing countries there is limited access to diagnostics and antiviral treatment mainly due to high costs and insufficient healthcare infrastructure. Although the current diagnostic technologies can reliably detect HBV, they are relatively laborious, impractical and require expensive resources that are not suitable for resource-limited settings. Advances in micro/nanotechnology are pioneering the development of new generation methodologies in diagnosis and screening of HBV. Owing to combination of nanomaterials (metal/inorganic nanoparticles, carbon nanotubes, etc.) with microfabrication technologies, utilization of miniaturized sensors detecting HBV and other viruses from ultra-low volume of blood, serum and plasma is realized. The state-of-the-art microfluidic devices with integrated nanotechnologies potentially allow for inexpensive HBV screening at low cost. This review aims to highlight recent advances in nanotechnology and microfabrication processes that are employed for developing point-of-care (POC) HBV assays.
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Affiliation(s)
- U Hakan Yildiz
- Demirci Bio-Acoustic-MEMS in Medicine (BAMM) Laboratory, Stanford University School of Medicine, Canary Center at Stanford for Cancer Early Detection, Palo Alto, CA 94304, United States
| | - Fatih Inci
- Demirci Bio-Acoustic-MEMS in Medicine (BAMM) Laboratory, Stanford University School of Medicine, Canary Center at Stanford for Cancer Early Detection, Palo Alto, CA 94304, United States
| | - ShuQi Wang
- Demirci Bio-Acoustic-MEMS in Medicine (BAMM) Laboratory, Stanford University School of Medicine, Canary Center at Stanford for Cancer Early Detection, Palo Alto, CA 94304, United States
| | - Mehlika Toy
- Department of Surgery, Stanford School of Medicine, Palo Alto, CA, United States
| | - H Cumhur Tekin
- Demirci Bio-Acoustic-MEMS in Medicine (BAMM) Laboratory, Stanford University School of Medicine, Canary Center at Stanford for Cancer Early Detection, Palo Alto, CA 94304, United States
| | - Asad Javaid
- Department of Medicine, Beth Israel Deaconnes Medical Center, Harvard Medical School, Boston, MA, United States
| | - Daryl T-Y Lau
- Department of Medicine, Beth Israel Deaconnes Medical Center, Harvard Medical School, Boston, MA, United States
| | - Utkan Demirci
- Demirci Bio-Acoustic-MEMS in Medicine (BAMM) Laboratory, Stanford University School of Medicine, Canary Center at Stanford for Cancer Early Detection, Palo Alto, CA 94304, United States.
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Retrospective study of the prevalence and clinical significance of hepatitis B virus precore and basal core promoter variants. Can J Gastroenterol Hepatol 2015; 29:e1-6. [PMID: 26401823 PMCID: PMC4699608 DOI: 10.1155/2015/940825] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.1] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 11/17/2022] Open
Abstract
BACKGROUND Hepatitis B virus (HBV) precore (PC) and basal core promoter (BCP) variants are well known; however, their prevalence in North America is unclear, especially among hepatitis B e antigen-negative patients. OBJECTIVE To investigate the prevalence of PC⁄BCP mutations and their clinical significance. METHODS One hundred twenty-eight patients positive for both hepatitis B surface antigen and hepatitis B e antibody were selected, and PC⁄BCP mutations were identified using a line probe assay. The subjects' charts were reviewed for race⁄ethnicity, HBV genotype, HBV viral load, sex, liver enzyme levels, imaging and biopsy results up to 10 years before the study. RESULTS The prevalence of PC and BCP variants were 47.6% and 62.5%, respectively. Older age was associated with aspartate aminotransferase-to-platelet index ratio (APRI) ≥0.7 (P=0.011) and abnormal imaging⁄biopsy results (P=0.0008). Although the presence of BCP variant(s) was associated with APRI ≥0.7 (P=0.029), it was not associated with abnormal imaging⁄biopsy results. The combination of age ≥50 years and the presence of BCP variant(s) was associated with abnormal imaging⁄biopsy results, suggestive of either cirrhosis or hepatocellular carcinoma (not observed with PC mutation). Neither sex or genotype, or median HBV viral load showed significant influence on any of these outcomes. CONCLUSIONS The present study suggests that the prevalence of PC and BCP mutations are higher than what has been previously reported. One potential explanation would be increased immigration in the past decade. Considering the potential public health and clinical implications of these variants, long-term multicentre and prospective studies could further unravel the uncertainty around these variants.
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Constantinescu I, Dinu AA, Boscaiu V, Niculescu M. Hepatitis B virus core promoter mutations in patients with chronic hepatitis B and hepatocellular carcinoma in bucharest, romania. HEPATITIS MONTHLY 2014; 14:e22072. [PMID: 25477976 PMCID: PMC4250966 DOI: 10.5812/hepatmon.22072] [Citation(s) in RCA: 9] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Subscribe] [Scholar Register] [Received: 07/25/2014] [Revised: 09/10/2014] [Accepted: 09/26/2014] [Indexed: 12/11/2022]
Abstract
BACKGROUND Accurate and personalized molecular virological diagnosis of hepatitis B virus (HBV) infection is crucial for individualized selection of patients for antiviral therapy in Romania. OBJECTIVES We aimed to investigate HBV mutations in Romanian patients with chronic HBV infection, also to match HBV genotypes with HBV mutations identified and clinical outcomes. PATIENTS AND METHODS This was a cross-sectional study. A total of 484 Romanian patients with chronic HBV infection and hepatocellular carcinoma (HCC) were investigated. This was performed in Fundeni Clinical Institute, Bucharest, Romania during January 2005 to August 2010. HBsAg positive patients with chronic HBV infection admitted to Fundeni Clinical Institute were randomly enrolled in the study. Analysis was performed in the Centre for Immunogenetics and Virology, Fundeni Clinical Institute, Bucharest, Romania. Indirect diagnosis was performed with enhanced chemiluminescence method using Architect i2000SR and HBV-DNA was quantified with COBAS TaqMan HBV PCR. Direct sequencing of the PCR-products was performed with the PCR-product sequencing kit. HBV genotyping was performed with INNO-LiPA DR Amplification and INNO-LiPA HBV precore-core. RESULTS We detected two HBV genotypes; A (8.1%) and D (60.5%), and a mixture of genotypes A and D (31.4%) (P < 0.001). Basal core promoter (BCP) A1762T/G1764A and precore (PC) G1896A mutations were detected in these Romanian patients with chronic HBV infection. HBV chronic carriers had mainly genotype D (54.4%) and HBV WT (64.0%). BCP A1762T, G1764A and PC G1896A were significantly associated with HCC-tissue HBV sequencing (75.3%) (P < 0.001). PC G1896A alone was detected in HCC-serum HBV sequencing group (66.7%). CONCLUSIONS Genotype D was the main genotype detected in Romanian patients with chronic HBV infection. Genotype D presented both BCP and PC mutations more frequently.
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Affiliation(s)
- Ileana Constantinescu
- Immunology of Transplantation Discipline, Faculty of Medicine, Carol Davila University of Medicine and Pharmacy, Bucharest, Romania
- Center for Immunogenetics and Virology, Fundeni Clinical Institute, Bucharest, Romania
- Corresponding Author: Ileana Constantinescu, Centre for Immunogenetics and Virology, Fundeni Clinical Institute, Bucharest, Romania. Tel: +40-744341984, Fax: +40-213180448, E-mail:
| | - Andrei-Antoniu Dinu
- Center for Immunogenetics and Virology, Fundeni Clinical Institute, Bucharest, Romania
| | - Voicu Boscaiu
- “Gheorghe Mihoc-Caius Iacob” Institute of Statistics and Applied Mathematics, Bucharest, Romania
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Menéndez-Arias L, Álvarez M, Pacheco B. Nucleoside/nucleotide analog inhibitors of hepatitis B virus polymerase: mechanism of action and resistance. Curr Opin Virol 2014; 8:1-9. [PMID: 24814823 DOI: 10.1016/j.coviro.2014.04.005] [Citation(s) in RCA: 112] [Impact Index Per Article: 10.2] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/02/2014] [Revised: 04/14/2014] [Accepted: 04/16/2014] [Indexed: 02/07/2023]
Abstract
Hepatitis B virus (HBV) polymerase and human immunodeficiency virus (HIV) reverse transcriptase are structurally related. However, the HBV enzyme has a protein priming activity absent in the HIV enzyme. Approved nucleoside/nucleotide inhibitors of the HBV polymerase include lamivudine, adefovir, telbivudine, entecavir and tenofovir. Although most of them target DNA elongation, guanosine and adenosine analogs (e.g. entecavir and tenofovir, respectively) also impair protein priming. Major mutational patterns conferring nucleoside/nucleotide analog resistance include the combinations rtL180M/rtM204(I/V) (for lamivudine, entecavir, telbivudine and clevudine) and rtA181V/rtN236T (for adefovir and tenofovir). However, development of drug resistance is very slow for entecavir and tenofovir. Novel nucleoside/nucleotide analogs in advanced clinical trials include phosphonates similar to adefovir or tenofovir, and new tenofovir derivatives with improved pharmacological properties.
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Affiliation(s)
- Luis Menéndez-Arias
- Centro de Biología Molecular "Severo Ochoa" (Consejo Superior de Investigaciones Científicas & Universidad Autónoma de Madrid), c/Nicolás Cabrera, 1, Campus de Cantoblanco, Madrid 28049, Spain.
| | - Mar Álvarez
- Centro de Biología Molecular "Severo Ochoa" (Consejo Superior de Investigaciones Científicas & Universidad Autónoma de Madrid), c/Nicolás Cabrera, 1, Campus de Cantoblanco, Madrid 28049, Spain
| | - Beatriz Pacheco
- Centro de Biología Molecular "Severo Ochoa" (Consejo Superior de Investigaciones Científicas & Universidad Autónoma de Madrid), c/Nicolás Cabrera, 1, Campus de Cantoblanco, Madrid 28049, Spain
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Azmi AN, Tan SS, Mohamed R. Practical approach in hepatitis B e antigen-negative individuals to identify treatment candidates. World J Gastroenterol 2014; 20:12045-12055. [PMID: 25232242 PMCID: PMC4161793 DOI: 10.3748/wjg.v20.i34.12045] [Citation(s) in RCA: 3] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 10/27/2013] [Revised: 01/17/2014] [Accepted: 04/09/2014] [Indexed: 02/06/2023] Open
Abstract
The natural history of chronic hepatitis B is characterized by different phases of infection, and patients may evolve from one phase to another or may revert to a previous phase. The hepatitis B e antigen (HBeAg)-negative form is the predominant infection worldwide, which consists of individuals with a range of viral replication and liver disease severity. Although alanine transaminase (ALT) remains the most accessible test available to clinicians for monitoring the liver disease status, further evaluations are required for some patients to assess if treatment is warranted. Guidance from practice guidelines together with thorough investigations and classifications of patients ensure recognition of who needs which level of care. This article aims to assist physicians in the assessment of HBeAg-negative individuals using liver biopsy or non-invasive tools such as hepatitis B s antigen quantification and transient elastography in addition to ALT and hepatitis B virus DNA, to identify who will remain stable, who will reactivate or at risk of disease progression hence will benefit from timely initiation of anti-viral therapy.
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Fan J, Wang Y, Xiong H, Guo X, Cheng YC. Impact of the rtI187V polymerase substitution of hepatitis B virus on viral replication and antiviral drug susceptibility. J Gen Virol 2014; 95:2523-2530. [PMID: 25028473 DOI: 10.1099/vir.0.066886-0] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.2] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/25/2022] Open
Abstract
A high prevalence of the rtI187V polymerase substitution of hepatitis B virus (HBV) was detected in nucleoside/nucleotide-analogue-naive and -treated chronic hepatitis B (CHB) patients. We aimed at assessing the replicative capacity and susceptibility to lamivudine (LAM) and adefovir (ADV) in vitro of HBV harbouring rtI187V alone or in conjunction with LAM- or ADV-resistant mutations. The reverse transcriptase region of HBV isolates was directly sequenced from a cohort of 300 CHB patients from China. Replication-competent HBV constructs containing rtI187V and combined with LAM-resistant (rtM204I, rtL180M/rtM204V) mutations were generated, and compared with WT, LAM-resistant single (rtM204I) or double (rtL180M/rtM204V) and ADV-resistant (rtN236T) clones. In a Chinese cohort of 300 CHB patients, 8.7 % (26/300) showed substitution of rtI187 with V. Of note, the rtI187V prevalence in HBV genotype B was significantly higher than that in HBV genotype C (95.2 vs 4.8 %). In vitro phenotypic assays showed that the viruses bearing the rtI187V substitution had impaired replication efficacy when compared with the WT and the virus carrying rtI187V combined with LAM-resistant single or double mutations showed even more significantly impaired replicative capacities. Furthermore, rtI187V HBV remained susceptible towards treatment with LAM or ADV in vitro whereas the combination of the rtI187V substitution with LAM-resistant mutations rendered HBV resistant to LAM but still sensitive to ADV. Our study revealed that the rtI187V substitution in the HBV polymerase frequently occurred in CHB patients, particularly those with HBV genotype B. However, the emergence of the rtI187V substitution significantly impaired viral replication but without affecting drug sensitivity in vitro.
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Affiliation(s)
- Jiyun Fan
- Department of Microbiology and Immunology, Institutes of Medical Sciences, Shanghai Jiao Tong University School of Medicine, Shanghai, PR China
| | - Ying Wang
- Department of Microbiology and Immunology, Institutes of Medical Sciences, Shanghai Jiao Tong University School of Medicine, Shanghai, PR China
| | - Hui Xiong
- Chinese National Human Genome Center, Shanghai, PR China
| | - Xiaokui Guo
- Department of Microbiology and Immunology, Institutes of Medical Sciences, Shanghai Jiao Tong University School of Medicine, Shanghai, PR China
| | - Yung-Chi Cheng
- Department of Pharmacology, Yale University School of Medicine, New Haven, CT, USA
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Wang YZ, Zhu Z, Zhang HY, Zhu MZ, Xu X, Chen CH, Liu LG. Detection of hepatitis B virus A1762T/G1764A mutant by amplification refractory mutation system. Braz J Infect Dis 2014; 18:261-5. [PMID: 24389280 PMCID: PMC9427444 DOI: 10.1016/j.bjid.2013.09.005] [Citation(s) in RCA: 9] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/10/2013] [Revised: 08/03/2013] [Accepted: 09/11/2013] [Indexed: 01/25/2023] Open
Affiliation(s)
- Yong-Zhong Wang
- Institute for the Study of Liver Diseases, The Third People's Hospital of Changzhou, Changzhou, China
| | - Zhen Zhu
- Institute for the Study of Liver Diseases, The Third People's Hospital of Changzhou, Changzhou, China
| | - Hong-Yu Zhang
- Institute for the Study of Liver Diseases, The Third People's Hospital of Changzhou, Changzhou, China
| | - Min-Zhi Zhu
- Institute for the Study of Liver Diseases, The Third People's Hospital of Changzhou, Changzhou, China
| | - Xin Xu
- Institute for the Study of Liver Diseases, The Third People's Hospital of Changzhou, Changzhou, China
| | - Chun-Hua Chen
- Institute for the Study of Liver Diseases, The Third People's Hospital of Changzhou, Changzhou, China
| | - Long-Gen Liu
- Institute for the Study of Liver Diseases, The Third People's Hospital of Changzhou, Changzhou, China.
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Moradi A, Zhand S, Ghaemi A, Javid N, Bazouri M, Tabarraei A. Mutations in pre-core and basal-core promoter regions of hepatitis B virus in chronic HBV patients from Golestan, Iran. IRANIAN JOURNAL OF BASIC MEDICAL SCIENCES 2014; 17:370-7. [PMID: 24967066 PMCID: PMC4069846] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Download PDF] [Figures] [Subscribe] [Scholar Register] [Received: 04/27/2013] [Accepted: 10/05/2013] [Indexed: 11/19/2022]
Abstract
OBJECTIVES It has been reported that the mutation of the pre-core (PC) and basal-core promoter (BCP) may play an important role in the development of HBV-related hepatocellular carcinoma (HCC). In this study the PC and BCP mutations were investigated in chronic HBV patients. MATERIALS AND METHODS In this study, 120 chronic HBV patients from Golestan, Northeast of Iran who were not vaccinated against HBV, were recruited from the year 2008 to 2012. HBV-DNA extraction from plasma and PCR were performed and positive PCR products were subjected to automated sequencing. RESULTS One hundred out of 120 (83.3%) patients were HBeAg negative. Comparison of our nucleotide sequences with reference sequence showed high rate mutation in BCP and PC region (96.66%). Frame shift mutation was found in 78 (65%) of patients in BCP region, among them 8 (6.6%) patients showed mutation in PC region. CONCLUSION Our results demonstrated high rate of mutations in BCP and PC regions among HBV chronic patients in Northeast of Iran.
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Affiliation(s)
- Abdolvahab Moradi
- Infectious Diseases Research Center, Department of Microbiology, School of Medicine, Golestan University of Medical Sciences, Gorgan, Iran
| | - Sareh Zhand
- Infectious Diseases Research Center, Department of Microbiology, School of Medicine, Golestan University of Medical Sciences, Gorgan, Iran
| | - Amir Ghaemi
- Infectious Diseases Research Center, Department of Microbiology, School of Medicine, Golestan University of Medical Sciences, Gorgan, Iran
| | - Naeme Javid
- Infectious Diseases Research Center, Department of Microbiology, School of Medicine, Golestan University of Medical Sciences, Gorgan, Iran
| | - Masoud Bazouri
- Infectious Diseases Research Center, Department of Microbiology, School of Medicine, Golestan University of Medical Sciences, Gorgan, Iran
| | - Alijan Tabarraei
- Infectious Diseases Research Center, Department of Microbiology, School of Medicine, Golestan University of Medical Sciences, Gorgan, Iran,Corresponding author: AlijanTabarraei. Infectious Diseases Research Centre, Department of Microbiology, School of Medicine, Golestan University of Medical Sciences, Gorgan, Iran. Tel: +98-9112733321; +98-171-4421289; Fax: +98-171-4440225; ;
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Tuteja A, Siddiqui AB, Madan K, Goyal R, Shalimar, Sreenivas V, Kaur N, Panda SK, Narayanasamy K, Subodh S, Acharya SK. Mutation profiling of the hepatitis B virus strains circulating in North Indian population. PLoS One 2014; 9:e91150. [PMID: 24637457 PMCID: PMC3956465 DOI: 10.1371/journal.pone.0091150] [Citation(s) in RCA: 17] [Impact Index Per Article: 1.5] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/10/2014] [Accepted: 02/07/2014] [Indexed: 12/11/2022] Open
Abstract
AIMS The aim of this study was to investigate the genomic mutations in the circulating Hepatitis B virus strains causing infection in the Indian population. Further, we wanted to analyze the biological significance of these mutations in HBV mediated disease. METHODS 222 HBsAg positive patients were enrolled in the study. The genotype and mutation profile was determined for the infecting HBV isolate by sequencing overlapping fragments. These sequences were analyzed by using different tools and compared with previously available HBV sequence information. Mutation Frequency Index (MFI) for the Genes and Diagnosis group was also calculated. RESULTS HBV Genotype D was found in 55% (n = 121) of the patient group and genotype A was found in 30% (n = 66) of samples. The majority (52%) of the HBV-infected individuals in the present study were HBeAg-negative in all the age groups studied. Spontaneous drug associated mutations implicated in resistance to antiviral therapy were also identified in about quarter of our patients, which is of therapeutic concern. The MFI approach used in the study indicated that Core peptide was the most conserved region in both genotypes and Surface peptide had highest mutation frequency. Few mutations in X gene (T36A and G50R) showed high frequency of association with HCC. A rare recombinant strain of HBV genotype A and D was also identified in the patient group. CONCLUSIONS HBV genotype D was found out to be most prevalent. More than half of the patients studied had HBeAg negative disease. Core region was found to be most conserved. Drug Associated mutations were detected in 22% of the patient group and T36A and G50R mutations in X gene were found to be associated with HCC.
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Affiliation(s)
- Amit Tuteja
- Institute of Molecular Medicine, New Delhi, India
- Amity Institute of Biotechnology, Amity University, Noida, India
| | | | - Kaushal Madan
- Department of Gastroenterology, All India Institute of Medical Sciences, New Delhi, India
| | - Rohit Goyal
- Department of Gastroenterology, All India Institute of Medical Sciences, New Delhi, India
| | - Shalimar
- Department of Gastroenterology, All India Institute of Medical Sciences, New Delhi, India
| | | | - Navkiran Kaur
- Amity Institute of Biotechnology, Amity University, Noida, India
| | - Subrat K. Panda
- Department of Pathology, All India Institute of Medical Sciences, New Delhi, India
| | | | - Swati Subodh
- Institute of Molecular Medicine, New Delhi, India
- Open Source Drug Discovery Unit, Council of Scientific & Industrial Research, New Delhi, India
| | - Subrat K. Acharya
- Department of Gastroenterology, All India Institute of Medical Sciences, New Delhi, India
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Khan A, Al Balwi MA, Tanaka Y, Hajeer A, Sanai FM, Al Abdulkarim I, Al Ayyar L, Badri M, Saudi D, Tamimi W, Mizokami M, Al Knawy B. Novel point mutations and mutational complexes in the enhancer II, core promoter and precore regions of hepatitis B virus genotype D1 associated with hepatocellular carcinoma in Saudi Arabia. Int J Cancer 2013; 133:2864-2871. [PMID: 23740667 DOI: 10.1002/ijc.28307] [Citation(s) in RCA: 19] [Impact Index Per Article: 1.6] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/22/2013] [Accepted: 05/14/2013] [Indexed: 10/10/2023]
Abstract
In this study, a cohort of 182 patients [55 hepatocellular carcinoma (HCC) and 127 non-HCC] infected with hepatitis B virus (HBV) in Saudi Arabia was investigated to study the relationship between sequence variation in the enhancer II (EnhII), basal core promoter (BCP) and precore regions of HBV genotype D (HBV/D) and the risk of HCC. HBV genotypes were determined by sequencing analysis and/or enzyme-linked immunosorbent assay. Variations in the EnhII, BCP and precore regions were compared between 107 non-HCC and 45 HCC patients infected with HBV/D, followed by age-matched analysis of 40 cases versus equal number of controls. Age and male gender were significantly associated with HCC (p = 0.0001 and p = 0.03, respectively). Serological markers such as aspartate aminotransferase, albumin and anti-HBe were significantly associated with HCC (p = 0.0001 for all), whereas HBeAg positivity was associated with non-HCC (p = 0.0001). The most prevalent HBV genotype was HBV/D (94%), followed by HBV/E (4%), HBV/A (1.6%) and HBV/C (0.5%). For HBV/D1, genomic mutations associated with HCC were T1673/G1679, G1727, C1741, C1761, A1757/T1764/G1766, T1773, T1773/G1775 and C1909. Age- and gender-adjusted stepwise logistic regression analysis indicated that mutations G1727 [odds ratio (OR) = 18.3; 95% confidence interval (CI) = 2.8-118.4; p = 0.002], A1757/T1764/G1766 (OR = 4.7; 95% CI = 1.3-17.2; p = 0.01) and T1773 (OR = 14.06; 95% CI = 2.3-84.8; p = 0.004) are independent predictors of HCC development. These results implicate novel individual and combination patterns of mutations in the X/precore region of HBV/D1 as predictors of HCC. Risk stratification based on these mutation complexes would be useful in determining high-risk patients and improving diagnostic and treatment strategies for HBV/D1.
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Affiliation(s)
- Anis Khan
- King Abdullah International Medical Research Center, King Saud bin Abdulaziz University for Health Sciences, Riyadh, Saudi Arabia; Department of Virology and Liver Unit, Graduate School of Medical Sciences, Nagoya City University, Nagoya, Japan
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Rodriguez-Frias F, Buti M, Tabernero D, Homs M. Quasispecies structure, cornerstone of hepatitis B virus infection: mass sequencing approach. World J Gastroenterol 2013; 19:6995-7023. [PMID: 24222943 PMCID: PMC3819535 DOI: 10.3748/wjg.v19.i41.6995] [Citation(s) in RCA: 30] [Impact Index Per Article: 2.5] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 06/28/2013] [Revised: 08/23/2013] [Accepted: 09/15/2013] [Indexed: 02/06/2023] Open
Abstract
Hepatitis B virus (HBV) is a DNA virus with complex replication, and high replication and mutation rates, leading to a heterogeneous viral population. The population is comprised of genomes that are closely related, but not identical; hence, HBV is considered a viral quasispecies. Quasispecies variability may be somewhat limited by the high degree of overlapping between the HBV coding regions, which is especially important in the P and S gene overlapping regions, but is less significant in the X and preCore/Core genes. Despite this restriction, several clinically and pathologically relevant variants have been characterized along the viral genome. Next-generation sequencing (NGS) approaches enable high-throughput analysis of thousands of clonally amplified regions and are powerful tools for characterizing genetic diversity in viral strains. In the present review, we update the information regarding HBV variability and present a summary of the various NGS approaches available for research in this virus. In addition, we provide an analysis of the clinical implications of HBV variants and their study by NGS.
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Mese S, Arikan M, Cakiris A, Abaci N, Gumus E, Kursun O, Onel D, Ustek D, Kaymakoglu S, Badur S, Yenen OS, Bozkaya E. Role of the line probe assay INNO-LiPA HBV DR and ultradeep pyrosequencing in detecting resistance mutations to nucleoside/nucleotide analogues in viral samples isolated from chronic hepatitis B patients. J Gen Virol 2013; 94:2729-2738. [PMID: 24045109 DOI: 10.1099/vir.0.053041-0] [Citation(s) in RCA: 9] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 02/06/2023] Open
Abstract
Despite the effectiveness of nucleoside/nucleotide analogues in the treatment of chronic hepatitis B (CHB), their long-term administration is associated with the emergence of resistant hepatitis B virus (HBV) mutants. In this study, mutations resulting in antiviral resistance in HBV DNA samples isolated from 23 CHB patients (nine treatment naïve and 14 treated previously) were studied using a line probe assay (INNO-LiPA HBV DR; Innogenetics) and ultradeep pyrosequencing (UDPS) methods. Whilst the INNO-LiPA HBV DR showed no resistance mutations in HBV DNA samples from treatment-naive patients, mutations mediating lamivudine resistance were detected in three samples by UDPS. Among patients who were treated previously, 19 mutations were detected in eight samples using the INNO-LiPA HBV DR and 29 mutations were detected in 12 samples using UDPS. All mutations detected by the INNO-LiPA HBV DR were also detected by UDPS. There were no mutations that could be detected by INNO-LiPA HBV DR but not by UDPS. A total of ten mutations were detected by UDPS but not by INNO-LiPA HBV DR, and the mean frequency of these mutations was 14.7 %. It was concluded that, although INNO-LiPA HBV DR is a sensitive and practical method commonly used for the detection of resistance mutations in HBV infection, UDPS may significantly increase the detection rate of genotypic resistance in HBV at an early stage.
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Affiliation(s)
- Sevim Mese
- Department of Virology and Fundamental Immunology, Istanbul Medical Faculty, Istanbul University, Istanbul, Turkey
| | - Muzaffer Arikan
- Department of Genetics, Institute for Experimental Medicine, Istanbul University, Istanbul, Turkey
| | - Aris Cakiris
- Department of Genetics, Institute for Experimental Medicine, Istanbul University, Istanbul, Turkey
| | - Neslihan Abaci
- Department of Genetics, Institute for Experimental Medicine, Istanbul University, Istanbul, Turkey
| | - Ergun Gumus
- Department of Computer Engineering, Istanbul University, Istanbul, Turkey
| | - Olcay Kursun
- Department of Computer Engineering, Istanbul University, Istanbul, Turkey
| | - Derya Onel
- Department of Virology and Fundamental Immunology, Istanbul Medical Faculty, Istanbul University, Istanbul, Turkey
| | - Duran Ustek
- Department of Genetics, Institute for Experimental Medicine, Istanbul University, Istanbul, Turkey
| | - Sabahattin Kaymakoglu
- Department of Gastroenterology, Istanbul Medical Faculty, Istanbul University, Istanbul, Turkey
| | - Selim Badur
- Department of Virology and Fundamental Immunology, Istanbul Medical Faculty, Istanbul University, Istanbul, Turkey
| | - Osman Sadi Yenen
- Department of Virology and Fundamental Immunology, Istanbul Medical Faculty, Istanbul University, Istanbul, Turkey
| | - Emel Bozkaya
- Department of Virology and Fundamental Immunology, Istanbul Medical Faculty, Istanbul University, Istanbul, Turkey
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Herbers U, Amini-Bavil-Olyaee S, Mueller A, Luedde T, Trautwein C, Tacke F. Hepatitis B e antigen-suppressing mutations enhance the replication efficiency of adefovir-resistant hepatitis B virus strains. J Viral Hepat 2013; 20:141-8. [PMID: 23301549 DOI: 10.1111/j.1365-2893.2012.01639.x] [Citation(s) in RCA: 6] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 12/12/2022]
Abstract
Hepatitis B e antigen (HBeAg) negative hepatitis B virus (HBV) infections caused by precore (PC) or basal core promoter (BCP) mutations are associated with disease progression and complications. PC or BCP mutations may enhance the replication capacity of distinct drug-resistance-associated polymerase mutations, but their effect on adefovir-resistant HBV mutants is unclear. Importantly, BCP mutations were an independent risk factor for virological breakthrough in lamivudine-resistant patients treated with adefovir. We aimed at addressing the functional consequences of PC and BCP mutations on the replication and drug susceptibility of adefovir-resistant HBV mutants. Therefore, HBV constructs with wild type (WT) or adefovir-resistant rtN236T, rtA181V and rtA181T mutations, with or without concomitant PC or BCP mutations, were analysed in vitro using molecular assays. The adefovir-resistant polymerase mutations rtN236T, rtA181V and rtA181T showed a drastically reduced viral replication compared with WT. Interestingly, additional PC or BCP mutations enhanced the reduced replication efficacy of adefovir-resistant constructs and restored HBV replication to WT level. HBV rtA181T mutants displayed abolished hepatitis B surface antigen (HBsAg) secretion, owing to a sW172* stop codon in the overlapping envelope gene. All rtN236T- or rtA181V/T-containing constructs, regardless of concomitant PC or BCP mutations, were resistant to adefovir, but remained susceptible to telbivudine, entecavir and tenofovir. In conclusion, adefovir drug resistance mutations reduced viral replication, which can be significantly increased by additional HBeAg-suppressing PC or BCP mutations. Because increased HBV replication in HBeAg-negative patients has been associated with an unfavourable clinical course, close monitoring appears indispensable during adefovir treatment in HBeAg-negative patients.
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Affiliation(s)
- U Herbers
- Department of Medicine III, RWTH-University Hospital Aachen, Aachen, Germany
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Abstract
During hepatitis B virus (HBV) infection, at least four antigen-antibody systems are observed: HBsAg and anti-HBs; preS antigen and anti-preS antibody; HBcAg and anti-HBc; and HBeAg and anti-HBe. Through the examination of these antigen-antibody systems, hepatitis B infection is diagnosed and the course of the disorder may be observed. Although the serologic findings that allow both the diagnosis of HBV infection as well as assessing of its clinical course are already well established, the dynamics of viral proteins expression and of the antibodies production may vary during the infection natural course. This causes the HBV infection to be occasionally associated with the presence of uncommon serological profiles, which could lead to doubts in the interpretation of results or suspicion of a serological result being incorrect. This paper is dedicated to the discussion of some of these profiles and their significance.
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Yan CH, Zhao CY, Ding H, Peng YQ, Jin PY, Yan L, Zhuang H, Li T. Hepatitis B virus basal core promoter mutations A1762T/G1764A are associated with genotype C and a low serum HBsAg level in chronically-infected HBeAg-positive Chinese patients. Antiviral Res 2012; 96:108-14. [PMID: 22960603 DOI: 10.1016/j.antiviral.2012.08.009] [Citation(s) in RCA: 16] [Impact Index Per Article: 1.2] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/09/2012] [Revised: 08/22/2012] [Accepted: 08/23/2012] [Indexed: 12/16/2022]
Abstract
The present study was aimed to obtain baseline information of basal core promoter A1762T/G1764A and precore G1896A mutations of hepatitis B virus (HBV) in 192 HBeAg-positive chronically-infected Chinese patients, who were potential candidates for antiviral treatment. The detection of these mutations (including minor mutant subpopulations) was achieved by direct sequencing, whose sensitivity for minor mutant subpopulations identification was confirmed by clone sequencing. Patients enrolled were infected with either genotype B (46.35%) or C (53.65%) HBV identified by routine tests in our laboratory. The A1762T/G1764A or G1896A mutations were detected in 125specimens (125/192, 65.10%), in which 77 (77/125, 61.60%) existed as subpopulations. The A1762T/G1764A mutations were found to be more prevalent in genotype C than that in genotype B HBV [62.14% (64/103) vs. 20.22% (18/89), P<0.0001]. There is no statistically significant link between G1896A and genotypes. The emergence of A1762T/G1764A mutations was also found to be associated with an older age, an elevated ALT/AST level, and a lower HBsAg level in serum [wild-type vs. mutant: 4.57 (3.46-5.42) vs. 3.93 (2.51-5.36), P<0.0001]. In conclusion, HBV basal core promoter mutations A1762T/G1764A are associated with genotype C and a low serum HBsAg level in chronically-infected HBeAg-positive Chinese patients.
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Affiliation(s)
- Chun-Hui Yan
- Department of Microbiology and Infectious Disease Center, School of Basic Medical Sciences, Peking University, 38 Xueyuan Road, Haidian District, Beijing 100191, PR China
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Malmström S, Larsson SB, Hannoun C, Lindh M. Hepatitis B viral DNA decline at loss of HBeAg is mainly explained by reduced cccDNA load--down-regulated transcription of PgRNA has limited impact. PLoS One 2012; 7:e36349. [PMID: 22911677 PMCID: PMC3401194 DOI: 10.1371/journal.pone.0036349] [Citation(s) in RCA: 36] [Impact Index Per Article: 2.8] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/06/2012] [Accepted: 04/03/2012] [Indexed: 01/01/2023] Open
Abstract
Background Quantification of hepatitis B virus (HBV) DNA and surface antigen (HBsAg) serum levels have become increasingly important for the assessment of clinical stage and response to treatment for chronic hepatitis B. Effective immune clearance results in reduction of viremia by 4–5 log units and HBsAg levels by 2 log, but these processes are not well understood. Thus, it is uncertain to what extent mechanisms that inhibit transcription of the pregenomic RNA (pgRNA), an RNA intermediate, contribute to suppression of viremia. Likewise, it is unclear if transcriptional regulation is important for the excessive production of surface antigen (HBsAg) that is a hallmark of HBV infection. Methods HBV RNA and cccDNA were quantified in 19 liver biopsies from patients with chronic HBV infection, as well as in transfected Huh7.5 cells and in PLC/PRF/5 cells carrying integrated HBV genome. Results Patients negative for HBeAg had 2.15 log lower levels of cccDNA in liver tissue, 4.84 log lower serum levels of HBV DNA and 1.45 log lower serum levels of HBsAg, than HBeAg-positive patients. The pgRNA in liver tissue correlated strongly with cccDNA (R2 = 0.87, p<0.0001) and HBV DNA levels in serum (R2 = 0.81, p<0.0001), whereas S-RNA correlated strongly with cccDNA (R2 = 0.65, p<0.0001) and HBsAg levels (R2 = 0.57, p = 0.0003). The S-RNA/pgRNA ratio was higher in HBeAg-negative patients (ratio 40 vs. 3, p = 0.01) and in PLC/PRF/5 cells, and was in transfected Huh7.5 cells not influenced by mutations in the HBV core promoter. Conclusion The reduction of viremia that is observed after loss of HBeAg was mainly explained by reduced cccDNA load in the liver, whereas the contribution of down-regulation of pgRNA transcription was relatively small. Enhanced transcription of S-RNA does not explain excessive production of HBsAg.
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Affiliation(s)
- Sebastian Malmström
- Department of Infectious Diseases, University of Gothenburg, Gothenburg, Sweden
| | - Simon B. Larsson
- Department of Infectious Diseases, University of Gothenburg, Gothenburg, Sweden
| | - Charles Hannoun
- Department of Infectious Diseases, University of Gothenburg, Gothenburg, Sweden
| | - Magnus Lindh
- Department of Infectious Diseases, University of Gothenburg, Gothenburg, Sweden
- * E-mail:
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