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Zhang J, Wang Q, Yuan W, Li J, Yuan Q, Zhang J, Xia N, Wang Y, Li J, Tong S. Both middle and large envelope proteins can mediate neutralization of hepatitis B virus infectivity by anti-preS2 antibodies: escape by naturally occurring preS2 deletions. J Virol 2024; 98:e0192923. [PMID: 39078152 PMCID: PMC11334434 DOI: 10.1128/jvi.01929-23] [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: 12/09/2023] [Accepted: 07/02/2024] [Indexed: 07/31/2024] Open
Abstract
Hepatitis B virus (HBV) expresses co-terminal large (L), middle (M), and small (S) envelope proteins containing preS1/preS2/S, preS2/S, and S domain alone, respectively. S and preS1 domains mediate sequential virion attachment to heparan sulfate proteoglycans and sodium taurocholate cotransporting polypeptide (NTCP), respectively, which can be blocked by anti-S and anti-preS1 antibodies. How anti-preS2 antibodies neutralize HBV infectivity remains enigmatic. The late stage of chronic HBV infection often selects for mutated preS2 translation initiation codon to prevent M protein expression, or in-frame preS2 deletions to shorten both L and M proteins. When introduced to infectious clone of genotype C or D, both M-minus mutations and most 5' preS2 deletions sustained virion production. Such mutant progeny viral particles were infectious in NTCP-reconstituted HepG2 cells. Neutralization experiments were performed on the genotype D clone. Although remaining susceptible to anti-preS1 and anti-S neutralizing antibodies, M-minus mutants were only partially neutralized by two anti-preS2 antibodies tested while preS2 deletion mutants were resistant. By infection experiments using viral particles with lost versus increased M protein expression, or a neutralization escaping preS2 deletion only present on L or M protein, we found that both full-length L and M proteins contributed to virus neutralization by the two anti-preS2 antibodies. Thus, immune escape could be a driving force for the selection of M-minus mutations, and especially preS2 deletions. The fact that both L and M proteins could mediate neutralization by anti-preS2 antibodies may shed light on the underlying molecular mechanism.IMPORTANCEThe large (L), middle (M), and small (S) envelope proteins of hepatitis B virus (HBV) contain preS1/preS2/S, preS2/S, and S domain alone, respectively. The discovery of heparan sulfate proteoglycans and sodium taurocholate cotransporting polypeptide (NTCP) as the low- and high-affinity HBV receptors could explain neutralizing potential of anti-S and anti-preS1 antibodies, respectively, but how anti-preS2 neutralizing antibodies work remains enigmatic. In this study, we found two M-minus mutants in the context of genotype D partially escaped two anti-preS2 neutralizing antibodies in NTCP-reconstituted HepG2 cells, while several naturally occurring preS2 deletion mutants escaped both antibodies. By point mutations to eliminate or enhance M protein expression, and by introducing preS2 deletion selectively to L or M protein, we found binding of anti-preS2 antibodies to both L and M proteins contributed to neutralization of wild-type HBV infectivity. Our finding may shed light on the possible mechanism(s) whereby anti-preS2 antibodies neutralize HBV infectivity.
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Affiliation(s)
- Jing Zhang
- Department of Pathobiology, Key Laboratory of Medical Molecular Virology, School of Basic Medical Sciences, Fudan University, Shanghai, China
| | - Qianru Wang
- Department of Pathobiology, Key Laboratory of Medical Molecular Virology, School of Basic Medical Sciences, Fudan University, Shanghai, China
| | - Wenqing Yuan
- Liver Research Center, Rhode Island Hospital, The Warren Alpert School of Medicine, Brown University, Providence, Rhode Island, USA
| | - Jing Li
- Liver Research Center, Rhode Island Hospital, The Warren Alpert School of Medicine, Brown University, Providence, Rhode Island, USA
- Department of Infectious Diseases, Huashan Hospital, Fudan University, Shanghai, China
| | - Quan Yuan
- State Key Laboratory of Molecular Vaccinology and Molecular Diagnostics, School of Public Health, Xiamen University, Xiamen, China
| | - Jiming Zhang
- Department of Infectious Diseases, Huashan Hospital, Fudan University, Shanghai, China
| | - Ningshao Xia
- State Key Laboratory of Molecular Vaccinology and Molecular Diagnostics, School of Public Health, Xiamen University, Xiamen, China
| | - Yongxiang Wang
- Department of Pathobiology, Key Laboratory of Medical Molecular Virology, School of Basic Medical Sciences, Fudan University, Shanghai, China
| | - Jisu Li
- Liver Research Center, Rhode Island Hospital, The Warren Alpert School of Medicine, Brown University, Providence, Rhode Island, USA
| | - Shuping Tong
- Department of Pathobiology, Key Laboratory of Medical Molecular Virology, School of Basic Medical Sciences, Fudan University, Shanghai, China
- Liver Research Center, Rhode Island Hospital, The Warren Alpert School of Medicine, Brown University, Providence, Rhode Island, USA
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Dopico E, Vila M, Tabernero D, Gregori J, Rando-Segura A, Pacín-Ruíz B, Guerrero L, Ubillos I, Martínez MJ, Costa J, Quer J, Pérez-Garreta J, González-Sánchez A, Antón A, Pumarola T, Riveiro-Barciela M, Ferrer-Costa R, Buti M, Rodríguez-Frías F, Cortese MF. Genotyping Hepatitis B virus by Next-Generation Sequencing: Detection of Mixed Infections and Analysis of Sequence Conservation. Int J Mol Sci 2024; 25:5481. [PMID: 38791519 PMCID: PMC11122360 DOI: 10.3390/ijms25105481] [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: 03/29/2024] [Revised: 05/01/2024] [Accepted: 05/13/2024] [Indexed: 05/26/2024] Open
Abstract
Our aim was to develop an accurate, highly sensitive method for HBV genotype determination and detection of genotype mixtures. We examined the preS and 5' end of the HBV X gene (5X) regions of the HBV genome using next-generation sequencing (NGS). The 1852 haplotypes obtained were subjected to genotyping via the Distance-Based discrimination method (DB Rule) using two sets of 95 reference sequences of genotypes A-H. In clinical samples from 125 patients, the main genotypes were A, D, F and H in Caucasian, B and C in Asian and A and E in Sub-Saharan patients. Genotype mixtures were identified in 28 (22.40%) cases, and potential intergenotypic recombination was observed in 29 (23.20%) cases. Furthermore, we evaluated sequence conservation among haplotypes classified into genotypes A, C, D, and E by computing the information content. The preS haplotypes exhibited limited shared conserved regions, whereas the 5X haplotypes revealed two groups of conserved regions across the genotypes assessed. In conclusion, we developed an NGS-based HBV genotyping method utilizing the DB Rule for genotype classification. We identified two regions conserved across different genotypes at 5X, offering promising targets for RNA interference-based antiviral therapies.
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Affiliation(s)
- Eva Dopico
- Department of Microbiology, Metropolitana Sud Territorial Clinical Laboratory, Bellvitge University Hospital, Institut Català de la Salut (ICS), 08907 Hospitalet de Llobregat, Spain; (E.D.); (L.G.); (I.U.)
- Bellvitge Biomedical Research Institute (IDIBELL), 08908 Hospitalet de Llobregat, Spain
| | - Marta Vila
- Liver Unit, Microbiology Department, Vall d’Hebron Institut de Recerca (VHIR), Vall d’Hebron Hospital Universitari, Vall d’Hebron Barcelona Hospital Campus, 08035 Barcelona, Spain; (M.V.); (A.R.-S.); (B.P.-R.); (J.P.-G.); (M.F.C.)
| | - David Tabernero
- Liver Unit, Microbiology Department, Vall d’Hebron Institut de Recerca (VHIR), Vall d’Hebron Hospital Universitari, Vall d’Hebron Barcelona Hospital Campus, 08035 Barcelona, Spain; (M.V.); (A.R.-S.); (B.P.-R.); (J.P.-G.); (M.F.C.)
- Centro de Investigación Biomédica en Red de Enfermedades Hepáticas y Digestivas (CIBERehd), Instituto de Salud Carlos III, 28029 Madrid, Spain; (J.Q.); (M.R.-B.); (M.B.); (F.R.-F.)
- Liver Diseases-Viral Hepatitis, Liver Unit, Vall d’Hebron Institut de Recerca (VHIR), Vall d’Hebron Hospital Universitari, Vall d’Hebron Barcelona Hospital Campus, 08035 Barcelona, Spain;
| | - Josep Gregori
- Liver Diseases-Viral Hepatitis, Liver Unit, Vall d’Hebron Institut de Recerca (VHIR), Vall d’Hebron Hospital Universitari, Vall d’Hebron Barcelona Hospital Campus, 08035 Barcelona, Spain;
| | - Ariadna Rando-Segura
- Liver Unit, Microbiology Department, Vall d’Hebron Institut de Recerca (VHIR), Vall d’Hebron Hospital Universitari, Vall d’Hebron Barcelona Hospital Campus, 08035 Barcelona, Spain; (M.V.); (A.R.-S.); (B.P.-R.); (J.P.-G.); (M.F.C.)
- Centro de Investigación Biomédica en Red de Enfermedades Hepáticas y Digestivas (CIBERehd), Instituto de Salud Carlos III, 28029 Madrid, Spain; (J.Q.); (M.R.-B.); (M.B.); (F.R.-F.)
- Virology Section, Microbiology Department, Vall d’Hebron Hospital Universitari, Vall d’Hebron Barcelona Hospital Campus, 08035 Barcelona, Spain; (A.G.-S.); (A.A.); (T.P.)
| | - Beatriz Pacín-Ruíz
- Liver Unit, Microbiology Department, Vall d’Hebron Institut de Recerca (VHIR), Vall d’Hebron Hospital Universitari, Vall d’Hebron Barcelona Hospital Campus, 08035 Barcelona, Spain; (M.V.); (A.R.-S.); (B.P.-R.); (J.P.-G.); (M.F.C.)
- Centro de Investigación Biomédica en Red de Enfermedades Hepáticas y Digestivas (CIBERehd), Instituto de Salud Carlos III, 28029 Madrid, Spain; (J.Q.); (M.R.-B.); (M.B.); (F.R.-F.)
| | - Laura Guerrero
- Department of Microbiology, Metropolitana Sud Territorial Clinical Laboratory, Bellvitge University Hospital, Institut Català de la Salut (ICS), 08907 Hospitalet de Llobregat, Spain; (E.D.); (L.G.); (I.U.)
| | - Itziar Ubillos
- Department of Microbiology, Metropolitana Sud Territorial Clinical Laboratory, Bellvitge University Hospital, Institut Català de la Salut (ICS), 08907 Hospitalet de Llobregat, Spain; (E.D.); (L.G.); (I.U.)
| | - Miguel J. Martínez
- Department of Microbiology, Hospital Clínic, Universitat de Barcelona, 08036 Barcelona, Spain; (M.J.M.); (J.C.)
- ISGlobal, Hospital Clínic, Universitat de Barcelona, 08036 Barcelona, Spain
- Centro de Investigación Biomédica en Red de Enfermedades Infecciosas (CIBERINFEC), Instituto de Salud Carlos III, 28029 Madrid, Spain
| | - Josep Costa
- Department of Microbiology, Hospital Clínic, Universitat de Barcelona, 08036 Barcelona, Spain; (M.J.M.); (J.C.)
| | - Josep Quer
- Centro de Investigación Biomédica en Red de Enfermedades Hepáticas y Digestivas (CIBERehd), Instituto de Salud Carlos III, 28029 Madrid, Spain; (J.Q.); (M.R.-B.); (M.B.); (F.R.-F.)
- Liver Diseases-Viral Hepatitis, Liver Unit, Vall d’Hebron Institut de Recerca (VHIR), Vall d’Hebron Hospital Universitari, Vall d’Hebron Barcelona Hospital Campus, 08035 Barcelona, Spain;
- Biochemistry and Molecular Biology Department, Universitat Autònoma de Barcelona (UAB), 08193 Bellaterra, Spain
| | - Javier Pérez-Garreta
- Liver Unit, Microbiology Department, Vall d’Hebron Institut de Recerca (VHIR), Vall d’Hebron Hospital Universitari, Vall d’Hebron Barcelona Hospital Campus, 08035 Barcelona, Spain; (M.V.); (A.R.-S.); (B.P.-R.); (J.P.-G.); (M.F.C.)
| | - Alejandra González-Sánchez
- Virology Section, Microbiology Department, Vall d’Hebron Hospital Universitari, Vall d’Hebron Barcelona Hospital Campus, 08035 Barcelona, Spain; (A.G.-S.); (A.A.); (T.P.)
- Centro de Investigación Biomédica en Red de Enfermedades Infecciosas (CIBERINFEC), Instituto de Salud Carlos III, 28029 Madrid, Spain
- Respiratory Virus Unit, Microbiology Department, Vall d’Hebron Institut de Recerca (VHIR), Vall d’Hebron Hospital Universitari, Vall d’Hebron Barcelona Hospital Campus, 08035 Barcelona, Spain
| | - Andrés Antón
- Virology Section, Microbiology Department, Vall d’Hebron Hospital Universitari, Vall d’Hebron Barcelona Hospital Campus, 08035 Barcelona, Spain; (A.G.-S.); (A.A.); (T.P.)
- Centro de Investigación Biomédica en Red de Enfermedades Infecciosas (CIBERINFEC), Instituto de Salud Carlos III, 28029 Madrid, Spain
- Respiratory Virus Unit, Microbiology Department, Vall d’Hebron Institut de Recerca (VHIR), Vall d’Hebron Hospital Universitari, Vall d’Hebron Barcelona Hospital Campus, 08035 Barcelona, Spain
| | - Tomás Pumarola
- Virology Section, Microbiology Department, Vall d’Hebron Hospital Universitari, Vall d’Hebron Barcelona Hospital Campus, 08035 Barcelona, Spain; (A.G.-S.); (A.A.); (T.P.)
- Centro de Investigación Biomédica en Red de Enfermedades Infecciosas (CIBERINFEC), Instituto de Salud Carlos III, 28029 Madrid, Spain
| | - Mar Riveiro-Barciela
- Centro de Investigación Biomédica en Red de Enfermedades Hepáticas y Digestivas (CIBERehd), Instituto de Salud Carlos III, 28029 Madrid, Spain; (J.Q.); (M.R.-B.); (M.B.); (F.R.-F.)
- Liver Unit, Internal Medicine Department, Vall d’Hebron Hospital Universitari, Vall d’Hebron Barcelona Hospital Campus, 08035 Barcelona, Spain
| | - Roser Ferrer-Costa
- Clinical Biochemistry, Drug Delivery and Therapy (CB-DDT) Research Group, Vall d’Hebron Institut de Recerca (VHIR), Vall d’Hebron Hospital Universitari, Vall d’Hebron Barcelona Hospital Campus, 08035 Barcelona, Spain;
- Biochemistry Department, Vall d’Hebron Hospital Universitari, Vall d’Hebron Barcelona Hospital Campus, 08035 Barcelona, Spain
| | - Maria Buti
- Centro de Investigación Biomédica en Red de Enfermedades Hepáticas y Digestivas (CIBERehd), Instituto de Salud Carlos III, 28029 Madrid, Spain; (J.Q.); (M.R.-B.); (M.B.); (F.R.-F.)
- Liver Unit, Internal Medicine Department, Vall d’Hebron Hospital Universitari, Vall d’Hebron Barcelona Hospital Campus, 08035 Barcelona, Spain
| | - Francisco Rodríguez-Frías
- Centro de Investigación Biomédica en Red de Enfermedades Hepáticas y Digestivas (CIBERehd), Instituto de Salud Carlos III, 28029 Madrid, Spain; (J.Q.); (M.R.-B.); (M.B.); (F.R.-F.)
- Clinical Biochemistry, Drug Delivery and Therapy (CB-DDT) Research Group, Vall d’Hebron Institut de Recerca (VHIR), Vall d’Hebron Hospital Universitari, Vall d’Hebron Barcelona Hospital Campus, 08035 Barcelona, Spain;
- Department of Basic Sciences, Universitat Internacional de Catalunya, 08017 Barcelona, Spain
| | - Maria Francesca Cortese
- Liver Unit, Microbiology Department, Vall d’Hebron Institut de Recerca (VHIR), Vall d’Hebron Hospital Universitari, Vall d’Hebron Barcelona Hospital Campus, 08035 Barcelona, Spain; (M.V.); (A.R.-S.); (B.P.-R.); (J.P.-G.); (M.F.C.)
- Centro de Investigación Biomédica en Red de Enfermedades Hepáticas y Digestivas (CIBERehd), Instituto de Salud Carlos III, 28029 Madrid, Spain; (J.Q.); (M.R.-B.); (M.B.); (F.R.-F.)
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Han JJ, Hu YA, Nan Y, Chen Y, Yang YL. Decreased expression of HBV surface antigen (HBsAg) with sK122R and sV96A co-mutation is associated with an ineffective antibody response in a chronic hepatitis B patient. INFECTION, GENETICS AND EVOLUTION : JOURNAL OF MOLECULAR EPIDEMIOLOGY AND EVOLUTIONARY GENETICS IN INFECTIOUS DISEASES 2023; 111:105431. [PMID: 37015318 DOI: 10.1016/j.meegid.2023.105431] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Subscribe] [Scholar Register] [Received: 04/02/2022] [Revised: 02/01/2023] [Accepted: 03/29/2023] [Indexed: 04/04/2023]
Abstract
BACKGROUND Emergence and predominance of hepatitis B virus (HBV) variants carrying S gene mutations frequently occur in HBV-infected individuals. Here, coexistent serum anti-HBsAg antibody (HBsAb) and HBV surface antigen (HBsAg) were detected in a chronic HBV patient. The patient's HBsAg proteins possessed amino acid substitutions sK122R and sV96A. We reported this case and conducted relevant studies to investigate differences in expression levels and antibody neutralization of HBsAg proteins bearing sK122R and sV96A amino acid substitutions to explore causes of antigen-antibody coexistence in a chronic hepatitis B patient. STUDY DESIGN We first sequenced the S gene from HBV present within the patient's serum. Based on the S gene sequence, we cloned wild-type and mutated S gene sequences via site-directed mutagenesis to construct expression plasmids pJW4303-WT (wild-type), pJW4303-sV96A, pJW4303-sK122R, and pJW4303-sV96A-sK122R. Plasmids were transfected into HEK 293 T cells then culture supernatants and cells were collected. Collected cells and supernatants were next subjected to a series of quantitative and functional tests to assess expression and neutralization characteristics of wild-type and mutant HBsAg proteins. RESULTS Based on quantification of HBsAg expression in cells transfected with the four plasmids, HBsAg-sK122R-sV96A was more intracellularly retained and less secreted than HBsAg-sV96A single-mutant protein and WT. Neutralization ability of serum from chronic HBV patient against culture supernatants containing recombinant HBsAg proteins were ranked from highest to lowest as HBsAg-sV96A, HBsAg-sV96A-sK122R, and HBsAg-sK122R. However, no significant differences of neutralization efficiency by high-potency antibodies from HBV-vaccinees against these three mutant proteins were observed. CONCLUSIONS The levels of HBsAg proteins with amino acid substitutions sV96A-sK122R were greatly reduced in culture supernatants but were apparently increased in the intracellular fraction. This may account for the higher levels of HBV replication in patients. HBsAg neutralization by HBsAb in this patient may have been compromised by the HBsAg sK122R amino acid substitution, suggesting that antibodies produced by the patient had lost their HBV-neutralizing effect.
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Affiliation(s)
- Jing-Jing Han
- Department of Anesthesiology, The First Affiliated Hospital of Nanjing Medical University, Nanjing 210029, Jiangsu Province, China
| | - Yu-An Hu
- Basic Medical Laboratory, Institute of Clinical Laboratory Science, Nanjing Jinling Hospital, Affiliated Hospital of Medical School, Nanjing University, Nanjing 210002, Jiangsu Province, China
| | - Yuchen Nan
- Department of Preventive Veterinary Medicine, College of Veterinary Medicine, Northwest A&F University, Yangling 712100, Shaanxi Province, China
| | - Yu Chen
- Department of Anesthesiology, The First Affiliated Hospital of Nanjing Medical University, Nanjing 210029, Jiangsu Province, China
| | - Yong-Lin Yang
- Department of Infectious Disease, The Affiliated Taizhou People's Hospital of Nanjing Medical University, Taizhou 225300, Jiangsu Province, China.
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Efficient degradation of hydroquinone by a metabolically engineered Pseudarthrobacter sulfonivorans strain. Arch Microbiol 2022; 204:588. [PMID: 36048304 DOI: 10.1007/s00203-022-03214-z] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.7] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/06/2022] [Revised: 08/18/2022] [Accepted: 08/22/2022] [Indexed: 11/02/2022]
Abstract
Pseudarthrobacter sulfonivorans strain Ar51 can degrade crude oil and multi-substituted benzene compounds efficiently at low temperatures. However, it cannot degrade hydroquinone, which is a key intermediate in the degradation of several other compounds of environmental importance, such as 4-nitrophenol, g-hexachlorocyclohexane, 4-hydroxyacetophenone and 4-aminophenol. Here we co-expressed the two subunits of hydroquinone dioxygenase from Sphingomonas sp. strain TTNP3 with different promoters in the strain Ar51. The strain with 2 hdnO promoters exhibited the strongest hydroquinone catabolic activity. However, in the absence of antibiotic selection this ability to degrade hydroquinone was lost due to plasmid instability. Consequently, we constructed a hisD knockout strain, which was unable to synthesise histidine. By introducing the hisD gene onto the plasmid, the ability to degrade hydroquinone in the absence of antibiotic selection was stabilised. In addition, to make the strain more stable for industrial applications, we knocked out the recA gene and integrated the hydroquinone dioxygenase genes at this chromosomal locus. This strain exhibited the strongest activity in catabolizing hydroquinone, up to 470 mg/L in 16 h without antibiotic selection. In addition, this activity was shown to be stable when the strain has cultured in medium without antibiotic selection after 20 passages.
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Jia JA, Zhang S, Bai X, Fang M, Chen S, Liang X, Zhu S, Wong DKH, Zhang A, Feng J, Sun F, Gao C. Sparse logistic regression revealed the associations between HBV PreS quasispecies and hepatocellular carcinoma. Virol J 2022; 19:114. [PMID: 35765099 PMCID: PMC9238101 DOI: 10.1186/s12985-022-01836-9] [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: 10/06/2021] [Accepted: 05/25/2022] [Indexed: 11/10/2022] Open
Abstract
BACKGROUND Chronic infection with hepatitis B virus (HBV) has been proved highly associated with the development of hepatocellular carcinoma (HCC). AIMS The purpose of the study is to investigate the association between HBV preS region quasispecies and HCC development, as well as to develop HCC diagnosis model using HBV preS region quasispecies. METHODS A total of 104 chronic hepatitis B (CHB) patients and 117 HBV-related HCC patients were enrolled. HBV preS region was sequenced using next generation sequencing (NGS) and the nucleotide entropy was calculated for quasispecies evaluation. Sparse logistic regression (SLR) was used to predict HCC development and prediction performances were evaluated using receiver operating characteristic curves. RESULTS Entropy of HBV preS1, preS2 regions and several nucleotide points showed significant divergence between CHB and HCC patients. Using SLR, the classification of HCC/CHB groups achieved a mean area under the receiver operating characteristic curve (AUC) of 0.883 in the training data and 0.795 in the test data. The prediction model was also validated by a completely independent dataset from Hong Kong. The 10 selected nucleotide positions showed significantly different entropy between CHB and HCC patients. The HBV quasispecies also classified three clinical parameters, including HBeAg, HBVDNA, and Alkaline phosphatase (ALP) with the AUC value greater than 0.6 in the test data. CONCLUSIONS Using NGS and SLR, the association between HBV preS region nucleotide entropy and HCC development was validated in our study and this could promote the understanding of HCC progression mechanism.
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Affiliation(s)
- Jian-an Jia
- Department of Laboratory Medicine, Eastern Hepatobiliary Surgery Hospital, Navy Military Medical University, Shanghai, 200438 China
- Department of Laboratory Medicine, The 901th Hospital of Joint Logistics Support Force of Chinese People’s Liberation Army, Hefei, 230031 China
| | - Shuqin Zhang
- Centre for Computational Systems Biology, School of Mathematical Sciences, Fudan University, Shanghai, 200433 China
| | - Xin Bai
- Molecular and Computational Program, Department of Biological Sciences, University of Southern California, 1050 Childs Way, Los Angeles, 90089 USA
| | - Meng Fang
- Department of Laboratory Medicine, Eastern Hepatobiliary Surgery Hospital, Navy Military Medical University, Shanghai, 200438 China
| | - Shipeng Chen
- Department of Laboratory Medicine, Eastern Hepatobiliary Surgery Hospital, Navy Military Medical University, Shanghai, 200438 China
| | - Xiaotao Liang
- Department of Computer Science, Fudan University, Shanghai, 200433 China
| | - Shanfeng Zhu
- Department of Computer Science, Fudan University, Shanghai, 200433 China
- Institute of Science and Technology for Brain-Inspired Intelligence, Fudan University, Shanghai, 200433 China
| | - Danny Ka-Ho Wong
- State Key Laboratory for Liver Research, The University of Hong Kong, Hong Kong SAR, China
| | - Anye Zhang
- Department of Medicine, The Eighth Affiliated Hospital, Sun Yat-Sen University, Shenzhen, 518033 China
| | - Jianfeng Feng
- Institute of Science and Technology for Brain-Inspired Intelligence, Fudan University, Shanghai, 200433 China
- Department of Computer Science, University of Warwick, Coventry, CV4 7AL UK
| | - Fengzhu Sun
- Molecular and Computational Program, Department of Biological Sciences, University of Southern California, 1050 Childs Way, Los Angeles, 90089 USA
| | - Chunfang Gao
- Department of Laboratory Medicine, Eastern Hepatobiliary Surgery Hospital, Navy Military Medical University, Shanghai, 200438 China
- Clinical Laboratory Medicine Center, Yueyang Hospital of Integrated Traditional Chinese and Western Medicine, Shanghai University of Traditional Chinese Medicine, Shanghai, 200437 China
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Hepatitis B Virus Variants with Multiple Insertions and/or Deletions in the X Open Reading Frame 3′ End: Common Members of Viral Quasispecies in Chronic Hepatitis B Patients. Biomedicines 2022; 10:biomedicines10051194. [PMID: 35625929 PMCID: PMC9139148 DOI: 10.3390/biomedicines10051194] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/31/2022] [Revised: 05/18/2022] [Accepted: 05/19/2022] [Indexed: 11/16/2022] Open
Abstract
Deletions in the 3′ end region of the hepatitis B virus (HBV) X open reading frame (HBX) may affect the core promoter (Cp) and have been frequently associated with hepatocellular carcinoma (HCC). The aim of this study was to investigate the presence of variants with deletions and/or insertions (Indels) in this region in the quasispecies of 50 chronic hepatitis B (CHB) patients without HCC. We identified 103 different Indels in 47 (94%) patients, in a median of 3.4% of their reads (IQR, 1.3–8.4%), and 25% (IQR, 13.1–40.7%) of unique sequences identified in each quasispecies (haplotypes). Of those Indels, 101 (98.1%) caused 44 different altered stop codons, the most commonly observed were at positions 128, 129, 135, and 362 (putative position). Moreover, 39 (37.9%) Indels altered the TATA-like box (TA) sequences of Cp; the most commonly observed caused TA2 + TA3 fusion, creating a new putative canonical TATA box. Four (8%) patients developed negative clinical outcomes after a median follow-up of 9.4 (8.7–12) years. In conclusion, we observed variants with Indels in the HBX 3′ end in the vast majority of our CHB patients, some of them encoding alternative versions of HBx with potential functional roles, and/or alterations in the regulation of transcription.
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Nishida Y, Imamura M, Teraoka Y, Morio K, Fujino H, Ono A, Nakahara T, Murakami E, Yamauchi M, Kawaoka T, Miki D, Tsuge M, Hiramatsu A, Abe-Chayama H, Hayes CN, Aikata H, Sasaki N, Sekiguchi T, Kinukawa H, Yoshimura T, Chayama K. Serum PreS1 and HBsAg ratio reflects liver fibrosis and predicts the development of hepatocellular carcinoma in chronic hepatitis B patients. J Viral Hepat 2021; 28:1304-1311. [PMID: 34105859 DOI: 10.1111/jvh.13557] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 03/10/2021] [Revised: 05/11/2021] [Accepted: 05/20/2021] [Indexed: 12/09/2022]
Abstract
While the preS1 region of the large hepatitis B surface protein plays an essential role in hepatitis B virus (HBV) infection, the effect of preS1 on liver fibrosis and hepatocarcinogenesis in chronic hepatitis B (CHB) patients is not well known. In this study, we measured serum preS1 levels by chemiluminescent immunoassay technology in 690 CHB patients and evaluated the correlation between serum preS1 levels and HBV, liver function markers and liver inflammation, fibrosis assessed by histological findings. Predictive factors for hepatocellular carcinoma (HCC) development in patients who had no previous history of HCC at the time of preS1 level measurement were also analysed. Median hepatitis B surface antigen (HBsAg) and preS1 levels were 3.08 log IU/mL and 98 ng/mL, respectively. PreS1 values were significantly correlated with serum HBsAg (p <0.001), hepatitis B core-related antigen (HBcrAg) (p <0.001) and HBV DNA levels (p <0.01). PreS1 values were also significantly correlated with serum alanine aminotransferase levels (p <0.001) and were significantly higher in patients who had higher grading of liver inflammatory activity (p <0.05). HBsAg level was correlated, but preS1/HBsAg ratio reflected liver fibrosis staging more directly than HBsAg alone. Multivariate analysis identified age ≥53 years (hazard ratio [HR], 18.360 for <53 years; p = 0.021) and preS1/HBsAg ratio ≥0.12 (HR, 6.205 for <0.12; p = 0.040) as significant and independent factors for HCC development in CHB patients. The preS1/HBsAg ratio directly reflects liver fibrosis, and the ratio might be a predictive marker for HCC development in CHB patients.
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Affiliation(s)
- Yuno Nishida
- Department of Gastroenterology and Metabolism, Graduate School of Biomedical and Health Science, Hiroshima, Japan.,Research Center for Hepatology and Gastroenterology, Hiroshima University, Hiroshima, Japan
| | - Michio Imamura
- Department of Gastroenterology and Metabolism, Graduate School of Biomedical and Health Science, Hiroshima, Japan.,Research Center for Hepatology and Gastroenterology, Hiroshima University, Hiroshima, Japan
| | - Yuji Teraoka
- Department of Gastroenterology and Metabolism, Graduate School of Biomedical and Health Science, Hiroshima, Japan.,Research Center for Hepatology and Gastroenterology, Hiroshima University, Hiroshima, Japan
| | - Kei Morio
- Department of Gastroenterology and Metabolism, Graduate School of Biomedical and Health Science, Hiroshima, Japan.,Research Center for Hepatology and Gastroenterology, Hiroshima University, Hiroshima, Japan
| | - Hatsue Fujino
- Department of Gastroenterology and Metabolism, Graduate School of Biomedical and Health Science, Hiroshima, Japan.,Research Center for Hepatology and Gastroenterology, Hiroshima University, Hiroshima, Japan
| | - Atsushi Ono
- Department of Gastroenterology and Metabolism, Graduate School of Biomedical and Health Science, Hiroshima, Japan.,Research Center for Hepatology and Gastroenterology, Hiroshima University, Hiroshima, Japan
| | - Takashi Nakahara
- Department of Gastroenterology and Metabolism, Graduate School of Biomedical and Health Science, Hiroshima, Japan.,Research Center for Hepatology and Gastroenterology, Hiroshima University, Hiroshima, Japan
| | - Eisuke Murakami
- Department of Gastroenterology and Metabolism, Graduate School of Biomedical and Health Science, Hiroshima, Japan.,Research Center for Hepatology and Gastroenterology, Hiroshima University, Hiroshima, Japan
| | - Masami Yamauchi
- Department of Gastroenterology and Metabolism, Graduate School of Biomedical and Health Science, Hiroshima, Japan.,Research Center for Hepatology and Gastroenterology, Hiroshima University, Hiroshima, Japan
| | - Tomokazu Kawaoka
- Department of Gastroenterology and Metabolism, Graduate School of Biomedical and Health Science, Hiroshima, Japan.,Research Center for Hepatology and Gastroenterology, Hiroshima University, Hiroshima, Japan
| | - Daiki Miki
- Department of Gastroenterology and Metabolism, Graduate School of Biomedical and Health Science, Hiroshima, Japan.,Research Center for Hepatology and Gastroenterology, Hiroshima University, Hiroshima, Japan
| | - Masataka Tsuge
- Research Center for Hepatology and Gastroenterology, Hiroshima University, Hiroshima, Japan.,Natural Science Center for Basic Research and Development, Hiroshima University, Hiroshima, Japan
| | - Akira Hiramatsu
- Department of Gastroenterology and Metabolism, Graduate School of Biomedical and Health Science, Hiroshima, Japan.,Research Center for Hepatology and Gastroenterology, Hiroshima University, Hiroshima, Japan
| | - Hiromi Abe-Chayama
- Research Center for Hepatology and Gastroenterology, Hiroshima University, Hiroshima, Japan.,Center for Medical Specialist Graduate Education and Research, Institute of Biomedical and Health Sciences, Hiroshima University, Hiroshima, Japan
| | - C Nelson Hayes
- Department of Gastroenterology and Metabolism, Graduate School of Biomedical and Health Science, Hiroshima, Japan.,Research Center for Hepatology and Gastroenterology, Hiroshima University, Hiroshima, Japan
| | - Hiroshi Aikata
- Department of Gastroenterology and Metabolism, Graduate School of Biomedical and Health Science, Hiroshima, Japan.,Research Center for Hepatology and Gastroenterology, Hiroshima University, Hiroshima, Japan
| | - Noriko Sasaki
- Diagnostics Division, Abbott Japan LLC, Matsudo, Japan
| | | | | | | | - Kazuaki Chayama
- Research Center for Hepatology and Gastroenterology, Hiroshima University, Hiroshima, Japan.,Collaborative Research Laboratory of Medical Innovation, Graduate School of Biomedical and Health Sciences, Hiroshima University, Yokohama, Japan.,Institute of Physical and Chemical Research (RIKEN) Center for Integrative Medical Sciences, Yokohama, Japan
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8
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Jiang X, Chang L, Yan Y, Wang L. Paradoxical HBsAg and anti-HBs coexistence among Chronic HBV Infections: Causes and Consequences. Int J Biol Sci 2021; 17:1125-1137. [PMID: 33867835 PMCID: PMC8040313 DOI: 10.7150/ijbs.55724] [Citation(s) in RCA: 19] [Impact Index Per Article: 4.8] [Reference Citation Analysis] [Abstract] [Key Words] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/10/2020] [Accepted: 03/11/2021] [Indexed: 12/11/2022] Open
Abstract
Hepatitis B surface antigen (HBsAg) and Hepatitis B surface antibody (anti-HBs) were reported simultaneously among Hepatitis B virus (HBV) infections. HBsAg is a specific indicator of acute or chronic HBV infections, while anti-HBs is a protective antibody reflecting the recovery and immunity of hosts. HBsAg and anti-HBs coexist during seroconversion and then form immune complex, which is rare detected in clinical cases. However, with the promotion of vaccination and the application of various antiviral drugs, along with the rapid development of medical technology, the coexistence of HBsAg and anti-HBs has become more prevalent. Mutations in the viral genomes, immune status and genetic factors of hosts may contribute to the coexistence. Novel HBsAg assays, with higher sensitivity and ability to detect mutations or immune complexes, can also yield HBsAg/anti-HBs coexistence. The discovery of coexistence has shattered the idea of traditional serological patterns and raised questions about the effectiveness of vaccines. Worth noting is that HBsAg/anti-HBs double positivity is strongly associated with progressive liver diseases, especially hepatocellular carcinoma. In conclusion, viral mutations, host factors, and methodology impacts can all lead to the coexistence of HBsAg and anti-HBs. This coexistence is not an indicator of improvement, as an increased risk of adverse clinical outcomes still exists.
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Affiliation(s)
- Xinyi Jiang
- National Center for Clinical Laboratories, Beijing Hospital, National Center of Gerontology; Institute of Geriatric Medicine, Chinese Academy of Medical Sciences, Beijing, P.R. China.,Beijing Engineering Research Center of Laboratory Medicine, Beijing Hospital, P.R. China.,Graduate School, Peking Union Medical College, Chinese Academy of Medical Sciences, Beijing, P.R. China
| | - Le Chang
- National Center for Clinical Laboratories, Beijing Hospital, National Center of Gerontology; Institute of Geriatric Medicine, Chinese Academy of Medical Sciences, Beijing, P.R. China.,Beijing Engineering Research Center of Laboratory Medicine, Beijing Hospital, P.R. China.,Graduate School, Peking Union Medical College, Chinese Academy of Medical Sciences, Beijing, P.R. China
| | - Ying Yan
- National Center for Clinical Laboratories, Beijing Hospital, National Center of Gerontology; Institute of Geriatric Medicine, Chinese Academy of Medical Sciences, Beijing, P.R. China.,Beijing Engineering Research Center of Laboratory Medicine, Beijing Hospital, P.R. China.,Graduate School, Peking Union Medical College, Chinese Academy of Medical Sciences, Beijing, P.R. China
| | - Lunan Wang
- National Center for Clinical Laboratories, Beijing Hospital, National Center of Gerontology; Institute of Geriatric Medicine, Chinese Academy of Medical Sciences, Beijing, P.R. China.,Beijing Engineering Research Center of Laboratory Medicine, Beijing Hospital, P.R. China.,Graduate School, Peking Union Medical College, Chinese Academy of Medical Sciences, Beijing, P.R. China
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9
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Li J, Li J, Chen S, Yuan Q, Zhang J, Wu J, Jiang Q, Wang Q, Xia NS, Zhang J, Tong S. Naturally occurring 5' preS1 deletions markedly enhance replication and infectivity of HBV genotype B and genotype C. Gut 2021; 70:575-584. [PMID: 32571971 DOI: 10.1136/gutjnl-2019-320096] [Citation(s) in RCA: 11] [Impact Index Per Article: 2.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 10/16/2019] [Revised: 05/08/2020] [Accepted: 05/26/2020] [Indexed: 12/19/2022]
Abstract
BACKGROUND AND AIMS Deletion of 15-nucleotide or 18-nucleotide (nt) covering preS1 ATG frequently arises during chronic infection with HBV genotypes B and C. Since the second ATG is 33nt downstream, they truncate large (L) envelope protein by 11 residues like wild-type genotype D. This study characterised their functional consequences. METHODS HBV genomes with or without deletion were amplified from a patient with advanced liver fibrosis and assembled into replication competent 1.1mer construct. Deletion, insertion or point mutation was introduced to additional clones of different genotypes. Viral particles concentrated from transfected HepG2 cells were inoculated to sodium taurocholate cotransporting polypeptide (NTCP)-reconstituted HepG2 (HepG2/NTCP) cells or differentiated HepaRG cells, and HBV RNA, DNA, proteins were monitored. RESULTS From transfected HepG2 cells, the 15-nt and 18-nt deletions increased HBV RNA, replicative DNA and extracellular virions. When same number of viral particles was inoculated to HepG2/NTCP cells, the deletion mutants showed higher infectivity. Conversely, HBV infectivity was diminished by putting back the 18nt into naturally occurring genotype C deletion mutants and by adding 33nt to genotype D. Infectivity of full-length genotype C clones was also enhanced by mutating the first ATG codon of the preS1 region but diminished by mutating the second in-frame ATG. Removing N-terminal 11 residues from preS1 peptide 2-59 of genotype C potentiated inhibition of HBV infection and enhanced binding to HepG2/NTCP cells. CONCLUSIONS The 15-nt and 18-nt deletions somehow increase HBV RNA, replicative DNA and virion production. Shortened L protein is more efficient at mediating HBV infection.
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Affiliation(s)
- Jing Li
- Department of Infectious Diseases, Huashan Hospital, Fudan University, Shanghai, China.,Liver Research Center, Rhode Island Hospital and Warren Alpert Medical School of Brown University, Providence, RI, USA
| | - Jisu Li
- Liver Research Center, Rhode Island Hospital and Warren Alpert Medical School of Brown University, Providence, RI, USA
| | - Shiqi Chen
- Department of Infectious Diseases, Huashan Hospital, Fudan University, Shanghai, China
| | - Quan Yuan
- State Key Laboratory of Molecular Vaccinology and Molecular Diagnostics, School of Publich Health, Xiamen University, Xiamen, China
| | - Jing Zhang
- Department of Pathobiology, Key Laboratory of Medical Molecular Virology, School of Basic Medical Sciences, Fudan University, Shanghai, China
| | - Jingwen Wu
- Department of Infectious Diseases, Huashan Hospital, Fudan University, Shanghai, China
| | - Qirong Jiang
- Department of Infectious Diseases, Huashan Hospital, Fudan University, Shanghai, China
| | - Qianru Wang
- Department of Pathobiology, Key Laboratory of Medical Molecular Virology, School of Basic Medical Sciences, Fudan University, Shanghai, China
| | - Ning-Shao Xia
- State Key Laboratory of Molecular Vaccinology and Molecular Diagnostics, School of Publich Health, Xiamen University, Xiamen, China
| | - Jiming Zhang
- Department of Infectious Diseases, Huashan Hospital, Fudan University, Shanghai, China
| | - Shuping Tong
- Liver Research Center, Rhode Island Hospital and Warren Alpert Medical School of Brown University, Providence, RI, USA .,Department of Pathobiology, Key Laboratory of Medical Molecular Virology, School of Basic Medical Sciences, Fudan University, Shanghai, China
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10
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Yuen L, Revill PA, Rosenberg G, Wagner J, Littlejohn M, Bayliss J, Jackson K, Tan SK, Gaggar A, Kitrinos K, Subramanian M, Gane E, Chan HLY, Li X, Bowden S, Locarnini S, Thompson A. HBV variants are common in the 'immune-tolerant' phase of chronic hepatitis B. J Viral Hepat 2020; 27:1061-1070. [PMID: 32384174 DOI: 10.1111/jvh.13318] [Citation(s) in RCA: 5] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 01/08/2020] [Revised: 04/09/2020] [Accepted: 04/23/2020] [Indexed: 12/13/2022]
Abstract
Nucleos(t)ide analogues (NUC) treatment prevents progression of liver fibrosis in subjects with chronic hepatitis B (CHB). However, risk of hepatocellular carcinoma (HCC) persists despite viral suppression. Specific HBV variants have been associated with adverse outcomes, including HCC; however, the frequency of these variants during the seemingly benign immunotolerant (IT) phase is unknown. Next-generation sequencing and detailed virological characterization on a cohort of treatment-naïve IT subjects were performed to determine the frequency of clinically relevant viral variants. Samples from 97 subjects (genotype B/C 55%/45%, median HBV-DNA 8.5 log10 IU/mL, median HBsAg 4.8 log10 IU/mL, median HBeAg 3.6 log10 PEIU/mL) were analysed. Despite subjects being in the IT phase, clinically relevant HBV variants were common at baseline, particularly in the basal core promoter (BCP, overlaps the hepatitis B X (HBx) gene), precore and PreS regions. BCP/HBx variants were independently associated with lower baseline HBeAg, HBsAg and HBV-DNA titres. Precore variants were independently associated with higher baseline ALT. Increased viral diversity was associated with increased age and lower HBV-DNA, HBsAg and HBeAg levels. Low-level (<5%) drug resistance-associated amino acid substitutions in the HBV reverse transcriptase were detected in 9 (9%) subjects at pre-treatment but were not associated with reduced antiviral activity. Future studies should evaluate whether the detection of HBV variant during IT CHB is predictive of progression to immune clearance and poor prognosis, and whether early initiation of antiviral therapy during IT CHB to prevent the selection of HBV variants is clinically beneficial.
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Affiliation(s)
- Lilly Yuen
- Victorian Infectious Diseases Reference Laboratory, Royal Melbourne Hospital, at the Peter Doherty Institute for Infection and Immunity, Melbourne, Victoria, Australia
| | - Peter A Revill
- Victorian Infectious Diseases Reference Laboratory, Royal Melbourne Hospital, at the Peter Doherty Institute for Infection and Immunity, Melbourne, Victoria, Australia.,Department of Microbiology and Immunology, University of Melbourne, Parkville, Victoria, Australia
| | | | - Josef Wagner
- Victorian Infectious Diseases Reference Laboratory, Royal Melbourne Hospital, at the Peter Doherty Institute for Infection and Immunity, Melbourne, Victoria, Australia
| | - Margaret Littlejohn
- Victorian Infectious Diseases Reference Laboratory, Royal Melbourne Hospital, at the Peter Doherty Institute for Infection and Immunity, Melbourne, Victoria, Australia
| | - Julianne Bayliss
- Victorian Infectious Diseases Reference Laboratory, Royal Melbourne Hospital, at the Peter Doherty Institute for Infection and Immunity, Melbourne, Victoria, Australia
| | - Kathy Jackson
- Victorian Infectious Diseases Reference Laboratory, Royal Melbourne Hospital, at the Peter Doherty Institute for Infection and Immunity, Melbourne, Victoria, Australia
| | | | | | | | | | - Ed Gane
- New Zealand Transplant Unit, Auckland, New Zealand
| | - Henry L Y Chan
- Department of Medicine and Therapeutics, The Chinese University of Hong Kong, Hong Kong, Hong Kong SAR
| | - Xin Li
- Victorian Infectious Diseases Reference Laboratory, Royal Melbourne Hospital, at the Peter Doherty Institute for Infection and Immunity, Melbourne, Victoria, Australia
| | - Scott Bowden
- Victorian Infectious Diseases Reference Laboratory, Royal Melbourne Hospital, at the Peter Doherty Institute for Infection and Immunity, Melbourne, Victoria, Australia
| | - Stephen Locarnini
- Victorian Infectious Diseases Reference Laboratory, Royal Melbourne Hospital, at the Peter Doherty Institute for Infection and Immunity, Melbourne, Victoria, Australia
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11
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Lazarevic I, Banko A, Miljanovic D, Cupic M. Biological features of hepatitis B virus strains associated with fulminant hepatitis. Future Virol 2020. [DOI: 10.2217/fvl-2020-0011] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/21/2022]
Abstract
Accumulating evidence suggests that hepatitis B virus (HBV) biological features may influence the course and clinical manifestations of infection and possibly the development of fulminant hepatitis (FH). Since HBV is not a cytocidal virus, virus-induced liver damage results from an interplay between the virus replication and the host's defense. Therefore, viral factors contributing to enhanced replication, induction of a stronger immune attack or apoptosis of hepatocytes could be crucial in development of FH. Numerous mutations in basal core promoter, pre-C, C and S regions of the HBV genome contribute to development of FH by different mechanisms, including enhanced viral replication, the loss of a decoy for immune response, unbalanced expression of viral proteins and retention of unprocessed cytotoxic proteins in hepatocytes.
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Affiliation(s)
- Ivana Lazarevic
- Institute of Microbiology & Immunology, Faculty of Medicine, University of Belgrade, Dr Subotica 1, 11000 Belgrade, Serbia
| | - Ana Banko
- Institute of Microbiology & Immunology, Faculty of Medicine, University of Belgrade, Dr Subotica 1, 11000 Belgrade, Serbia
| | - Danijela Miljanovic
- Institute of Microbiology & Immunology, Faculty of Medicine, University of Belgrade, Dr Subotica 1, 11000 Belgrade, Serbia
| | - Maja Cupic
- Institute of Microbiology & Immunology, Faculty of Medicine, University of Belgrade, Dr Subotica 1, 11000 Belgrade, Serbia
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12
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Jiang B, Wu Q, Kuhnhenn L, Akhras S, Spengler C, Boller K, Peiffer KH, Hildt E. Formation of semi-enveloped particles as a unique feature of a hepatitis B virus PreS1 deletion mutant. Aliment Pharmacol Ther 2019; 50:940-954. [PMID: 31240738 DOI: 10.1111/apt.15381] [Citation(s) in RCA: 8] [Impact Index Per Article: 1.3] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 03/24/2019] [Revised: 04/29/2019] [Accepted: 05/29/2019] [Indexed: 12/12/2022]
Abstract
BACKGROUND Naturally occurring variants with deletions or mutations in the C-terminal PreS1 domain from hepatitis B virus (HBV) chronically infected patients have been shown to promote HBsAg retention, inhibit HBsAg secretion and change the extracellular appearance of PreS1-containing HBV particles (filaments and virions). AIMS To study the impact of N-terminal deletion in preS1 domain on viral secretion and morphogenesis. METHODS An HBV mutant with 15 amino acids (aa 25-39) deletion in N-terminal preS1 was isolated. Intracellular and extracellular HBsAg were quantified by Western blot. Subcellular HBsAg distribution was analysed by confocal laser scanning microscopy. The viral morphology was characterised by sucrose density gradient ultracentrifugation, Western blot, electron microscopy, HBV mixed ELISA and HBV particle gel essay. RESULTS Expression of this mutant genome released higher amounts of HBsAg in the form of shorter filaments. A significant fraction of semi-enveloped virions was observed in the supernatant that has been unprecedented so far. Stepwise insertion of aa 25-31, aa 32-39 and aa 25-39 increased the length of filaments. The rescue of aa 25-31 and aa 25-39 drastically reduced the amounts of extracellular HBsAg and semi-enveloped virions, while such effects could not be observed after insertion of aa 32-39, arguing against a simple spacer function of this region. The deletion and rescued mutants do not differ in subcellular HBsAg distribution and colocalisation with ER, Golgi and multivesicular bodies markers arguing against differences in release pathways. CONCLUSION N-terminal PreS1-domain (aa 25-31) determines HBsAg secretion and triggers proper assembly of PreS1-containing particles.
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Affiliation(s)
- Bingfu Jiang
- Division of Virology, Paul-Ehrlich-Institut, Langen, Germany
| | - Qingyan Wu
- Division of Virology, Paul-Ehrlich-Institut, Langen, Germany
| | - Lisa Kuhnhenn
- Division of Virology, Paul-Ehrlich-Institut, Langen, Germany.,Department of Gastroenterology and Hepatology, J. W. Goethe University, Frankfurt, Germany
| | - Sami Akhras
- Division of Virology, Paul-Ehrlich-Institut, Langen, Germany
| | | | - Klaus Boller
- Department of Immunology, Paul-Ehrlich-Institut, Langen, Germany
| | - Kai-Henrik Peiffer
- Division of Virology, Paul-Ehrlich-Institut, Langen, Germany.,Department of Gastroenterology and Hepatology, J. W. Goethe University, Frankfurt, Germany
| | - Eberhard Hildt
- Division of Virology, Paul-Ehrlich-Institut, Langen, Germany.,German Center for Infection Research (DZIF), Germany
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13
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Suzuki Y, Maekawa S, Komatsu N, Sato M, Tatsumi A, Miura M, Matsuda S, Muraoka M, Nakakuki N, Amemiya F, Takano S, Fukasawa M, Nakayama Y, Yamaguchi T, Inoue T, Sato T, Sakamoto M, Yamashita A, Moriishi K, Enomoto N. HBV preS deletion mapping using deep sequencing demonstrates a unique association with viral markers. PLoS One 2019; 14:e0212559. [PMID: 30794632 PMCID: PMC6386350 DOI: 10.1371/journal.pone.0212559] [Citation(s) in RCA: 4] [Impact Index Per Article: 0.7] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/30/2018] [Accepted: 02/05/2019] [Indexed: 12/18/2022] Open
Abstract
Aim Deletions are observed frequently in the preS1/S2 region of hepatitis B virus (HBV) genome, in association with liver disease advancement. However, the most significant preS1/S2 region and its influences on viral markers are unclear. Methods The preS1/S2 HBV regions of 90 patients without antiviral therapy were subjected to deep sequencing and deleted regions influencing viral markers were investigated. Results From the deletion frequency analysis in each patient, deletions were observed most frequently in the preS2 codon 132–141 region. When the patients were divided into three groups (0–0.1%: n = 27, 0.1%-10%: n = 34, 10–100%: n = 29), based on the deletion frequency, FIB-4 (p < 0.01), HBV DNA (p < 0.01), HBcrAg (p < 0.01) and preS1/S2 start codon mutations (p < 0.01, both) were significantly associated with the deletion. When clinical and viral markers were investigated by multivariate analysis for their association with the deletion, FIB-4 (p < 0.05), HBcrAg (p < 0.05), and preS1 start codon mutation (p < 0.01) were extracted as independent variables. When the influence of the preS codon 132-141deletions on HBsAg and HBcrAg, relative to HBV DNA, was investigated, the HBsAg/HBV DNA ratio was lower (0–10% vs. 10%-100%, p<0.05), while the HBcrAg/HBV DNA rati o was higher (0–0.1% vs. 10%-100%, p<0.05) in the presence of the preS codon 132-141deletions. Conclusion The preS codon.132-141 deletions have a significant influence on the clinical characteristics and viral markers, even when present as a minor population. Importantly, the preS codon 132–141 deletions have a clear influence on the viral life cycle and pathogenesis.
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Affiliation(s)
- Yuichiro Suzuki
- First Department of Internal Medicine, Faculty of Medicine, University of Yamanashi, Chuo, Yamanashi, Japan
| | - Shinya Maekawa
- First Department of Internal Medicine, Faculty of Medicine, University of Yamanashi, Chuo, Yamanashi, Japan
- * E-mail:
| | - Nobutoshi Komatsu
- First Department of Internal Medicine, Faculty of Medicine, University of Yamanashi, Chuo, Yamanashi, Japan
| | - Mitsuaki Sato
- First Department of Internal Medicine, Faculty of Medicine, University of Yamanashi, Chuo, Yamanashi, Japan
| | - Akihisa Tatsumi
- First Department of Internal Medicine, Faculty of Medicine, University of Yamanashi, Chuo, Yamanashi, Japan
| | - Mika Miura
- First Department of Internal Medicine, Faculty of Medicine, University of Yamanashi, Chuo, Yamanashi, Japan
| | - Shuya Matsuda
- First Department of Internal Medicine, Faculty of Medicine, University of Yamanashi, Chuo, Yamanashi, Japan
| | - Masaru Muraoka
- First Department of Internal Medicine, Faculty of Medicine, University of Yamanashi, Chuo, Yamanashi, Japan
| | - Natsuko Nakakuki
- First Department of Internal Medicine, Faculty of Medicine, University of Yamanashi, Chuo, Yamanashi, Japan
| | - Fumitake Amemiya
- First Department of Internal Medicine, Faculty of Medicine, University of Yamanashi, Chuo, Yamanashi, Japan
| | - Shinichi Takano
- First Department of Internal Medicine, Faculty of Medicine, University of Yamanashi, Chuo, Yamanashi, Japan
| | - Mitsuharu Fukasawa
- First Department of Internal Medicine, Faculty of Medicine, University of Yamanashi, Chuo, Yamanashi, Japan
| | - Yasuhiro Nakayama
- First Department of Internal Medicine, Faculty of Medicine, University of Yamanashi, Chuo, Yamanashi, Japan
| | - Tatsuya Yamaguchi
- First Department of Internal Medicine, Faculty of Medicine, University of Yamanashi, Chuo, Yamanashi, Japan
| | - Taisuke Inoue
- First Department of Internal Medicine, Faculty of Medicine, University of Yamanashi, Chuo, Yamanashi, Japan
| | - Tadashi Sato
- First Department of Internal Medicine, Faculty of Medicine, University of Yamanashi, Chuo, Yamanashi, Japan
| | - Minoru Sakamoto
- First Department of Internal Medicine, Faculty of Medicine, University of Yamanashi, Chuo, Yamanashi, Japan
| | - Atsuya Yamashita
- Department of Microbiology, University of Yamanashi, Chuo, Yamanashi, Japan
| | - Kohji Moriishi
- Department of Microbiology, University of Yamanashi, Chuo, Yamanashi, Japan
| | - Nobuyuki Enomoto
- First Department of Internal Medicine, Faculty of Medicine, University of Yamanashi, Chuo, Yamanashi, Japan
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14
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Nomura M, Tsuge M, Uchida T, Hiraga N, Kurihara M, Tsushima K, Fujino H, Nakahara T, Murakami E, Abe-Chayama H, Kawaoka T, Miki D, Hiramatsu A, Imamura M, Kawakami Y, Aikata H, Ochi H, Zhang Y, Makokha GN, Hayes CN, Tanaka S, Chayama K. CTL-associated and NK cell-associated immune responses induce different HBV DNA reduction patterns in chronic hepatitis B patients. J Viral Hepat 2018; 25:1555-1564. [PMID: 29998562 DOI: 10.1111/jvh.12970] [Citation(s) in RCA: 5] [Impact Index Per Article: 0.7] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 03/30/2018] [Accepted: 06/07/2018] [Indexed: 12/23/2022]
Abstract
The activation of hepatitis B virus (HBV)-related hepatitis is associated with both natural killer (NK) cells and cytotoxic T lymphocytes (CTLs). We analyzed the association between the immune response and changes in the proportion of Pre-S deletion variants. We quantified Pre-S deleted HBV (HBV-del) and wild-type HBV (HBV-wt) DNA levels in sera obtained from HBV-infected mice and chronic hepatitis B patients. In chronic hepatitis B patients, the HBV-del proportion usually increased during or after ALT elevation but did not occur during all ALT elevations. To clarify this difference in the immunological responses, we performed in vivo analyses using HBV-infected human hepatocyte chimeric mice. Although HBV-del proportions did not change in mice with NK cell-associated hepatitis or in mice treated with entecavir, the proportions sharply increased in mice with CTL-associated hepatitis. Furthermore, the number of patients in which HBV-del proportions were greater than 5% was significantly higher in chronic hepatitis B patients than in asymptomatic carriers (P = 0.023). We identified associations between virological response in chronic hepatitis B patients and two different immune responses. The proportion of HBV-del variants could be a useful biomarker for distinguishing between chronic hepatitis and asymptomatic carriers.
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Affiliation(s)
- Motonobu Nomura
- Department of Gastroenterology and Metabolism, Division of Frontier Medical Science, Programs for Biomedical Research Graduate School of Biomedical Science, Hiroshima University, Hiroshima, Japan.,Liver Research Project Center, Hiroshima University, Hiroshima, Japan
| | - Masataka Tsuge
- Department of Gastroenterology and Metabolism, Division of Frontier Medical Science, Programs for Biomedical Research Graduate School of Biomedical Science, Hiroshima University, Hiroshima, Japan.,Liver Research Project Center, Hiroshima University, Hiroshima, Japan.,Natural Science Center for Basic Research and Development, Hiroshima University, Hiroshima, Japan
| | - Takuro Uchida
- Department of Gastroenterology and Metabolism, Division of Frontier Medical Science, Programs for Biomedical Research Graduate School of Biomedical Science, Hiroshima University, Hiroshima, Japan.,Liver Research Project Center, Hiroshima University, Hiroshima, Japan
| | - Nobuhiko Hiraga
- Department of Gastroenterology and Metabolism, Division of Frontier Medical Science, Programs for Biomedical Research Graduate School of Biomedical Science, Hiroshima University, Hiroshima, Japan.,Liver Research Project Center, Hiroshima University, Hiroshima, Japan
| | - Mio Kurihara
- Department of Gastroenterology and Metabolism, Division of Frontier Medical Science, Programs for Biomedical Research Graduate School of Biomedical Science, Hiroshima University, Hiroshima, Japan.,Liver Research Project Center, Hiroshima University, Hiroshima, Japan
| | - Ken Tsushima
- Department of Gastroenterology and Metabolism, Division of Frontier Medical Science, Programs for Biomedical Research Graduate School of Biomedical Science, Hiroshima University, Hiroshima, Japan.,Liver Research Project Center, Hiroshima University, Hiroshima, Japan
| | - Hatsue Fujino
- Department of Gastroenterology and Metabolism, Division of Frontier Medical Science, Programs for Biomedical Research Graduate School of Biomedical Science, Hiroshima University, Hiroshima, Japan.,Liver Research Project Center, Hiroshima University, Hiroshima, Japan
| | - Takashi Nakahara
- Department of Gastroenterology and Metabolism, Division of Frontier Medical Science, Programs for Biomedical Research Graduate School of Biomedical Science, Hiroshima University, Hiroshima, Japan.,Liver Research Project Center, Hiroshima University, Hiroshima, Japan
| | - Eisuke Murakami
- Department of Gastroenterology and Metabolism, Division of Frontier Medical Science, Programs for Biomedical Research Graduate School of Biomedical Science, Hiroshima University, Hiroshima, Japan.,Liver Research Project Center, Hiroshima University, Hiroshima, Japan
| | - Hiromi Abe-Chayama
- Liver Research Project Center, Hiroshima University, Hiroshima, Japan.,Center for Medical Specialist Graduate Education and Research, Institute of Biomedical and Health Sciences, Hiroshima University, Hiroshima, Japan
| | - Tomokazu Kawaoka
- Department of Gastroenterology and Metabolism, Division of Frontier Medical Science, Programs for Biomedical Research Graduate School of Biomedical Science, Hiroshima University, Hiroshima, Japan.,Liver Research Project Center, Hiroshima University, Hiroshima, Japan
| | - Daiki Miki
- Department of Gastroenterology and Metabolism, Division of Frontier Medical Science, Programs for Biomedical Research Graduate School of Biomedical Science, Hiroshima University, Hiroshima, Japan.,Liver Research Project Center, Hiroshima University, Hiroshima, Japan.,Laboratory for Digestive Diseases, RIKEN Center for Integrative Medical Sciences, Hiroshima, Japan
| | - Akira Hiramatsu
- Department of Gastroenterology and Metabolism, Division of Frontier Medical Science, Programs for Biomedical Research Graduate School of Biomedical Science, Hiroshima University, Hiroshima, Japan.,Liver Research Project Center, Hiroshima University, Hiroshima, Japan
| | - Michio Imamura
- Department of Gastroenterology and Metabolism, Division of Frontier Medical Science, Programs for Biomedical Research Graduate School of Biomedical Science, Hiroshima University, Hiroshima, Japan.,Liver Research Project Center, Hiroshima University, Hiroshima, Japan
| | - Yoshiiku Kawakami
- Department of Gastroenterology and Metabolism, Division of Frontier Medical Science, Programs for Biomedical Research Graduate School of Biomedical Science, Hiroshima University, Hiroshima, Japan.,Liver Research Project Center, Hiroshima University, Hiroshima, Japan
| | - Hiroshi Aikata
- Department of Gastroenterology and Metabolism, Division of Frontier Medical Science, Programs for Biomedical Research Graduate School of Biomedical Science, Hiroshima University, Hiroshima, Japan.,Liver Research Project Center, Hiroshima University, Hiroshima, Japan
| | - Hidenori Ochi
- Department of Gastroenterology and Metabolism, Division of Frontier Medical Science, Programs for Biomedical Research Graduate School of Biomedical Science, Hiroshima University, Hiroshima, Japan.,Liver Research Project Center, Hiroshima University, Hiroshima, Japan.,Laboratory for Digestive Diseases, RIKEN Center for Integrative Medical Sciences, Hiroshima, Japan
| | - Yizhou Zhang
- Department of Gastroenterology and Metabolism, Division of Frontier Medical Science, Programs for Biomedical Research Graduate School of Biomedical Science, Hiroshima University, Hiroshima, Japan.,Liver Research Project Center, Hiroshima University, Hiroshima, Japan
| | - Grace Naswa Makokha
- Department of Gastroenterology and Metabolism, Division of Frontier Medical Science, Programs for Biomedical Research Graduate School of Biomedical Science, Hiroshima University, Hiroshima, Japan.,Liver Research Project Center, Hiroshima University, Hiroshima, Japan
| | - Clair Nelson Hayes
- Department of Gastroenterology and Metabolism, Division of Frontier Medical Science, Programs for Biomedical Research Graduate School of Biomedical Science, Hiroshima University, Hiroshima, Japan.,Liver Research Project Center, Hiroshima University, Hiroshima, Japan
| | - Shinji Tanaka
- Department of Gastroenterology and Metabolism, Division of Frontier Medical Science, Programs for Biomedical Research Graduate School of Biomedical Science, Hiroshima University, Hiroshima, Japan.,Department of Endoscopy, Hiroshima University Hospital, Hiroshima, Japan
| | - Kazuaki Chayama
- Department of Gastroenterology and Metabolism, Division of Frontier Medical Science, Programs for Biomedical Research Graduate School of Biomedical Science, Hiroshima University, Hiroshima, Japan.,Liver Research Project Center, Hiroshima University, Hiroshima, Japan.,Laboratory for Digestive Diseases, RIKEN Center for Integrative Medical Sciences, Hiroshima, Japan
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15
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Genetic variability in coding regions of the surface antigen and reverse transcriptase domain of hepatitis B virus polymerase, Colombia, 2002-2014. BIOMEDICA 2018; 38:37-50. [PMID: 30184362 DOI: 10.7705/biomedica.v38i3.3871] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Subscribe] [Scholar Register] [Received: 04/07/2017] [Revised: 11/10/2017] [Indexed: 11/21/2022]
Abstract
Introduction: Despite the availability of an effective vaccine and treatment to reduce the viral load and progressive hepatocellular injury, approximately 240 million people worldwide are chronically infected with the hepatitis B virus (HBV). In Colombia, the circulation of different viral genotypes has been confirmed. Mutations in the genome have been associated to antiviral therapy resistance, viral escape to neutralizing antibodies, occult infection and progression to hepatocellular carcinoma.
Objective: To identify the genotypes and the presence of mutations in the coding region of the surface (S) antigen and the reverse transcriptase (RT) domain of the polymerase of HBV obtained from serum samples for hepatitis B diagnosis received by the Instituto Nacional de Salud during the period 2002-2014.
Materials and methods: A total of 495 serum samples with previous HBsAg reactive result were used for molecular detection. A fragment of 1,591 nucleotides was sequenced, and the corresponding phylogenetic analysis was performed.
Results: We detected the viral genome of HBV in 66 samples and 28 were successfully sequenced. The phylogenetic analysis allowed the identification of subgenotypes F3 and A2. The L180M and M204V resistance mutations were simultaneously identified in one sample, while the I169L resistance mutation was identified in another one. A single escape mutation, P120Q, was identified in one more. Two samples showed a deletion of 105 nucleotides in the preS1-preS2 region.
Conclusions: The circulation of genotypes/subgenotypes F3 and A2 of HBV in Colombia was corroborated, as well as the presence of some resistance and escape mutations. The present study constitutes a contribution to the molecular epidemiology of HBV in Colombia.
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16
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Rajoriya N, Combet C, Zoulim F, Janssen HLA. How viral genetic variants and genotypes influence disease and treatment outcome of chronic hepatitis B. Time for an individualised approach? J Hepatol 2017; 67:1281-1297. [PMID: 28736138 DOI: 10.1016/j.jhep.2017.07.011] [Citation(s) in RCA: 106] [Impact Index Per Article: 13.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 01/03/2017] [Revised: 06/27/2017] [Accepted: 07/12/2017] [Indexed: 12/12/2022]
Abstract
Chronic hepatitis B virus (HBV) infection remains a global problem. Several HBV genotypes exist with different biology and geographical prevalence. Whilst the future aim of HBV treatment remains viral eradication, current treatment strategies aim to suppress the virus and prevent the progression of liver disease. Current strategies also involve identification of patients for treatment, namely those at risk of progressive liver disease. Identification of HBV genotype, HBV mutants and other predictive factors allow for tailoured treatments, and risk-surveillance pathways, such as hepatocellular cancer screening. In the future, these factors may enable stratification not only of treatment decisions, but also of patients at risk of higher relapse rates when current therapies are discontinued. Newer technologies, such as next-generation sequencing, to assess drug-resistant or immune escape variants and quasi-species heterogeneity in patients, may allow for more information-based treatment decisions between the clinician and the patient. This article serves to discuss how HBV genotypes and genetic variants impact not only upon the disease course and outcomes, but also current treatment strategies. Adopting a personalised genotypic approach may play a role in future strategies to combat the disease. Herein, we discuss new technologies that may allow more informed decision-making for response guided therapy in the battle against HBV.
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Affiliation(s)
- Neil Rajoriya
- Toronto Centre for Liver Diseases, Toronto General Hospital, 200 Elizabeth Street, Toronto, Ontario M5G 2C4, Canada
| | - Christophe Combet
- Univ Lyon, Université Claude Bernard Lyon 1, INSERM 1052, CNRS 5286, Centre Léon Bérard, Centre de recherche en cancérologie de Lyon, Lyon 69XXX, France
| | - Fabien Zoulim
- Univ Lyon, Université Claude Bernard Lyon 1, INSERM 1052, CNRS 5286, Centre Léon Bérard, Centre de recherche en cancérologie de Lyon, Lyon 69XXX, France; Department of Hepatology, Groupement Hospitalier Nord, Hospices Civils de Lyon, Lyon, France
| | - Harry L A Janssen
- Toronto Centre for Liver Diseases, Toronto General Hospital, 200 Elizabeth Street, Toronto, Ontario M5G 2C4, Canada.
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17
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Bayliss J, Yuen L, Rosenberg G, Wong D, Littlejohn M, Jackson K, Gaggar A, Kitrinos KM, Subramanian GM, Marcellin P, Buti M, Janssen HLA, Gane E, Sozzi V, Colledge D, Hammond R, Edwards R, Locarnini S, Thompson A, Revill PA. Deep sequencing shows that HBV basal core promoter and precore variants reduce the likelihood of HBsAg loss following tenofovir disoproxil fumarate therapy in HBeAg-positive chronic hepatitis B. Gut 2017; 66:2013-2023. [PMID: 27534671 DOI: 10.1136/gutjnl-2015-309300] [Citation(s) in RCA: 38] [Impact Index Per Article: 4.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 02/01/2015] [Revised: 07/20/2016] [Accepted: 07/21/2016] [Indexed: 12/21/2022]
Abstract
OBJECTIVE Hepatitis B e antigen (HBeAg) seroconversion and hepatitis B surface antigen (HBsAg) loss are important clinical outcomes for patients with chronic hepatitis B (CHB) treated with antiviral therapy. To date, there have been few studies that have evaluated viral sequence markers predicting serological response to nucleos(t)ide analogue (NA) treatment. DESIGN We used next-generation sequencing (NGS) and quantitative HBV serology (HBeAg and HBsAg) to identify viral sequence markers associated with serological response to long-term tenofovir disoproxil fumarate therapy among HBeAg-positive patients. In the GS-US-174-0103 study, approximately half the patients seroconverted to anti-HBe by week 192 and 11% of patients exhibited HBsAg loss, the closest outcome to functional cure. The frequency of HBV variants that have previously been associated with HBV clinical outcomes was evaluated. HBV viral diversity in baseline sequences generated by NGS was calculated using Shannon entropy. RESULTS NGS analysis of HBV sequences from 157 patients infected with genotypes A to D showed the frequency of variants in the basal core promoter (BCP) and precore (PC) regions varied by genotype and that these mutations were associated with the absence of HBsAg loss. This was the case even when mutations were present at frequencies below the threshold of detection by population sequencing. Increased viral diversity across the HBV genome as determined by NGS was also associated with reduced likelihood of HBsAg loss. CONCLUSION Patients with detectable BCP and/or PC variants and higher viral diversity have a lower probability of HBsAg loss during long-term NA therapy. Strategies to achieve functional cure of HBV infection through combination therapy should consider using NGS to stratify patients according to BCP/PC sequence. Consideration should also be given to earlier initiation of therapy prior to the emergence of BCP/PC variants. TRIAL REGISTRATION NUMBER NCT00116805; Post result.
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Affiliation(s)
- Julianne Bayliss
- Division of Molecular Research and Development, Victorian Infectious Diseases Reference Laboratory, The Royal Melbourne Hospital, Peter Doherty Institute for Infection and Immunity, Melbourne, Victoria, Australia
| | - Lilly Yuen
- Division of Molecular Research and Development, Victorian Infectious Diseases Reference Laboratory, The Royal Melbourne Hospital, Peter Doherty Institute for Infection and Immunity, Melbourne, Victoria, Australia
| | - Gillian Rosenberg
- Division of Molecular Research and Development, Victorian Infectious Diseases Reference Laboratory, The Royal Melbourne Hospital, Peter Doherty Institute for Infection and Immunity, Melbourne, Victoria, Australia
| | - Darren Wong
- Division of Molecular Research and Development, Victorian Infectious Diseases Reference Laboratory, The Royal Melbourne Hospital, Peter Doherty Institute for Infection and Immunity, Melbourne, Victoria, Australia.,Department of Gastroenterology, St. Vincent's Hospital, Melbourne, Victoria, Australia
| | - Margaret Littlejohn
- Division of Molecular Research and Development, Victorian Infectious Diseases Reference Laboratory, The Royal Melbourne Hospital, Peter Doherty Institute for Infection and Immunity, Melbourne, Victoria, Australia
| | - Kathleen Jackson
- Division of Molecular Research and Development, Victorian Infectious Diseases Reference Laboratory, The Royal Melbourne Hospital, Peter Doherty Institute for Infection and Immunity, Melbourne, Victoria, Australia
| | - Anuj Gaggar
- Gilead Sciences, Foster City, California, USA
| | | | | | | | - Maria Buti
- Liver Unit, Valle d'Hebron (Ciberehd) University Hospital, Barcelona, Spain
| | - Harry L A Janssen
- Toronto Center for Liver Diseases, Toronto Western and General Hospital, University Health Network, University of Toronto, Toronto, Ontario, Canada
| | - Ed Gane
- New Zealand Liver Transplant Unit, Auckland City Hospital, Auckland, New Zealand
| | - Vitina Sozzi
- Division of Molecular Research and Development, Victorian Infectious Diseases Reference Laboratory, The Royal Melbourne Hospital, Peter Doherty Institute for Infection and Immunity, Melbourne, Victoria, Australia
| | - Danni Colledge
- Division of Molecular Research and Development, Victorian Infectious Diseases Reference Laboratory, The Royal Melbourne Hospital, Peter Doherty Institute for Infection and Immunity, Melbourne, Victoria, Australia
| | - Rachel Hammond
- Division of Molecular Research and Development, Victorian Infectious Diseases Reference Laboratory, The Royal Melbourne Hospital, Peter Doherty Institute for Infection and Immunity, Melbourne, Victoria, Australia
| | - Rosalind Edwards
- Division of Molecular Research and Development, Victorian Infectious Diseases Reference Laboratory, The Royal Melbourne Hospital, Peter Doherty Institute for Infection and Immunity, Melbourne, Victoria, Australia
| | - Stephen Locarnini
- Division of Molecular Research and Development, Victorian Infectious Diseases Reference Laboratory, The Royal Melbourne Hospital, Peter Doherty Institute for Infection and Immunity, Melbourne, Victoria, Australia
| | - Alexander Thompson
- Department of Gastroenterology, St. Vincent's Hospital, Melbourne, Victoria, Australia
| | - Peter A Revill
- Division of Molecular Research and Development, Victorian Infectious Diseases Reference Laboratory, The Royal Melbourne Hospital, Peter Doherty Institute for Infection and Immunity, Melbourne, Victoria, Australia
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18
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Jia J, Liang X, Chen S, Wang H, Li H, Fang M, Bai X, Wang Z, Wang M, Zhu S, Sun F, Gao C. Next-generation sequencing revealed divergence in deletions of the preS region in the HBV genome between different HBV-related liver diseases. J Gen Virol 2017; 98:2748-2758. [PMID: 29022863 DOI: 10.1099/jgv.0.000942] [Citation(s) in RCA: 12] [Impact Index Per Article: 1.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/13/2022] Open
Abstract
In order to investigate if deletion patterns of the preS region can predict liver disease advancement, the preS region of the hepatitis B virus (HBV) genome in 45 chronic hepatitis B (CHB) and 94 HBV-related hepatocellular carcinoma (HCC) patients was sequenced by next-generation sequencing (NGS) and the percentages of nucleotide deletion in the preS region were analysed. Hierarchical clustering and heatmaps based on deletion percentages of preS revealed different deletion patterns between CHB and HCC patients. Intergenotype comparison also indicated divergence in preS deletions between HBV genotype B and C. No significant difference was found in preS deletion patterns between sera and matched adjacent non-tumour tissues. Based on hierarchical clustering, HCC patients were classed into two groups with different preS deletion patterns and different clinical features. Finally, the support vector machine (SVM) model was trained on preS nucleotide deletion percentages and used to predict HCC versus CHB patients. The prediction performance was assessed with fivefold cross-validation and independent cohort validation. The median area under the curve (AUC) was 0.729 after repeating SVM 500 times with fivefold cross-validations. After parameter optimization, the SVM model was used to predict an independent cohort with 51 CHB patients and 72 HCC patients and the AUC was 0.727. In conclusion, the use of the NGS method revealed a prominent divergence in preS deletion patterns between disease groups and virus genotypes, but not between different tissue types. Quantitative NGS data combined with a machine learning method could be a powerful approach for prediction of the status of different diseases.
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Affiliation(s)
- Jian'an Jia
- Department of Laboratory Medicine, Eastern Hepatobiliary Surgery Hospital, Second Military Medical University, Shanghai, 200438, PR China.,Department of Laboratory Medicine, The 105th Hospital of PLA, Hefei 230031, PR China
| | - Xiaotao Liang
- Shanghai Key Lab of Intelligent Information Processing and School of Computer Science, Fudan University, Shanghai, PR China.,Centre for Computational Systems Biology, School of Mathematical Sciences, Fudan University, Shanghai, PR China
| | - Shipeng Chen
- Department of Laboratory Medicine, Eastern Hepatobiliary Surgery Hospital, Second Military Medical University, Shanghai, 200438, PR China
| | - Hui Wang
- Department of Laboratory Medicine, Eastern Hepatobiliary Surgery Hospital, Second Military Medical University, Shanghai, 200438, PR China.,Department of Clinical Laboratory, The First Affiliated Hospital of Chinese PLA's General Hospital, Beijing 100048, PR China
| | - Huiming Li
- Department of Laboratory Medicine, Eastern Hepatobiliary Surgery Hospital, Second Military Medical University, Shanghai, 200438, PR China
| | - Meng Fang
- Department of Laboratory Medicine, Eastern Hepatobiliary Surgery Hospital, Second Military Medical University, Shanghai, 200438, PR China
| | - Xin Bai
- Centre for Computational Systems Biology, School of Mathematical Sciences, Fudan University, Shanghai, PR China
| | - Ziyi Wang
- Shanghai Institute of Technology, Shanghai 201418, PR China
| | - Mengmeng Wang
- Department of Laboratory Medicine, Eastern Hepatobiliary Surgery Hospital, Second Military Medical University, Shanghai, 200438, PR China
| | - Shanfeng Zhu
- Shanghai Key Lab of Intelligent Information Processing and School of Computer Science, Fudan University, Shanghai, PR China.,Centre for Computational Systems Biology, School of Mathematical Sciences, Fudan University, Shanghai, PR China
| | - Fengzhu Sun
- Centre for Computational Systems Biology, School of Mathematical Sciences, Fudan University, Shanghai, PR China.,Molecular and Computational Program Department of Biological Sciences, University of Southern California, LA 90089, USA
| | - Chunfang Gao
- Department of Laboratory Medicine, Eastern Hepatobiliary Surgery Hospital, Second Military Medical University, Shanghai, 200438, PR China
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19
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Zhu HL, Li X, Li J, Zhang ZH. Genetic variation of occult hepatitis B virus infection. World J Gastroenterol 2016; 22:3531-3546. [PMID: 27053845 PMCID: PMC4814639 DOI: 10.3748/wjg.v22.i13.3531] [Citation(s) in RCA: 59] [Impact Index Per Article: 6.6] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 10/29/2015] [Revised: 12/13/2015] [Accepted: 12/30/2015] [Indexed: 02/06/2023] Open
Abstract
Occult hepatitis B virus infection (OBI), characterized as the persistence of hepatitis B virus (HBV) surface antigen (HBsAg) seronegativity and low viral load in blood or liver, is a special form of HBV infection. OBI may be related mainly to mutations in the HBV genome, although the underlying mechanism of it remains to be clarified. Mutations especially within the immunodominant “α” determinant of S protein are “hot spots” that could contribute to the occurrence of OBI via affecting antigenicity and immunogenicity of HBsAg or replication and secretion of virion. Clinical reports account for a large proportion of previous studies on OBI, while functional analyses, especially those based on full-length HBV genome, are rare.
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20
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Gededzha MP, Muzeze M, Burnett RJ, Amponsah-Dacosta E, Mphahlele MJ, Selabe SG. Complete genome analysis of hepatitis B virus in human immunodeficiency virus infected and uninfected South Africans. J Med Virol 2016; 88:1560-6. [PMID: 26890489 DOI: 10.1002/jmv.24502] [Citation(s) in RCA: 6] [Impact Index Per Article: 0.7] [Reference Citation Analysis] [Abstract] [Key Words] [Journal Information] [Subscribe] [Scholar Register] [Accepted: 02/14/2016] [Indexed: 12/21/2022]
Abstract
Hepatitis B virus (HBV) and human immunodeficiency virus (HIV) infections are highly endemic in South Africa. Data on the complete genome sequences of HBV in HIV-positive patients in South Africa are scanty. This study characterized the complete HBV genome isolated from both HIV-positive and negative patients at the Dr George Mukhari Academic Hospital (DGMAH), Pretoria. Serum samples from nine (five HIV-positive and four HIV-negative) patients attending the DGMAH from 2007 to 2011 were serologically tested, amplified, and sequenced for complete genome. Phylogenetic tree was constructed using MEGA6.0. Mutations were analyzed by comparing the sequences with genotype-matched GenBank references. Eight patients were HBsAg positive, with only one from the HIV positive group being negative. Phylogenetic analysis of the complete genome sequences classified them into five genotypes; A1 (n = 4), A2 (n = 1), C1 (n = 2), D1 (n = 1), and D3 (n = 1). Deletions up to 35 nucleotides in length were identified in this study. No drug resistance mutations were identified in the P ORF, while the L217R mutation was identified in one subgenotype A2 sequence. The double (A1762T/G1764A) and triple (T1753C/A1762T/G1764A) mutations in the Basal core promoter were identified in four and two sequences, respectively. In the core region, mutation G1888A was identified in four of the subgenotype A1 sequences. In conclusion, this study has added to the limited South African data on HBV genotypes and mutations in HBV/HIV co-infected and HBV mono-infected patients, based on complete HBV genome analysis. Subgenotype A1 was predominant, and no drug-resistant mutants were detected in the study. J. Med. Virol. 88:1560-1566, 2016. © 2016 Wiley Periodicals, Inc.
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Affiliation(s)
- Maemu P Gededzha
- Department of Virology, HIV and Hepatitis Research Unit, Sefako Makgatho Health Sciences University and National Health Laboratory Service, Pretoria, South Africa
| | - Muxe Muzeze
- Department of Virology, HIV and Hepatitis Research Unit, Sefako Makgatho Health Sciences University and National Health Laboratory Service, Pretoria, South Africa
| | - Rosemary J Burnett
- Department of Virology, HIV and Hepatitis Research Unit, Sefako Makgatho Health Sciences University and National Health Laboratory Service, Pretoria, South Africa
| | - Edina Amponsah-Dacosta
- Department of Virology, HIV and Hepatitis Research Unit, Sefako Makgatho Health Sciences University and National Health Laboratory Service, Pretoria, South Africa
| | | | - Selokela G Selabe
- Department of Virology, HIV and Hepatitis Research Unit, Sefako Makgatho Health Sciences University and National Health Laboratory Service, Pretoria, South Africa
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21
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Li YW, Yang FC, Lu HQ, Zhang JS. Hepatocellular carcinoma and hepatitis B surface protein. World J Gastroenterol 2016; 22:1943-1952. [PMID: 26877602 PMCID: PMC4726670 DOI: 10.3748/wjg.v22.i6.1943] [Citation(s) in RCA: 44] [Impact Index Per Article: 4.9] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 04/11/2015] [Revised: 07/27/2015] [Accepted: 12/01/2015] [Indexed: 02/06/2023] Open
Abstract
The tumorigenesis of hepatitis B virus (HBV)-associated hepatocellular carcinoma (HCC) has been widely studied. HBV envelope proteins are important for the structure and life cycle of HBV, and these proteins are useful for judging the natural disease course and guiding treatment. Truncated and mutated preS/S are produced by integrated viral sequences that are defective for replication. The preS/S mutants are considered “precursor lesions” of HCC. Different preS/S mutants induce various mechanisms of tumorigenesis, such as transactivation of transcription factors and an immune inflammatory response, thereby contributing to HCC. The preS2 mutants and type II “Ground Glass” hepatocytes represent novel biomarkers of HBV-associated HCC. The preS mutants may induce the unfolded protein response and endoplasmic reticulum stress-dependent and stress-independent pathways. Treatments to inhibit hepatitis B surface antigen (HBsAg) and damage secondary to HBsAg or the preS/S mutants include antivirals and antioxidants, such as silymarin, resveratrol, and glycyrrhizin acid. Methods for the prevention and treatment of HCC should be comprehensive.
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22
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Akpinar F, Timm A, Yin J. High-Throughput Single-Cell Kinetics of Virus Infections in the Presence of Defective Interfering Particles. J Virol 2016; 90:1599-612. [PMID: 26608322 PMCID: PMC4719634 DOI: 10.1128/jvi.02190-15] [Citation(s) in RCA: 35] [Impact Index Per Article: 3.9] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/28/2015] [Accepted: 11/18/2015] [Indexed: 01/01/2023] Open
Abstract
UNLABELLED Defective interfering particles (DIPs) are virus mutants that lack essential genes for growth. In coinfections with helper virus, the diversion of viral proteins to the replication and packaging of DIP genomes can interfere with virus production. Mounting cases of DIPs and DIP-like genomes in clinical and natural isolates, as well as growing interest in DIP-based therapies, underscore a need to better elucidate how DIPs work. DIP activity is primarily measured by its inhibition of virus infection yield, an endpoint that masks the dynamic and potentially diverse individual cell behaviors. Using vesicular stomatitis virus (VSV) as a model, we coinfected BHK cells with VSV DIPs and recombinant helper virus carrying a gene encoding a red fluorescent protein (RFP) whose expression correlates with the timing and level of virus release. For single cells within a monolayer, 10 DIPs per cell suppressed the reporter expression in only 1.2% of the cells. In most cells, it slowed and reduced viral gene expression, manifested as a shift in mean latent time from 4 to 6 h and reduced virus yields by 10-fold. For single cells isolated in microwells, DIP effects were more pronounced, reducing virus yields by 100-fold and extending latent times to 12 h, including individual instances above 20 h. Together, these results suggest that direct or indirect cell-cell interactions prevent most coinfected cells from being completely suppressed by DIPs. Finally, a gamma distribution model captures well how the infection kinetics quantitatively depends on the DIP dose. Such models will be useful for advancing a predictive biology of DIP-associated virus growth and infection spread. IMPORTANCE During the last century, basic studies in virology have focused on developing a molecular mechanistic understanding of how infectious viruses reproduce in their living host cells. However, over the last 10 years, the advent of deep sequencing and other powerful technologies has revealed in natural and patient infections that viruses do not act alone. Instead, viruses are often accompanied by defective virus-like particles that carry large deletions in their genomes and fail to replicate on their own. Coinfections of viable and defective viruses behave in unpredictable ways, but they often interfere with normal virus growth, potentially enabling infections to evade host immune surveillance. In the current study, controlled levels of defective viruses are coinfected with viable viruses that have been engineered to express a fluorescent reporter protein during infection. Unique profiles of reporter expression acquired from thousands of coinfected cells reveal how interference acts at multiple stages of infection.
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Affiliation(s)
- Fulya Akpinar
- Systems Biology Theme, Wisconsin Institute for Discovery, Department of Chemical and Biological Engineering, University of Wisconsin-Madison, Madison, Wisconsin, USA
| | - Andrea Timm
- Systems Biology Theme, Wisconsin Institute for Discovery, Department of Chemical and Biological Engineering, University of Wisconsin-Madison, Madison, Wisconsin, USA
| | - John Yin
- Systems Biology Theme, Wisconsin Institute for Discovery, Department of Chemical and Biological Engineering, University of Wisconsin-Madison, Madison, Wisconsin, USA
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23
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Guerrieri F, Belloni L, Pediconi N, Levrero M. Pathobiology of Hepatitis B Virus-Induced Carcinogenesis. ACTA ACUST UNITED AC 2016. [DOI: 10.1007/978-3-319-22330-8_5] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.1] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/14/2022]
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24
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Coppola N, Onorato L, Minichini C, Di Caprio G, Starace M, Sagnelli C, Sagnelli E. Clinical significance of hepatitis B surface antigen mutants. World J Hepatol 2015; 7:2729-2739. [PMID: 26644816 PMCID: PMC4663392 DOI: 10.4254/wjh.v7.i27.2729] [Citation(s) in RCA: 63] [Impact Index Per Article: 6.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 06/29/2015] [Revised: 09/27/2015] [Accepted: 11/17/2015] [Indexed: 02/06/2023] Open
Abstract
Hepatitis B virus (HBV) infection is a major public health problem in many countries, with nearly 300 million people worldwide carrying HBV chronic infection and over 1 million deaths per year due to cirrhosis and liver cancer. Several hepatitis B surface antigen (HBsAg) mutations have been described, most frequently due to a single amino acid substitution and seldom to a nucleotide deletion. The majority of mutations are located in the S region, but they have also been found in the pre-S1 and pre-S2 regions. Single amino acid substitutions in the major hydrophilic region of HBsAg, called the “a” determinant, have been associated with immune escape and the consequent failure of HBV vaccination and HBsAg detection, whereas deletions in the pre-S1 or pre-S2 regions have been associated with the development of hepatocellular carcinoma. This review article will focus on the HBsAg mutants and their biological and clinical implications.
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25
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Chiu AP, Tschida BR, Lo LH, Moriarity BS, Rowlands DK, Largaespada DA, Keng VW. Transposon mouse models to elucidate the genetic mechanisms of hepatitis B viral induced hepatocellular carcinoma. World J Gastroenterol 2015; 21:12157-12170. [PMID: 26576100 PMCID: PMC4641133 DOI: 10.3748/wjg.v21.i42.12157] [Citation(s) in RCA: 5] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 08/12/2015] [Revised: 09/18/2015] [Accepted: 09/30/2015] [Indexed: 02/06/2023] Open
Abstract
The major type of human liver cancer is hepatocellular carcinoma (HCC), and there are currently many risk factors that contribute to this deadly disease. The majority of HCC occurrences are associated with chronic hepatitis viral infection, and hepatitis B viral (HBV) infection is currently a major health problem in Eastern Asia. Elucidating the genetic mechanisms associated with HBV-induced HCC has been difficult due to the heterogeneity and genetic complexity associated with this disease. A repertoire of animal models has been broadly used to study the pathophysiology and to develop potential treatment regimens for HBV-associated HCC. The use of these animal models has provided valuable genetic information and has been an important contributor to uncovering the factors involved in liver malignant transformation, invasion and metastasis. Recently, transposon-based mouse models are becoming more widely used in liver cancer research to interrogate the genome by forward genetics and also used to validate genes rapidly in a reverse genetic manner. Importantly, these transposon-based rapid reverse genetic mouse models could become crucial in testing potential therapeutic agents before proceeding to clinical trials in human. Therefore, this review will cover the use of transposon-based mouse models to address the problems of liver cancer, especially HBV-associated HCC occurrences in Asia.
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Characterization of Full-Length Genomes of Hepatitis B Virus Quasispecies in Sera of Patients at Different Phases of Infection. J Clin Microbiol 2015; 53:2203-14. [PMID: 25926495 DOI: 10.1128/jcm.00068-15] [Citation(s) in RCA: 25] [Impact Index Per Article: 2.5] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/13/2015] [Accepted: 04/24/2015] [Indexed: 12/17/2022] Open
Abstract
Hepatitis B virus (HBV) infection results in different clinical presentation due to different levels of immune response. Our study aimed to characterize HBV full-length genome quasispecies (QS) in patients with different phases of infection to better understand its pathogenesis. Forty treatment-naive HBV-infected patients were enrolled, including 10 cases of acute hepatitis B (AHB), 9 cases of immunotolerant (IT) HBV carriers, 11 cases of chronic hepatitis B (CHB), and 10 cases of acute-on-chronic liver failure (ACLF). The present study was conducted by clone-based sequencing. QS heterogeneity within each open reading frame was calculated. The mutation frequency index (MFI) and amino acid variations within the large HBsAg, HBcAg, and HBxAg regions were analyzed based on the different infection phases. In total, 606 HBV full-length sequences were obtained. HBV QS had higher heterogeneity in ACLF and CHB than that in IT among chronically infected individuals. AHB patients had the lower QS heterogeneity at onset than those with chronic infection. ACLF patients had the highest frequency of mutations in the core promoter and precore region. A triple mutation (A1762T/G1764A/G1896A) was observed more frequently in genotype C than in genotype B. The MFI indicated that specific peptides of the studied regions had more frequent mutations in ACLF. Furthermore, several amino acid variations, known as T- and B-cell epitopes, were potentially associated with the immunoactive phase of infection. More HBV genome mutations and deletions were observed in patients with more severe diseases, particularly in specific regions of the core and preS regions, the clinical significance and mechanism of which need to be further investigated.
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Li F, Zhang D, Li Y, Jiang D, Luo S, Du N, Chen W, Deng L, Zeng C. Whole genome characterization of hepatitis B virus quasispecies with massively parallel pyrosequencing. Clin Microbiol Infect 2015; 21:280-7. [PMID: 25658544 DOI: 10.1016/j.cmi.2014.10.007] [Citation(s) in RCA: 19] [Impact Index Per Article: 1.9] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/28/2014] [Revised: 10/04/2014] [Accepted: 10/10/2014] [Indexed: 01/19/2023]
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The emerging role of hepatitis B virus pre-S2 deletion mutant proteins in HBV tumorigenesis. J Biomed Sci 2014; 21:98. [PMID: 25316153 PMCID: PMC4200140 DOI: 10.1186/s12929-014-0098-7] [Citation(s) in RCA: 55] [Impact Index Per Article: 5.0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/28/2014] [Accepted: 10/07/2014] [Indexed: 02/07/2023] Open
Abstract
Chronic hepatitis B virus (HBV) infection can cause hepatocellular carcinoma (HCC). Several hypotheses have been proposed to explain the mechanisms of HBV tumorigenesis, including inflammation and liver regeneration associated with cytotoxic immune injuries and transcriptional activators of mutant HBV gene products. The mutant viral oncoprotein-driven tumorigenesis is prevailed at the advanced stage or anti-HBe-positive phase of chronic HBV infection. Besides HBx, the pre-S2 (deletion) mutant protein represents a newly recognized oncoprotein that is accumulated in the endoplasmic reticulum (ER) and manifests as type II ground glass hepatocytes (GGH). The retention of pre-S2 mutant protein in ER can induce ER stress and initiate an ER stress-dependent VEGF/Akt/mTOR and NFκB/COX-2 signal pathway. Additionally, the pre-S2 mutant large surface protein can induce an ER stress-independent pathway to transactivate JAB-1/p27/RB/cyclin A,D pathway, leading to growth advantage of type II GGH. The pre-S2 mutant protein-induced ER stress can also cause DNA damage, centrosome overduplication, and genomic instability. In 5-10% of type II GGHs, there is co-expression of pre-S2 mutant protein and HBx antigen which exhibited enhanced oncogenic effects in transgenic mice. The mTOR signal cascade is consistently activated throughout the course of pre-S2 mutant transgenic livers and in human HCC tissues, leading to metabolic disorders and HCC tumorigenesis. Clinically, the presence of pre-S2 deletion mutants in sera frequently develop resistance to nucleoside analogues anti-virals and predict HCC development. The pre-S2 deletion mutants and type II GGHs therefore represent novel biomarkers of HBV-related HCCs. A versatile DNA array chip has been developed to detect pre-S2 mutants in serum. Overall, the presence of pre-S2 mutants in serum has implications for anti-viral treatment and can predict HCC development. Targeting at pre-S2 mutant protein-induced, ER stress-dependent, mTOR signal cascade and metabolic disorders may offer potential strategy for chemoprevention or therapy in high risk chronic HBV carriers.
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Bang KB, Kim HJ. Management of antiviral drug resistance in chronic hepatitis B. World J Gastroenterol 2014; 20:11641-11649. [PMID: 25206270 PMCID: PMC4155356 DOI: 10.3748/wjg.v20.i33.11641] [Citation(s) in RCA: 18] [Impact Index Per Article: 1.6] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 10/21/2013] [Revised: 01/10/2014] [Accepted: 05/29/2014] [Indexed: 02/07/2023] Open
Abstract
Rescue antiviral treatment for patients with resistance to preexisting nucleos(t)ide analogues remains a clinical challenge. The correct choice of a first-line treatment of high potency and with a high genetic barrier to achieve sustained long-term suppression of viral replication provides the best chance of preventing treatment failure and the emergence of drug resistance. The management of treatment failure and drug resistance requires a precise and accurate clinical and virologic monitoring. Combination treatment with antiviral drugs that belong to different groups is associated with a lower chance of developing resistance to rescue drugs. To guarantee better control of viral replication in patients with drug resistance, the addition of another drug without a cross resistance profile should be given as early as possible, preferably at the time when genotypic resistance emerges. Long-term surveillance for treatment efficacy and possible emergence of drug resistance should be continued to prevent the emergence of multidrug-resistant strains.
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Hepatitis B virus PreS/S gene variants: pathobiology and clinical implications. J Hepatol 2014; 61:408-17. [PMID: 24801416 DOI: 10.1016/j.jhep.2014.04.041] [Citation(s) in RCA: 188] [Impact Index Per Article: 17.1] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 03/02/2014] [Revised: 04/21/2014] [Accepted: 04/24/2014] [Indexed: 12/16/2022]
Abstract
The emergence and takeover of hepatitis B virus (HBV) variants carrying mutation(s) in the preS/S genomic region is a fairly frequent event that may occur spontaneously or may be the consequence of immunoprophylaxis or antiviral treatments. Selection of preS/S mutants may have relevant pathobiological and clinical implications. Both experimental data and studies in humans show that several specific mutations in the preS/S gene may induce an imbalance in the synthesis of the surface proteins and their consequent retention within the endoplasmic reticulum (ER) of the hepatocytes. The accumulation of mutated surface proteins may cause ER stress with the consequent induction of oxidative DNA damage and genomic instability. Viral mutants with antigenically modified surface antigen may be potentially infectious to immune-prophylaxed patients and may account for cases of occult HBV infection. In addition, preS/S variants were reported to be associated with cases of fulminant hepatitis as well as of fibrosing cholestatic hepatitis, and they are associated with cirrhosis and hepatocellular carcinoma development.
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Chen X, Gao J, Ji Z, Zhang W, Zhang L, Xu R, Zhang J, Li F, Li S, Hu S, Shang L, Shao Z, Yan Y. A description of the hepatitis B virus genomic background in a high-prevalence area in China. Virol J 2014; 11:101. [PMID: 24884702 PMCID: PMC4074864 DOI: 10.1186/1743-422x-11-101] [Citation(s) in RCA: 10] [Impact Index Per Article: 0.9] [Reference Citation Analysis] [Abstract] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/25/2013] [Accepted: 05/20/2014] [Indexed: 01/15/2023] Open
Abstract
Background Hepatitis B (HB) is an important disease worldwide. Almost 350 million people are positive for Hepatitis B virus surface antigen (HBsAg), and one-third of them live in China. According to a nation-wide serosurvey in China in 2006, the prevalence of HBsAg was higher in Northwest China than in other areas. However, the epidemic HBV strains in this area are poorly studied. Results In this study, 242 complete hepatitis B virus (HBV) genome sequences were obtained from HBV asymptomatic carriers in major cities of Northwest China. The 242 HBV sequences clustered into genotypes B, C and D. Through comparison of the genotype consensus sequences, 158 genotype-dependent positions were observed in P, S and X ORFs. Clinically relevant mutation screening in this study revealed that no HBV antiviral drug resistance mutations were observed and the vaccination failure mutations were heavily underrepresented. Conclusions The role of genotype D strains in HBV prevalence should not be ignored in Northwest China. Due to low prevalence of vaccination failure mutations, it can be inferred that the genotype B, C and D strains in Northwest China may have less likelihood of vaccine escape.
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Affiliation(s)
| | | | | | | | | | | | | | | | | | | | | | - ZhongJun Shao
- Department of Epidemiology, School of Public Health, Fourth Military Medical University, No, 17, Changle west RD, Xi'an 710032, China.
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Analysis of complete nucleotide sequences of Angolan hepatitis B virus isolates reveals the existence of a separate lineage within genotype E. PLoS One 2014; 9:e92223. [PMID: 24632784 PMCID: PMC3954871 DOI: 10.1371/journal.pone.0092223] [Citation(s) in RCA: 12] [Impact Index Per Article: 1.1] [Reference Citation Analysis] [Abstract] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/21/2013] [Accepted: 02/19/2014] [Indexed: 02/07/2023] Open
Abstract
Hepatitis B virus genotype E (HBV/E) is highly prevalent in Western Africa. In this work, 30 HBV/E isolates from HBsAg positive Angolans (staff and visitors of a private hospital in Luanda) were genetically characterized: 16 of them were completely sequenced and the pre-S/S sequences of the remaining 14 were determined. A high proportion (12/30, 40%) of subjects tested positive for both HBsAg and anti-HBs markers. Deduced amino acid sequences revealed the existence of specific substitutions and deletions in the B- and T-cell epitopes of the surface antigen (pre-S1- and pre-S2 regions) of the virus isolates derived from 8/12 individuals with concurrent HBsAg/anti-HBs. Phylogenetic analysis performed with 231 HBV/E full-length sequences, including 16 from this study, showed that all isolates from Angola, Namibia and the Democratic Republic of Congo (n = 28) clustered in a separate lineage, divergent from the HBV/E isolates from nine other African countries, namely Cameroon, Central African Republic, Côte d'Ivoire, Ghana, Guinea, Madagascar, Niger, Nigeria and Sudan, with a Bayesian posterior probability of 1. Five specific mutations, namely small S protein T57I, polymerase Q177H, G245W and M612L, and X protein V30L, were observed in 79-96% of the isolates of the separate lineage, compared to a frequency of 0–12% among the other HBV/E African isolates.
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Xu Z, Wu G, Li F, Bai J, Xing W, Zhang D, Zeng C. Positive selection signals of hepatitis B virus and their association with disease stages and viral genotypes. INFECTION GENETICS AND EVOLUTION 2013; 19:176-87. [PMID: 23871771 DOI: 10.1016/j.meegid.2013.07.011] [Citation(s) in RCA: 7] [Impact Index Per Article: 0.6] [Reference Citation Analysis] [Track Full Text] [Subscribe] [Scholar Register] [Received: 04/02/2013] [Revised: 06/27/2013] [Accepted: 07/06/2013] [Indexed: 12/18/2022]
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