Whole genome HBV deletion profiles and the accumulation of preS deletion mutant during antiviral treatment

BMC Microbiology

Changqing Zeng¹  Huiguo Ding⁵  Ulrike Protzer⁴  Feifei Li²  Wei Chen²  Christian Bach⁴  Libin Deng³  Ke Zhang⁴  Peiling Dong⁶  Dake Zhang² 

[1]No.7 Beitucheng West Road, Chaoyang District, Beijing, 100029, China

[2]Laboratory of Disease Genomics and Individualized Medicine, Beijing Institute of Genomics, Chinese Academy of Sciences, Beijing, China

[3]Basic Medical Institute of Nanchang University, Nanchang, China

[4]Institute of Virology, Technische Universität München / Helmholtz Zentrum München - German Research Center for Environmental Health, München, Germany

[5]No.8 Xitoutiao, You An Men, Beijing, 100069, China

[6]Beijing Youan Hospital, Capital Medical University, Beijing, China

关键词: Nucleotide analog;    Antiviral therapy;    Chronic hepatitis;    PreS;    Deletion;    HBV;   

Others  :  1144664
DOI  :  10.1186/1471-2180-12-307

 received in 2012-01-17, accepted in 2012-12-22,  发布年份 2012

【 摘 要 】

Background

Hepatitis B virus (HBV), because of its error-prone viral polymerase, has a high mutation rate leading to widespread substitutions, deletions, and insertions in the HBV genome. Deletions may significantly change viral biological features complicating the progression of liver diseases. However, the clinical conditions correlating to the accumulation of deleted mutants remain unclear. In this study, we explored HBV deletion patterns and their association with disease status and antiviral treatment by performing whole genome sequencing on samples from 51 hepatitis B patients and by monitoring changes in deletion variants during treatment. Clone sequencing was used to analyze preS regions in another cohort of 52 patients.

Results

Among the core, preS, and basic core promoter (BCP) deletion hotspots, we identified preS to have the highest frequency and the most complex deletion pattern using whole genome sequencing. Further clone sequencing analysis on preS identified 70 deletions which were classified into 4 types, the most common being preS2. Also, in contrast to the core and BCP regions, most preS deletions were in-frame. Most deletions interrupted viral surface epitopes, and are possibly involved in evading immuno-surveillance. Among various clinical factors examined, logistic regression showed that antiviral medication affected the accumulation of deletion mutants (OR = 6.81, 95% CI = 1.296 ~ 35.817, P = 0.023). In chronic carriers of the virus, and individuals with chronic hepatitis, the deletion rate was significantly higher in the antiviral treatment group (Fisher exact test, P = 0.007). Particularly, preS2 deletions were associated with the usage of nucleos(t)ide analog therapy (Fisher exact test, P = 0.023). Dynamic increases in preS1 or preS2 deletions were also observed in quasispecies from samples taken from patients before and after three months of ADV therapy. In vitro experiments demonstrated that preS2 deletions alone were not responsible for antiviral resistance, implying the coordination between wild type and mutant strains during viral survival and disease development.

Conclusions

We present the HBV deletion distribution patterns and preS deletion substructures in viral genomes that are prevalent in northern China. The accumulation of preS deletion mutants during nucleos(t)ide analog therapy may be due to viral escape from host immuno-surveillance.

【 授权许可】

   
2012 Zhang et al.; licensee BioMed Central Ltd.

【 图 表 】

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【 参考文献 】

• [1]Locarnini S, Zoulim F: Molecular genetics of HBV infection. Antivir Ther 2010, 15(Suppl 3):3-14.
• [2]Kim BK, Revill PA, Ahn SH: HBV genotypes: relevance to natural history, pathogenesis and treatment of chronic hepatitis B. Antivir Ther 2011, 16(8):1169-1186.
• [3]Gunther S: Genetic variation in HBV infection: genotypes and mutants. J Clin Virol 2006, 36(Suppl 1):S3-S11.
• [4]Preikschat P, Gunther S, Reinhold S, Will H, Budde K, Neumayer HH, Kruger DH, Meisel H: Complex HBV populations with mutations in core promoter, C gene, and pre-S region are associated with development of cirrhosis in long-term renal transplant recipients. Hepatology 2002, 35(2):466-477.
• [5]Marschenz S, Brinckmann A, Nurnberg P, Kruger DH, Gunther S, Meisel H: Co-replication analyses of naturally occurring defective hepatitis B virus variants with wild-type. Virology 2008, 372(2):247-259.
• [6]Ferns RB, Naoumov NV, Gilson RJ, Tedder RS: Presence of hepatitis B virus core promoter mutations pre-seroconversion predict persistent viral replication after HBeAg loss. J Clin Virol 2007, 39(3):199-204.
• [7]Zhu P, Tan D, Peng Z, Liu F, Song L: Polymorphism analyses of hepatitis B virus X gene in hepatocellular carcinoma patients from southern China. Acta Biochim Biophys Sin (Shanghai) 2007, 39(4):265-272.
• [8]Liu XH, Lin J, Zhang SH, Zhang SM, Feitelson MA, Gao HJ, Zhu MH: COOH-terminal deletion of HBx gene is a frequent event in HBV-associated hepatocellular carcinoma. World J Gastroenterol 2008, 14(9):1346-1352.
• [9]Hung JH, Su IJ, Lei HY, Wang HC, Lin WC, Chang WT, Huang W, Chang WC, Chang YS, Chen CC, et al.: Endoplasmic reticulum stress stimulates the expression of cyclooxygenase-2 through activation of NF-kappaB and pp 38 mitogen-activated protein kinase. J Biol Chem 2004, 279(45):46384-46392.
• [10]Wang LH, Huang W, Lai MD, Su IJ: Aberrant cyclin A expression and centrosome overduplication induced by hepatitis B virus pre-S2 mutants and its implication in hepatocarcinogenesis. Carcinogenesis 2012, 33(2):466-472.
• [11]Wang HC, Huang W, Lai MD, Su IJ: Hepatitis B virus pre-S mutants, endoplasmic reticulum stress and hepatocarcinogenesis. Cancer Sci 2006, 97(8):683-688.
• [12]Yeung P, Wong DK, Lai CL, Fung J, Seto WK, Yuen MF: Association of hepatitis B virus pre-S deletions with the development of hepatocellular carcinoma in chronic hepatitis B. J Infect Dis 2011, 203(5):646-654.
• [13]Abe K, Thung SN, Wu HC, Tran TT, Le Hoang P, Truong KD, Inui A, Jang JJ, Su IJ: Pre-S2 deletion mutants of hepatitis B virus could have an important role in hepatocarcinogenesis in Asian children. Cancer Sci 2009, 100(12):2249-2254.
• [14]Fang ZL, Sabin CA, Dong BQ, Wei SC, Chen QY, Fang KX, Yang JY, Huang J, Wang XY, Harrison TJ: Hepatitis B virus pre-S deletion mutations are a risk factor for hepatocellular carcinoma: a matched nested case–control study. J Gen Virol 2008, 89(Pt 11):2882-2890.
• [15]Yuan TT, Lin MH, Chen DS, Shih C: A defective interference-like phenomenon of human hepatitis B virus in chronic carriers. J Virol 1998, 72(1):578-584.
• [16]Milich DR, McLachlan A, Moriarty A, Thornton GB: Immune response to hepatitis B virus core antigen (HBcAg): localization of T cell recognition sites within HBcAg/HBeAg. J Immunol 1987, 139(4):1223-1231.
• [17]Weber B: Genetic variability of the S gene of hepatitis B virus: clinical and diagnostic impact. J Clin Virol 2005, 32(2):102-112.
• [18]Chen BF, Liu CJ, Jow GM, Chen PJ, Kao JH, Chen DS: High prevalence and mapping of pre-S deletion in hepatitis B virus carriers with progressive liver diseases. Gastroenterology 2006, 130(4):1153-1168.
• [19]Fukuda R, Ishimura N, Kushiyama Y, Moriyama N, Ishihara S, Chowdhury A, Tokuda A, Sakai S, Akagi S, Watanabe M, et al.: Hepatitis B virus with X gene mutation is associated with the majority of serologically “silent” non-b, non-c chronic hepatitis. Microbiol Immunol 1996, 40(7):481-488.
• [20]Uchida T, Gotoh K, Shikata T: Complete nucleotide sequences and the characteristics of two hepatitis B virus mutants causing serologically negative acute or chronic hepatitis B. J Med Virol 1995, 45(3):247-252.
• [21]Moriyama K: Reduced antigen production by hepatitis B virus harbouring nucleotide deletions in the overlapping X gene and precore-core promoter. J Gen Virol 1997, 78(Pt 6):1479-1486.
• [22]Pal J, Nyarady Z, Marczinovits I, Par A, Ali YS, Berencsi G, Kvell K, Nemeth P: Comprehensive regression analysis of hepatitis B virus X antigen level and anti-HBx antibody titer in the sera of patients with HBV infection. Pathol Oncol Res 2006, 12(1):34-40.
• [23]Stemler M, Weimer T, Tu ZX, Wan DF, Levrero M, Jung C, Pape GR, Will H: Mapping of B-cell epitopes of the human hepatitis B virus X protein. J Virol 1990, 64(6):2802-2809.
• [24]Glebe D, Urban S: Viral and cellular determinants involved in hepadnaviral entry. World J Gastroenterol 2007, 13(1):22-38.
• [25]Locarnini S, McMillan J, Bartholomeusz A: The hepatitis B virus and common mutants. Semin Liver Dis 2003, 23(1):5-20.
• [26]Winters MA, Coolley KL, Cheng P, Girard YA, Hamdan H, Kovari LC, Merigan TC: Genotypic, phenotypic, and modeling studies of a deletion in the beta3-beta4 region of the human immunodeficiency virus type 1 reverse transcriptase gene that is associated with resistance to nucleoside reverse transcriptase inhibitors. J Virol 2000, 74(22):10707-10713.
• [27]Cho SW, Hahm KB, Kim JH: Reversion from precore/core promoter mutants to wild-type hepatitis B virus during the course of lamivudine therapy. Hepatology 2000, 32(5):1163-1169.
• [28]Ohkawa K, Takehara T, Kato M, Deguchi M, Kagita M, Hikita H, Sasakawa A, Kohga K, Uemura A, Sakamori R, et al.: Supportive role played by precore and preS2 genomic changes in the establishment of lamivudine-resistant hepatitis B virus. J Infect Dis 2008, 198(8):1150-1158.
• [29]Kondo Y, Asabe S, Kobayashi K, Shiina M, Niitsuma H, Ueno Y, Kobayashi T, Shimosegawa T: Recovery of functional cytotoxic T lymphocytes during lamivudine therapy by acquiring multi-specificity. J Med Virol 2004, 74(3):425-433.
• [30]Menne S, Tennant BC, Gerin JL, Cote PJ: Chemoimmunotherapy of chronic hepatitis B virus infection in the woodchuck model overcomes immunologic tolerance and restores T-cell responses to pre-S and S regions of the viral envelope protein. J Virol 2007, 81(19):10614-10624.
• [31]Park JH, Lee MK, Kim HS, Kim KL, Cho EW: Targeted destruction of the polymerized human serum albumin binding site within the preS2 region of the HBV surface antigen while retaining full immunogenicity for this epitope. J Viral Hepat 2003, 10(1):70-79.
• [32]Minami M, Okanoue T, Nakajima E, Yasui K, Kagawa K, Kashima K: Significance of pre-S region-defective hepatitis B virus that emerged during exacerbation of chronic type B hepatitis. Hepatology 1993, 17(4):558-563.
• [33]Chinese Society of Hepatology and Chinese Society of Infectious Diseases, Chinese Medical Association: Guideline on prevention and treatment of chronic hepatitis B in China (2010). Chin J Front Med Sci 2011, 3(1):16.
• [34]Gunther S, Li BC, Miska S, Kruger DH, Meisel H, Will H: A novel method for efficient amplification of whole hepatitis B virus genomes permits rapid functional analysis and reveals deletion mutants in immunosuppressed patients. J Virol 1995, 69(9):5437-5444.
• [35]Jazayeri SM, Carman WF: Virus Escape CTL or B Cell Epitopes? Hepatitis Monthly 2005, 5(4):133-136.