title: Identification of transforming hepatitis B virus S gene nonsense mutations derived from freely replicative viruses in hepatocellular carcinoma.
abstract: OBJECTIVE
  The correlation between chronic hepatitis B virus (HBV) infection and hepatocellular carcinoma (HCC) has been well-established. But the roles of viral factor remain uncertain. Only HBV X gene and nonsense mutations of S gene (C-terminal truncation of HBV surface protein) have been demonstrated to have transforming activity. Whether they play a significant role in hepatocarcinogenesis is still uncertain.
  
  
  METHODS
  Twenty-five HBV-related HCC patients were positive for hepatitis B core antigen (HBcAg) in the cancerous parts of their HCC liver tissues by immunohistochemistry studies, and had available tissue for whole HBV genome sequence analysis. The results were compared with 25 gender and age-matched HBcAg negative HCCs. Plasmids encoding HBV S gene nonsense mutations identified from HBcAg (+) HCC tissue were constructed to investigate their cell proliferation, transformation activity and the oncogenic potentials by xenograft study and in vivo migration assay.
  
  
  RESULTS
  HBcAg (+) HCC patients were significantly associated with cirrhosis and small tumor size (≦2 cm) when compared with HBcAg (-) HCC patients. Southern blot analyses revealed freely replicative forms of HBV in the cancerous parts of HBcAg(+) HCC. Three nonsense mutations of S gene (sL95*, sW182*, and sL216*) were identified in the HBcAg(+) HCC tumor tissues. sW182* and sL216* were recurrently found in the 25 HBcAg (-) HCC tumor tissue, too. Functional studies of the above 3 non-sense mutations all demonstrated higher cell proliferation activities and transformation abilities than wild type S, especially sW182*. Tumorigenicity analysis by xenograft experiments and in vitro migration assay showed potent oncogenic activity of sW182* mutant.
  
  
  CONCLUSIONS
  This study has demonstrated potent oncogenic activity of nonsense mutations of HBV S gene, suggesting they may play an important role in hepatocarcinogenesis.
authors.affiliation: Institute of Molecular and Genomic Medicine, National Health Research Institutes, Miaoli, Taiwan ; Department of Pathology, Chang-Gung Memorial Hospital, Taoyuan, Taiwan ; Department of Pathology, Tzu-Chi General Hospital, Taipei Branch, Tzu-Chi University School of Medicine, Hualien, Taiwan.
authors.forename: Shiu-Feng
authors.initials: SF
authors.lastname: Huang
authors.affiliation: Institute of Molecular and Genomic Medicine, National Health Research Institutes, Miaoli, Taiwan.
authors.forename: Ya-Ting
authors.initials: YT
authors.lastname: Chen
authors.affiliation: Department of General Surgery, Chang-Gung Memorial Hospital, Chang-Gung University College of Medicine, Taoyuan, Taiwan.
authors.forename: Wei-Chen
authors.initials: WC
authors.lastname: Lee
authors.affiliation: Institute of Molecular and Genomic Medicine, National Health Research Institutes, Miaoli, Taiwan.
authors.forename: Il-Chi
authors.initials: IC
authors.lastname: Chang
authors.affiliation: Institute of Molecular and Genomic Medicine, National Health Research Institutes, Miaoli, Taiwan.
authors.forename: Yu-Ting
authors.initials: YT
authors.lastname: Chiu
authors.affiliation: Institute of Molecular and Genomic Medicine, National Health Research Institutes, Miaoli, Taiwan.
authors.forename: Yu
authors.initials: Y
authors.lastname: Chang
authors.affiliation: Institute of Molecular and Genomic Medicine, National Health Research Institutes, Miaoli, Taiwan.
authors.forename: Hsiao-Chen
authors.initials: HC
authors.lastname: Tu
authors.affiliation: Institute of Molecular and Genomic Medicine, National Health Research Institutes, Miaoli, Taiwan.
authors.forename: Chiou-Hwa
authors.initials: CH
authors.lastname: Yuh
authors.affiliation: Institute of Molecular and Genomic Medicine, National Health Research Institutes, Miaoli, Taiwan.
authors.forename: Isao
authors.initials: I
authors.lastname: Matsuura
authors.affiliation: Department of Pathology, Chang-Gung Memorial Hospital, Taoyuan, Taiwan ; Department of Pathology, Tzu-Chi General Hospital, Dalin Branch, Chiayi, Taiwan.
authors.forename: Liang-Yu
authors.initials: LY
authors.lastname: Shih
authors.affiliation: Department of Pediatrics, Chang-Gung Memorial Hospital, Chang-Gung University College of Medicine, Taoyuan, Taiwan.
authors.forename: Ming-Wei
authors.initials: MW
authors.lastname: Lai
authors.affiliation: Department of Applied Mathematics and Institute of Statistics, National Chung-Hsing University, Taichung, Taiwan.
authors.forename: Hong-Dar Isaac
authors.initials: HD
authors.lastname: Wu
authors.affiliation: Department of General Surgery, Chang-Gung Memorial Hospital, Chang-Gung University College of Medicine, Taoyuan, Taiwan.
authors.forename: Miin-Fu
authors.initials: MF
authors.lastname: Chen
authors.affiliation: Liver Research Center, Chang-Gung Memorial Hospital, Chang-Gung University College of Medicine, Taoyuan, Taiwan.
authors.forename: Chau-Ting
authors.initials: CT
authors.lastname: Yeh
mesh_terms: D015139:Blotting, Southern; D006528:Carcinoma, Hepatocellular / Q000150:complications / Q000473:pathology / Q000821:virology*; D002471:Cell Transformation, Neoplastic / Q000235:genetics; D018389:Codon, Nonsense / Q000235:genetics; D004721:Endoplasmic Reticulum / Q000378:metabolism; D006509:Hepatitis B / Q000150:complications*; D006515:Hepatitis B virus / Q000235:genetics* / Q000378:metabolism; D006801:Humans; D007150:Immunohistochemistry; D008113:Liver Neoplasms / Q000150:complications / Q000473:pathology / Q000821:virology*; D010957:Plasmids / Q000235:genetics; D017422:Sequence Analysis, DNA; D014760:Viral Fusion Proteins / Q000235:genetics*
chemical_list: D018389:Codon, Nonsense; D014760:Viral Fusion Proteins