Malignant transformation of immortalized transgenic hepatocytes after transfection with hepatitis B virus DNA.

Abstract

Persistent infection by hepatitis B virus (HBV) is epidemiologically correlated with the prevalence of hepatocellular carcinoma, but its role in tumor development is not yet understood. To study the putative oncogenic potential of HBV, a non‐malignant immortal mouse hepatocyte line FMH202 harboring metallothionein promoter‐driven simian virus 40 large tumor antigen was transfected with HBV DNA. All stably transfected clones which replicated HBV displayed malignant growth characteristics in soft agar and were tumorigenic upon inoculation in nude mice. The nude mice tumors were histologically classified as differentiated or anaplastic hepatocellular carcinomas. As with human liver carcinomas, rearrangements of in vitro integrated HBV sequences were observed in the nude mouse tumors, and in tumor‐derived cell lines. In one case, expression of viral core and surface antigens was blocked in the tumors, correlating with hypermethylation of the HBV genome. However, the expression of X gene was maintained in most tumors and tumor‐derived cell lines. X protein was detected in nuclei by immune fluorescence and by immune blot. These results provide the first demonstration that HBV displays oncogenic potential in an experimental system. This system could be useful to functionally identify HBV genes which convey a tumorigenic phenotype.

Hepatitis B virus X mutants, present in hepatocellular carcinoma tissue abrogate both the antiproliferative and transactivation effects of HBx

“…However, in a di erent genetic context, HBx transgenic mice show either no obvious pathology (Lee et al, 1990), only increased susceptibility to chemical carcinogens (Slagle et al, 1996), or acceleration of the development of c-mycinduced HCC (Terradillos et al, 1997). Only a few studies have shown transformation of certain cell types by HBx in vitro (Hohne et al, 1990;Shirakata et al, 1989). Thus, the actual impact of HBx in liver carcinogenesis and the mechanisms of its action remain largely obscure.…”

Activated ras oncogene collaborates with HBx gene of hepatitis B virus to transform cells by suppressing HBx-mediated apoptosis

“…Numerous attempts have been made to examine the oncogenic potential of HBx in cell culture (Hohne et al, 1990;Oguey et al, 1996;Shirakata et al, 1989;Tarn et al, 1999). However, its transforming ability was barely measurable only when cells were immortalized by other oncogenes, such as SV40 T-antigen (Hohne et al, 1990) or TGF-a (Oguey et al, 1996;Tarn et al, 1999).…”

Partial characterization of the woodchuck tumor suppressor, p53, and its interaction with woodchuck hepatitis virus X antigen in hepatocarcinogenesis

“…The fact that HBxAg and WHxAg are also structurally and functionally similar (Galibert et al, 1982), as are both human and woodchuck p53s, further suggests that they may share common properties, possibly including some of the mechanistic steps whereby these viruses mediate hepatocellular transformation. Given the increasing importance of HBxAg to transformation 1994;Shirakata et al, 1989;Hohne et al, 1990;Koike et al, 1994), combined with its complexing to wild type p53 (Feitelson et al, 1993a;Ueda et al, 1995;Truant et al, 1995), it was of great interest to see whether this also occurred in the WHV system. The results of this study show that WHxAg and woodchuck p53 form complexes both in vitro and in vivo in a manner analogous to that which occurs in human carriers with HCC, providing at least part of a common mechanism whereby these viruses cause hepatocellular carcinoma.…”

“…The ®nding that HBxAg is often expressed from viral DNA fragments integrated into human liver or HCC nodules, and that the HBxAg made is still capable of trans-activation (Zahm et al, 1988;Wollersheim et al, 1988), suggest that functional HBxAg is associated with the development of these tumors, although it is not known whether HBxAg associated trans-activation is relevant to hepatocarcinogenesis. In addition, the ®nding that HBxAg transforms NIH3T3 cells (Shirakata et al, 1989) and a nontransformed mouse hepatocyte cell line (Hohne et al, 1990), suggests that under selected conditions, HBxAg can mediate transformation of cultured cells. Further, the ®nding that HBxAg complexes with the tumor suppressor product, p53, in the majority of HBV associated HCC Figure 6 Complex formation between woodchuck p53 and WHxAg in vivo.…”

“…For example, HB6Ag expression is sustained at high levels in chronic carriers who develop HCC, and in many cases, it is the only HBV encoded product detected by immunohistochemistry 1991a). Further, HB6Ag causes tumorigenic transformation of NIH3T3 cells (Shirakata et al, 1989), and of an untransformed mouse hepatocyte cell line (Hohne et al, 1990). More recently, HB6Ag has been shown to complex with and inactivate p53 both in vitro and in vivo (Feitelson et al, 1993a;Wang et al, 1994;Truant et al, 1995), suggesting that this relationship may be important for tumor development.…”