HEPATOMA-DERIVED INTEGRATED HBV DNA CAUSES MULTISTAGE TRANSFORMATION IN-VITRO

The hepatoma-derived hepatitis B virus (HBV) DNA insert HU-a has recen tly been shown to contain two viral transactivator genes, X and preS2/ S-t, We report here that HU-a induces malignant transformation after s table transfection of the fetal mouse hepatocyte line FMH202, as indic ated by soft agar growth and nude mouse tumorigenicity, Transfections with HU-a subclones, containing the X gene or the preS2/S-t gene alone or sequences without transactivator gene, respectively, suggested tha t the X gene is essential for transformation, Sequential stages of tra nsformation and tumor progression were analysed by injection of the st ably transfected FMH202 lines into nude mice, explantation of the resu lting tumors and re-establishment of cell lines from the tumors, Compa rison of two HU-a-transformed cell lines by HBV mRNA hybridization, So uthern analysis and chromosomal in situ hybridization revealed that in tegrated HBV DNAs were involved in major chromosomal rearrangements in both cases, Interestingly, recombination of the HBV DNA insert during the nude mouse passage had completely abolished HBV-specific transcri ption in one case, indicating that expression of integrated HBV genes, while presumably involved in early transformation, is dispensable at later stages of tumor progression, The sequential transformation obser ved in this experimental system suggests that expression of the X gene by integrated viral DNA and subsequent hepatocyte genome mutations mi ght both contribute to HBV-associated liver carcinogenesis.