The thrust of this study is to elucidate the genetic mechanism of neoplastic transformation of normal tissues. The experimental system centers in a 3.1 kilobasepair (kb), moderately cell- transforming human oncogene, hhcM, isolated from an African hepatocellular carcinoma line, Mahlavu. Using hhcM as probe, we isolated four other genomic clones of related DNA sequences from Chinese and Korean hepatomas; the cell-transformation capability of each on NIH3T3 cells had been ascertained. Transformation of Buffalo rat liver cell (BRL-1) has been achieved by transfection with hhcM 3.1 kb DNA, subcloned (rpNpM-1) in an expression vector carrying a neomycin resistance (Neor) marker and a SV40 promoter. Transformed BRL-1 cells, selected by colony-formation in soft-agar, were highly tumorigenic in athymic NIH Swiss nu/nu mice. A more diversified tumorigenesis was observed with rpNpM-1 DNA transformed NIH3T3 cells. Both solid tumor (75%) at the site of inoculation and B-cell lymphomas (40%) were induced with some challenged mice developed both tumors. As an on-going interest the role and relationship of hepatitis B virus (HBV) with hhcM in hepatocarcinogenesis were analyzed. Results of nucleotide homology search indicated that the human flanking sequence for HBV integration was seen in hhcM , involving a PstI recognition sequence. Only 4/6 nucleotides of PstI was seen at the same location of the normal liver homologue. HBV sequence was seen at a 2.0 kb site elsewhere in the hepatoma. Several aflatoxin B1 mutagenized hhcM clones were recovered from the tumor developed by inoculating athymic nu/nu mice with NIH3T3 cells transfected with AFB1-bound hhcM 3.1 kb DNA. These mutagenized hhcM clones would be analyzed to resolve the specific dG involved in the AFB1 induced mutation.
