CHARACTERIZATION OF HUMAN BUCCAL EPITHELIAL-CELLS TRANSFECTED WITH THE SIMIAN-VIRUS-40 T-ANTIGEN GENE

Serum-free cultures of normal human buccal epithelial cells were trans fected with a plasmid containing the SV40 T-antigen (SV40T) gene. Two major lines developed that showed extended lifespans (between 30 and 4 0 weeks) as compared with the controls (similar to 6 weeks). Continued growth through one or two crises generated several sublines. They exp ressed the epithelial marker keratin and also exhibited nuclear expres sion of SV40T. The lines showed abnormal karyotypes with both numerica l and structural aberrations and variably responded to agents that nor mally inhibit growth and/or induce terminal differentiation, i.e. tran sforming growth factor-beta 1 and fetal bovine serum. One of the lines , termed SVpgC2a, developed into an apparently immortal line, since it had undergone more than 700 population doublings from over 2 years in culture. Further characterization of this line demonstrated its clona l origin, with integration of two copies of SV40T at the same site and the presence of both normal retinoblastoma and wild-type p53 proteins . This line showed high resistance to growth inhibition by transformin g growth factor-pr and serum similar to that shown by buccal carcinoma cell line SqCC/Y1. Neither SVpgC2a nor its parental lines were tumori genic when injected into athymic nude mice, whereas the SqCC/Y1 cells induced tumors. The various lines with extended but finite lifespans, complemented by one immortalized line, which retained non-malignant pr operties upon extended culture, provide a battery of model systems tha t will be useful for studying mechanisms of human oral carcinogenesis.