Chromosomal and c-K-ras oncogene alterations induced by a chemical carcinogen and phorbol ester in skin fibroblasts of individuals with familial polyposis coli.

Skin fibroblasts derived from three patients with familial polyposis coli (FPC) were treated in vitro with N-methyl-N'-nitro-N-nitrosoguanidine (MNNG) alone or in combination with the tumor promoter, 12-O-tetradecanoylphorbol-13-acetate (TPA). None of the cultures treated four times with MNNG alone (1 microgram/ml) or in combination with TPA (eight applications, 0.1 microgram/ml each) showed either morphological transformation or anchorage-independent growth for 18 months after treatments. FPC cells treated with MNNG alone showed cell growth inhibition, breakage and loss of chromosomes, as well as the deletion of 5.7 kilobase (kb) EcoRI fragment in the c-K-ras locus as detected by Southern blot analysis with v-K-ras specific probe (clone KBE-2). The control cells contained two EcoRI fragments of 5.7 and 4.2 kb. On the other hand, cells treated with MNNG first and then followed by treatment with TPA not only showed an increase in the rate of cell growth but also exhibited two novel EcoRI fragments of 9.4 and 12 kb. Treatment with TPA alone appeared to stimulate cell division long after application and to induce chromosomal aberrations but had no effect on c-K-ras sequences. The most likely explanation for the appearance of chromosome pulverization and hyperploidy in FPC cells treated with TPA is the induction of premature chromosome condensation of the S phase cells due to fusion with the mitotic chromosomes.