Allele loss and point mutation in codons 12 and 61 of the c-Ha-ras oncogene in carcinogen-transformed human breast epithelial cells.

There is significant evidence that the ras oncogene plays a role in experimental mammary carcinogenesis; the evidence in human breast cancer, however, is more limited. We induced the expression of transformation phenotypes in the human breast epithelial cell line MCF-10F with the chemical carcinogens 7,12-dimethylbenz[a]anthracene, N-methyl-N-nitrosourea, N-methyl-N-nitro-N'-nitrosoguanidine, and benzo[a]pyrene. This work was designed to clarify whether chemically induced neoplastic transformation correlates with alterations in the ras gene. MCF-10F cells have two c-Ha-ras alleles, identified by 1.0-kb and 1.2-kb restriction fragments. Treatment with carcinogens resulted in the loss of one of the alleles (1.0 kb). Polymerase chain reaction-amplified DNA from all carcinogen-treated cells was analyzed for point mutations in c-Ha-ras at codons 12 and 61. All of the carcinogens induced a mutation of the remaining allele at the first position of codon 12 (GGC-->AGC). Another frequent mutation occurred at the first position of codon 61 (CAG-->GAG). The changes in c-Ha-ras were associated with the emergence of colony formation in agar-methocel, but no specific changes in this gene correlated with the emergence of invasiveness or tumorigenesis, indicating that other genes may be involved in the process.