LOSS OF P53 PROTEIN DURING RADIATION TRANSFORMATION OF PRIMARY HUMAN MAMMARY EPITHELIAL-CELLS

The causative factors leading to breast cancer are largely unknown. In creased incidence of breast cancer following diagnostic or therapeutic radiation suggests that radiation may contribute to mammary oncogenes is. This report describes the in vitro neoplastic transformation of a normal human mammary epithelial cell strain, 76N, by fractionated gamm a-irradiation at a clinically used dose (30 Gy). The transformed cells (76R-30) were immortal, had reduced growth factor requirements, and p roduced tumors in nude mice. Remarkably, the 76R-30 cells completely l acked the p53 tumor suppressor protein. Loss of p53 was due to deletio n of the gene on one allele and a 26-bp deletion within the third intr on on the second allele which resulted in abnormal splicing out of eit her the third or fourth exon from the mRNA. PCR with a mutation-specif ic primer showed that intron 3 mutation was present in irradiated cell s before selection for immortal phenotype. 76R-30 cells did not exhibi t G, arrest in response to radiation, indicating a loss of p53-mediate d function. Expression of the wild-type p53 gene in 76R-30 cells led t o their growth inhibition. Thus, loss of p53 protein appears to have c ontributed to neoplastic transformation of these cells. This unique mo del should facilitate analyses of molecular mechanisms of radiation-in duced breast cancer and allow identification of p53-regulated cellular genes in breast cells.