In this revised application renewal of funds is requested for continuation of the research on the mechanism of selenium and beta-carotene in prevention of 7, 12-dimethylbenz(a)anthracene (DMBA)-induced neoplastic transformation of the mouse mammary cells in vitro. The mammary epithelial cells transformed in organ culture by DMBA are potentially neoplastic, these cells produce hyperplastic alveolar outgrowth (HAG) and mammary tumors after transplantation into the mammary fat pad of syngeneic female mice. The process is also marked by the appearance of nodule-like alveolar lesion (NLAL) which serves as marker of transformation in the organ in a hormonally defined serum-free culture medium. Thus the mammary cells transformed by DMBA in vitro mimic the multistep mouse mammary carcinogenesis in vivo. This unique culture model of the whole mammary organ will be used to investigate the mechanism of the chemopreventive action of selenium and beta-carotene while acting as an inhibitor of the neoplastic transformation process. Both selenium and beta- carotene inhibits transformation of cells induced by physical, chemical and viral agents, acting in the animals and in the culture medium acting at the initiation and the promotional stages. The proposed research will involve investigation on a possible influence of the selenium and/or beta-carotene at the level of expression of the P1-450 genes, intervention at the levels of P-450 monooxygenase metabolic activation of the procarcinogen, generating the active carcinogens. Cellular concentration of the enzyme proteins, the electrophilic metabolites, and the pattern of adduct formation. Studies will also include determination whether DMBA entrapped in the mammary fat pad is metabolized by the monooxygenase system with resultant DNA adduct formation. Also influence of the free-radical modifier superoxide dismutase (SOD) acting individually or in combination with the dietary agents will be the DMBA transformation process promoted by TPA and/or hormones will be analyzed. A role of the dietary agents on carcinogen-DNA damage also will be measured. The results of the comprehensive and focused research will provide important knowledge in elucidation of the mode of action of selenium and/or beta-carotene in prevention of the neoplastic disease.
