Our long term objective is to induce preneoplastic and neoplastic transformation of human mammary epithelial cells, under defined in vitro conditions, with direct- and indirect-acting chemical carcinogens. An ability to consistently transform human mammary cells under defined in vitro conditions is essential to designing experiments to understand the cellular-molecular events underlying transformation and histogenesis of human breast cancer(s). Our strategy is to transform human epithelial cells grown under serum- free conditions in a collagen gel culture system, as well as in monolayer culture systems using various matrices. Specifically, we will: a) analyze the growth regulation of normal, benign (fibrocystic) and carcinomatous human mammary cells in collagen culture, under serum-free conditions in the presence of hormones (estrogen, progesterone, prolactin, etc.), growth factors (EGF, FGF, TGF alpha, TGF beta, etc.) and nonhormonal agents (lipids, TPA, Li+); b) use these endpoints to design culture conditions which facilitate human mammary epithelial cell transformation at high yield, in a manner similar to those developed for transformation of murine epithelial cells; c) use cDNA probes to determine whether amplification, rearrangement or expression of plasminogen activators (tPA, urokinase), TGF alpha, TGF beta, EGF receptor, glutathione-s-transferase-P, gamma-glutamyl transpeptidase and 0-6 alkyl-DNA-alkyltransferase are reliable, quantitative molecular markers for transformation and tumor histogenesis; d) determine whether changes at the molecular level correlate with phenotype changes (karyotypic, immunocytochemical, ultramicroscopic); e) develop suitable in vitro and in vivo assay systems to identify those molecular-cellular changes which represent preneoplastic and neoplastic (malignant) transformants. Our experience and approach suggest that development of a system(s) for transformation of human mammary epithelial cell in vitro is an achievable goal.
