The goals of the proposal are the investigation of the role played by insulin-like growth factors (IGFs) in the biology of human breast cancer and of the mechanism by which these factors stimulate mitogenesis in this system. the long term aim is to use these studies to define the biochemical events underlying tumor progression in this disease and to identify new targets for therapeutic intervention. Previous work has shown that IGF-I and IGF-II elicit a marked mitogenic response in some breast cancer cell lines by binding to the IGF-I receptor. Moreover, overexpression of IGF-II in an estrogen-dependent cell line confers some aspects of estrogen independence. In addition, stromal cells in breast tumors express IGF-II, whereas tumor cells do in only a minority of cases. Taken together, the data imply that IGFs are potential paracrine regulators of breast tumor cell growth and that expression of IGF-II by the tumor cell may be an event in tumor progression that allows the cell to become independent of stromal factors. The current proposal aims to define the effects of IGFs on breast cancer cell growth in more detail, exploring the interactions of IGFs with hormonal agents and retinoic acid and correlating growth effects with IGF-I receptor expression, tyrosine kinase activity and autophosphorylation. The system will be then used as a model with which to study the mechanistic details of the transduction of the proliferative signal by IGFs. Preliminary results reveal that IGFs activate the IGF-I receptor tyrosine kinase with subsequent phosphorylation of its beta- subunit and several other substrates, including pp185 (IRS1), GAP, and GAP-associated pp62. Current studies focus on further defining these events and in identifying their functional consequences. Interactions of pp185 and GAP with other proteins will be characterized and assessed in IGF-responsive and unresponsive cell lines, IGF-II transfectants, and cells treated with antiestrogens and differentiation agents. Specific events and interactions will be correlated with aspects of the proliferative and transformed phenotype. Transfections with GAP expression vectors will be performed to evaluate the role of the GAP-ras complex in mediating these events.