The loss of growth control characteristic of carcinogenesis is uniformly accompanied by alterations in normal pathways of differentiation and development. This implies an intrinsic relationship between these processes. The existence of endocrine risk factors for breast cancer that are related to the timing of normal developmental events such as menarche, menopause and age at first full-term pregnancy epitomizes the relationship between development and carcinogenesis in the breast. Understanding the mechanism by which reproductive events influence breast cancer susceptibility will undoubtedly require a far better understanding of normal mammary development than currently exists, particularly with respect to identifying the genes that control proliferation and differentiation. Since major insights into the molecular mechanisms of differentiation, development, and carcinogenesis have been obtained in a variety of systems through studies of protein kinases, we have chosen to focus on this family of molecules. Several members of the protein kinase family have been shown to contribute to mammary carcinogenesis both in humans and in rodent model systems. Moreover, over-expression of genes encoding certain protein kinases, such as HER2/Neu, has been shown to provide prognostic information relevant to clinical outcome and response to therapy. As such, further studies of the function of protein kinases in the breast may reveal significant features of the relationship between development and carcinogenesis in this organ, as well provide insight into how the decision to proliferate or differentiate is made in mammary epithelial cells. We have identified two novel protein kinases, Hunk and Punc, that are: differentially regulated in the mammary gland during pregnancy; expressed in spatially restricted subsets of epithelial cells during specific stages of pregnancy; and differentially expressed in murine breast cancers induced by the Neu and c-myc oncogenes. Our studies suggest that Hunk and Punc may play a role in normal mammary development, may represent markers for biologically important subsets of epithelial cells in the breast, and may contribute to the process of mammary carcinogenesis. We hypothesize that Hunk and Punc play critical roles in mammary development by mediating pregnancy-induced changes in proliferation and differentiation. These hypotheses will be tested by using a tetracycline-inducible system to conditionally express Hunk and Punc in nontransformed mammary epithelial cells in vitro and in transgenic animals in vivo. Mice bearing targeted deletions of Hunk and Punc will be used to determine the impact of loss of function mutations in these kinases on mammary development, specifically with respect to changes in cellular proliferation, apoptosis, and programs of differentiation that occur during pregnancy. In this application we propose to investigate the role played by two novel protein kinases in normal mammary development as a step toward understanding the relationship between development and carcinogenesis in the breast. Beyond the further elucidation of this important relationship, we believe that these studies will yield an improved understanding of the regulation of mammary epithelial proliferation and differentiation during pregnancy, and will thereby help illuminate a stage of mammary development that contributes to the determination of breast cancer susceptibility.
