Because growth of breast cancer is regulated by several hormones and growth factors, it is important to define the role that each growth-regulatory substance plays and how they interact in this multihormonal disease. Rodent tumors provide laboratory models to experimentally establish and define such roles as they relate to neoplastic growth. This research proposal will investigate insulin-like growth factors (IGFs) and the 6 different IGF binding proteins (IGFBPs), which can prevent or enhance the actions of IGFs. Subsequent studies to elucidate the mechanisms whereby IGFs and their IGFBPs affect tumor growth should provide the basis for exploiting this knowledge as potential therapeutic interventions. Initially, the goal of this project is to ascertain whether there are coordinate changes in tumor growth and the IGF-IGFR-IGFBP axis. Two experimental rodent mammary tumors will be used: primary, hormone-dependent carcinomas induced by 7,12-dimethylbenz(a)anthracene (DMBA) and the transplantable autonomous, hormone-responsive R3230AC adenocarcinoma. Tumor growth will be modified by selected hormonal perturbations, e.g., ovariectomy, induction of diabetes, estrogenic, progestagenic, insulin or an anti-estrogenic treatment, all known to affect growth of these neoplasms. Measurement of IGF receptors and IGFBPs in whole tumors, in primary cultured cells and in subcellular preparations will document changes preceding or concurrent with tumor growth. We will study the effect of estrogens and antiestrogens on IGFBP production in MCF-7 mammary tumors in vivo (as xenografts in nude mice) since this will provide data on a well studied estrogen and insulin-dependent human tumor cell line for comparison with the results from the rodent tumors. In situ hybridization, Northern and Western blot analysis, immunohistochemistry and run-on assays provide state-of-the-art technology to ascertain transcriptional and translational changes of the IGF axis. Mechanistic investigation of regulation of receptor function will then be undertaken by cell transfection assays. The ability of insulin and IGF-1 to alter transcriptional activation of an estrogen responsive CAT reporter gene will be determined. These experiments will be performed by transfecting primary cultures of cells dissociated from R3230AC mammary adenocarcinoma and DMBA- induced tumors as well as MCF-7 breast tumor cells with a ptkCAT plasmid containing an ERE consensus sequence. This new information on the regulation of growth of mammary tumors will provide insight into innovative therapeutic maneuvers to improve tumor treatment response.
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