The objective of the research is to continue to use the estrogen-induced carcinomas in hamster kidney and liver as experimental models to elucidate the mechanisms whereby estrogens, both natural and synthetic, ellicit tumorigenesis in the hamster. We have concluded that in addition to their hormonal properties, estrogen possess carcinogenic properties which can be partially dissociated from the former. To elucidate more precisely the role estrogens play in causing these carcinomas, the following studies are proposed. 1) At present, the most plausible reactive intermediates which might be involved in neoplastic changes in hamster kidney are the arene oxides and o-quinones of the catechol pathway. To test this hypothesis, 2- and 4- hydroxyestrogens will be synthesized and their carcinogenic activities assessed in the hamster kidney. U-0521, an inhibitor of COMT will be used concomitantly with DES or 17Beta-estradiol in an effort to potentiate the carcinogenic effects of these estrogens and to discern the significance of the proposed catechol pathway in estrogen carcinogenesis. Modified steroidal estrogens (deoxoestrone, 11Beta-methoxy, 11Beta-methy and 11Beta-ethyl) will be prepared to further elucidate the significance of the catechol pathway in renal tumorigenesis. 2) We propose to attempt to indirectly identify the presence of arene oxide formation by observing an NIH-shift with the aid of epoxide hydrolase inhibitor, TCPO, and employing the estrogen hydroxylase assay. 3) We will continue our assessment of estrogen-2/-4 hydroxylase activity and formation of catechol estrogens, particularly in regard to 2 hydroxyestrogen formation using HPLC analyses. COMT and demethylase activities will also be assessed and their substrate specificity determined. These latter enzymes will provide more precise information regarding the in-vivo concentration of catechol estrogens in these tissues. 4) Prevention of estrogen-induced renal tumorigenesis in the hamster by androgens, progestins, or antiestrogens may be mediated in part by nonhormonal mechanisms by inhibiting catechol estrogen formation and hence arene oxide and oquinone amounts. This hypothesis will be tested employing estrogen hydroxylase assay with hormonally treated microsomes. 5) The P-450 MMM will be examined in detail in the untreated kidney and liver, and nonhormonal probes (Beta-NF, AlphaNF and 3-MC) will be used to perturb the P-450 MMM activities to determine alterations which may occur in estrogen metabolism. 6) The nature of the induction of liver tumors with Alpha-NF/estrogen treatment will be studied to rigorously identity the metabolites formed by [14C]-17Beta-E2 and DES in hamster liver and kidney.
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