The effects of steroidal estrogens in ACI rat mammary carcinogenesis: 17beta-estradiol, 2-hydroxyestradiol, 4-hydroxyestradiol, 16alpha-hydroxyestradiol, and 4-hydroxyestrone.

Several investigators have suggested that certain hydroxylated metabolites of 17beta-estradiol (E2) are the proximate carcinogens that induce mammary carcinomas in estrogen-sensitive rodent models. The studies reported here were designed to examine the carcinogenic potential of different levels of E2 and the effects of genotoxic metabolites of E2 in an in vivo model sensitive to E2-induced mammary cancer. The potential induction of mammary tumors was determined in female ACI rats subcutaneously implanted with cholesterol pellets containing E2 (1, 2, or 3 mg), or 2-hydroxyestradiol (2-OH E2), 4-hydroxyestradiol (4-OH E2), 16alpha-hydroxyestradiol (16alpha-OH E2), or 4-hydoxyestrone (4-OH E1) (equimolar to 2 mg E2). Treatment with 1, 2, or 3 mg E2 resulted in the first appearance of a mammary tumor between 12 and 17 weeks, and a 50% incidence of mammary tumors was observed at 36, 19, and 18 weeks respectively. The final cumulative mammary tumor incidence in rats treated with 1, 2, or 3 mg E2 for 36 weeks was 50%, 73%, and 100% respectively. Treatment of rats with pellets containing 2-OH E2, 4-OH E2, 16alpha-OH E2, or 4-OH E1 did not induce any detectable mammary tumors. The serum levels of E2 in rats treated with a 1 or 3 mg E2 pellet for 12 weeks was increased 2- to 6-fold above control values (approximately 30 pg/ml). Treatment of rats with E2 enhanced the hepatic microsomal metabolism of E2 to E1, but did not influence the 2- or 4-hydroxylation of E2). In summary, we observed a dose-dependent induction of mammary tumors in female ACI rats treated continuously with E2; however, under these conditions 2-OH E2, 4-OH E2, 16alpha-OH E2, and 4-OH E1 were inactive in inducing mammary tumors.

Clotrimazole is a selective and potent inhibitor of rat cytochrome P450 3A subfamily-related testosterone metabolism.

In this study, clotrimazole (CTZ) and ketoconazole (KTZ) were evaluated for their inhibition of testosterone metabolism catalyzed by rat hepatic microsomes differentially expressing certain cytochrome P450 enzymes. The objective was to compare the inhibitory potencies using hepatic microsomes from adult female rats treated with dexamethasone (F-DEX) and hepatic microsomes from vehicle-treated adult male rats (M-VEH), which are known to contain high levels of isozymes CYP3A1 (3A23) and 3A2, respectively. The results demonstrate that CTZ is a very potent and selective inhibitor of the 6beta-hydroxylation of testosterone, a CYP3A-mediated reaction, in all rat metabolic systems tested. The IC(50) value was 9.7 nM in F-DEX, and 6.7 nM in M-VEH for CTZ. The in vitro inhibitory potency for CTZ significantly exceeds the same parameters for KTZ, a well established specific inhibitor of human CYP3A-mediated reactions. It was found that the IC(50) values of KTZ in F-DEX and M-VEH were 69 and 780 nM, respectively. These values for KTZ are 10-fold and 100-fold higher, respectively, than for CTZ. CTZ, at the concentration that inhibits 90% and more of CYP3A-mediated reactions (40 nM), has less than a 10% inhibitory effect on the activities of other rat liver enzymes, such as CYP1A1, -1A2, -2A1, -2B1, -2B2, -2C11, and -2E1. In summary, CTZ is a more potent and selective inhibitor of all CYP3A-mediated reactions than KTZ in rat hepatic microsomes.