B16

Estrogen exposure and metabolism are of current interest because they may play an important role in estrogen-sensitive cancers in postmenopausal women. We investigated the effects of long-term soy isoflavone (IF) consumption or premenopausal oral contraceptive (OC) use on postmenopausal expression of cytochrome P450 (CYP) mRNA, and whether IF directly affects CYP activity. One-hundred eighty-one female cynomolgus monkeys were randomized to receive OC or placebo for 28 months premenopausally, then ovariectomized and randomized to either an IF-free or IF-containing diet for 36 months. We measured mRNA levels of CYP1A1, CYP1B1, and CYP3A4 from total RNA isolated from liver and mammary samples obtained at necropsy using real-time RT-PCR. Long-term IF consumption did not significantly alter postmenopausal mammary CYP mRNA levels, while premenopausal OC administration reduced postmenopausal mRNA levels of all three enzymes assayed (50% reduction in both CYP1A1 and CYP1B1 mRNA and 80% reduction of CYP3A4 mRNA). Neither treatment changed any CYP mRNA level within the liver. To complement our studies in monkeys and increase our mechanistic understanding of IF effects, we also measured the effect of IF on CYP activity using a fluorogenic CYP activity assay in intact V79 cells stably transfected to express one of the three aforementioned CYP enzymes. Cells were preincubated with 1, 3, 10, or 30uM genistein, daidzein, or equol (a major metabolite of daidzein) added to the medium thirty minutes prior to addition of substrate for assay of activity. Overall, the activity of each CYP enzyme was altered in a dose-dependent and isoflavone-specific manner, with genistein exhibiting the most significant reduction of CYP1A1 and CYP1B1 activities. CYP1B1 activity was the enzyme most sensitive to IF pretreatment with each isoflavone achieving greater than 50% reduction in activity (approximate IC50 values of 2.1, 5.6, and 7.9uM for genistein, equol, and daidzein respectively). In summary, long-term mRNA expression of CYP isozymes involved in steroid hormone metabolism in the mammary tissues of postmenopausal non-human primates was reduced by prior OC exposure, but not by dietary IF consumption. However, our cell culture studies indicated that soy IF altered CYP activity acutely at concentrations achievable in the tissues of individuals on a diet including significant amounts of soy isoflavones. The strong reduction of CYP1B1 activity by IF could translate to reduced amounts of 4-OHE1, a precursor to mutagenic catecholestrogen quinones, produced in the tissue, thereby reducing the risk of estrogen-induced cancers. Finally, the reduction in CYP mRNA expression by OC exposure suggests that OC may actually attenuate the risk thought to be associated with metabolism of estrogens in this pathway.

Sixth AACR International Conference on Frontiers in Cancer Prevention Research-- Dec 5-8, 2007; Philadelphia, PA