The dietary isoflavones genistein, daidzein, formononetin, and biochanin A are derived from precursors primarily found in soybean products, bean sprouts and other legumes, and whole-grain cereals. It is postulated that the soy-based diets in Asian populations may contribute to the lower incidences of breast, colorectal, and prostate cancers in this group as compared to Western populations. The isoflavones are belived to play a significant inhibitory role during the promotional stage of the disease, and recent evidence suggests their role in the initiation stage of carcinogenesis. In addition, biotransformation of the isoflavones is believed to be necessary for some actions. Cytochromes P450 are a class of enzymes that metabolize a wide variety of compounds and are found in nearly every tissue. Several drug-drug and food-drug interactions have been attributed to the interaction of compounds at the active site of cytochrome P450. Many times, metabolism by P450 results in products that are excreted from the body, but metabolism can also result in reactive compounds that interact with cellular macromolecules with eventual detrimental effects including initiation of carcinogenesis. The isoflavones may bind at the active site of P450 and act as low Km substrates while inhibiting the activation of procarcinogens. The goal of this research is to investigate the metabolism of isoflavones by cytochromes P450 using rat and human liver microsomes, heterologously-expressed human cytochromes P450, primary cultured rat hepatocytes, and breast MCF-7, MCF-10A, and MCF-10AT cell lines. The metabolites will be characterized by HPLC, GC/MS, UV-Vis and fluorescence spectroscopy, and ESI/MS/MS. The enzymatically-generated metabolites will be compared with metabolites that are chemically synthesized. The long term goal is to determine which human cytochromes P450 metabolize these isoflavones, and identify and characterize the metabolites from subcellular fractions and cell cultures.
