Accumulating evidence suggests that specific metabolites of estrogens, namely, catechol estrogen quinones, react with DNA to form adducts and generate apurinic sites, which can lead to the mutations that induce breast cancer. Oxidation of estradiol (E2) produces 2 catechol estrogens, 4-hydroxyestradiol (4-OHE2) and 2-OHE2 among the major metabolites. These, in turn, are oxidized to the quinones, E2-3,4-quinone (E2-3,4-Q) and E2-2,3-Q, which can react with DNA. Oxidation of E2 to 2-OHE2 is mainly catalyzed by cytochrome P450 (CYP) 1A1, and CYP3A4, whereas oxidation of E2 to 4-OHE2 in extrahepatic tissues is mainly catalyzed by CYP1B1 as well as some CYP3As. The potential involvement of CYP isoforms in the further oxidation of catechols to semiquinones and quinones has, however, not been investigated in detail. In this project, to identify the potential function of various CYPs in oxidizing catechol estrogens to quinones, we used different recombinant human CYP isoforms, namely, CYP1A1, CYP1B1, and CYP3A4, with the scope of oxidizing the catechol estrogens 2-OHE2 and 4-OHE2 to their respective estrogen quinones, which then reacted with DNA. The depurinating adducts 2-OHE2-6-N3Ade, 4-OHE2-1-N3Ade, and 4-OHE2-1-N7Gua were observed in the respective reaction systems by ultraperformance liquid chromatography/tandem mass spectrometry. Furthermore, more than 100-fold higher levels of estrogen-glutathione (GSH) conjugates were detected in the reactions. Glutathione conjugates were observed, in much smaller amounts, when control microsomes were used. Depurinating adducts, as well as GSH conjugates, were obtained when E2-3,4-Q was incubated with CYP1B1 or control microsomes in a 30-minute reaction, further demonstrating that GSH is present in these recombinant enzyme preparations. These experiments demonstrated that CYP1A1, CYP1B1, and CYP3A4 are able to oxidize catechol estrogens to their respective quinones, which can further react with GSH, protein, and DNA, the last resulting in depurinating adducts that can lead to mutagenesis.
ASJC Scopus subject areas
- Endocrinology, Diabetes and Metabolism