Formation of estrogen metabolites that react with DNA is thought to be a mechanism of cancer initiation by estrogens. The estrogens estrone (E(1)) and estradiol (E(2)) can form catechol estrogen (CE) metabolites, catechol estrogen quinones [E(1)(E(2))-3,4-Q], which react with DNA to form predominantly depurinating adducts. This may lead to mutations that initiate cancer. Catechol-O-methyltransferase (COMT) catalyzes an inactivation (protective) pathway for CE. This study investigated the effect of inhibiting COMT activity on the levels of depurinating 4-OHE(1)(E(2))-1-N3Ade and 4-OHE(1)(E(2))-1-N7Gua adducts in human breast epithelial cells. MCF-10F cells were treated with TCDD, a cytochrome P450 inducer, then with E(2) and Ro41-0960, a COMT inhibitor. Estrogen metabolites and depurinating DNA adducts in culture medium were analyzed by HPLC with electrochemical detection. Pre-treatment of cells with TCDD increased E(2) metabolism to 4-OHE(1)(E(2)) and 4-OCH(3)E(1)(E(2)). Inclusion of Ro41-0960 and E(2) in the medium blocked formation of methoxy CE, and depurinating adducts were observed. With Ro41-0960, more adducts were detected in MCF-10F cells exposed to 1 microM E(2), whereas without the inhibitor, no increases in adducts were detected with E(2) < or =10 microM. We conclude that low COMT activity and increased formation of depurinating adducts can be critical factors leading to initiation of breast cancer.