Both 17β-estradiol-2,3-quinone (E₂-2,3-Q) and 17β-estradiol-3,4-quinone (E₂-3,4-Q) are reactive metabolites of estrogen that are thought to be responsible for the estrogen-induced genotoxicity. The aim of this study was to establish a methodology to analyze estrogen quinone-derived protein adducts and to measure the background levels of these adducts in human serum albumin (Alb) derived from female blood donors in Taiwan. Results from in vitro experiments confirmed that the production of estrogen quinone-derived adducts on serum Alb increased with increased concentration of estrogen quinones. Time-course experiments suggested that both E₂-2,3-Q- and E₂-3,4-Q-derived adducts rapidly reached maximum values at 10 min mark and remained constant thereafter for up to 24 h. Additionally, with 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) pretreatment, the production of estrogen quinone-derived protein adducts was detected in human MCF-7 breast cancer cells exposed to estrogen. Co-treatment of a catechol-O-methyl transferase inhibitor further enhanced the production of estrogen quinone-derived adducts in all cases. When we investigated the levels of estrogen quinone-derived adducts in human serum Alb, cysteinyl adducts of E₂-2,3-Q-1-S-Alb, E₂-2,3-Q-4-S-Alb, and E₂-3,4-Q-2-S-Alb were detected in all healthy female controls (n=10) with median levels at 147 (range 14.1-533), 197 (range 30.0-777), and 65.6 (range 17.6-1360) (pmol/g), respectively. We noticed that levels of E₂-2,3-Q-derived adducts were 2-fold greater than those of E₂-3,4-Q-2-S-Alb in controls whereas levels of E₂-3,4-Q-2-S-Alb were 2-fold higher than those of E₂-2,3-Q-derived adducts in patients (n = 20). Additionally, levels of E₂-2,3-Q-4-S-Alb correlated significantly with those of E₂-3,4-Q-2-S-Alb (correlation coefficient r = 0.684-0.850, p < 0.05). Overall, we conclude that cumulative body burden of E₂-3,4-Q is a significant predictor of breast cancer.
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