The Major Metabolite of Equilin, 4-Hydroxyequilin, Autoxidizes to an o-Quinone Which Isomerizes to the Potent Cytotoxin 4-Hydroxyequilenin-o-quinone

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• 作者：Fagen Zhang ; Yumei Chen ; Emily Pisha ; Li Shen ; Yansan Xiong ; Richard B. van Breemen ; and Judy L. Bolton
• 刊名：Chemical Research in Toxicology
• 出版年：1999
• 出版时间：February 1999
• 年：1999
• 卷：12
• 期：2
• 页码：204 - 213
• 全文大小：197K
• 年卷期：v.12,no.2(February 1999)
• ISSN：1520-5010

文摘

The risk factors for women developing breast and endometrial cancers are all associatedwith a lifetime of estrogen exposure. Estrogen replacement therapy in particular has beencorrelated with a slight increased cancer risk. Previously, we showed that equilenin, a minorcomponent of Premarin (Wyeth-Ayerst), was metabolized to highly cytotoxic quinoids whichcaused oxidative stress and alkylation of DNA in vitro [Bolton, J. L., Pisha, E., Zhang, F., andQiu, S. (1998) Chem. Res. Toxicol. 11, 1113-1127]. In this study, we have compared thechemistry of the major catechol metabolite of equilin (4-hydroxyequilin), which is found inseveral estrogen replacement formulations, to the equilenin catechol (4-hydroxyequilenin).Unlike endogenous catechol estrogens, both equilin and equilenin were primarily convertedby rat liver microsomes to 4-hydroxylated rather than 2-hydroxylated o-quinone GSHconjugates. With equilin, a small amount of 2-hydroxyequilin GSH quinoids were detected(4-hydroxyequilin:2-hydroxyequilin ratio of 6:1); however, no peaks corresponding to 2-hydroxyequilenin were observed in incubations with equilenin. These data suggest thatunsaturation in the B ring alters the regiochemistry of P450-catalyzed hydroxylation fromprimarily 2-hydroxylation for endogenous estrogens to 4-hydroxylation for equine estrogens.4-Hydroxyequilenin-o-quinone reacts with GSH to give two mono-GSH conjugates and onedi-adduct. The behavior of 4-hydroxyequilin was found to be more complex than 4-hydroxyequilenin as conjugates resulting from 4-hydroxyequilenin were detected in addition to the4-hydroxyequilin-GSH adducts. The mechanism of decomposition of 4-hydroxyequilin likelyinvolves isomerization to a quinone methide which readily aromatizes to 4-hydroxyequileninfollowed by autoxidation to 4-hydroxyequilenin-o-quinone. Similar results were obtained with2-hydroxyequilin, although, in contrast to 4-hydroxyequilenin, 2-hydroxyequilenin does notautoxidize and the reaction stops at the catechol. Since 4-hydroxyequilin is converted to4-hydroxyequilenin and 4-hydroxyequilenin-o-quinone, similar effects were observed for thisequine catechol, including consumption of NAD(P)H likely by the 4-hydroxyequilenin-o-quinone,depletion of molecular oxygen by 4-hydroxyequilenin or its semiquinone radical, and alkylationof deoxynucleosides and DNA by 4-hydroxyequilenin quinoids. Finally, preliminary studiesconducted with the human breast tumor cell line MCF-7 demonstrated that the cytotoxic effectsof the catechol estrogens from estrone, equilin, and 2-hydroxyequilenin were similar, whereas4-hydroxyequilenin was a much more potent cytotoxin (~30-fold). These results suggest thatthe catechol metabolites of equine estrogens have the ability to cause alkylation/redox damagein vivo primarily through formation of 4-hydroxyequilenin quinoids.