Metabolic Activation and Formation of DNA Adducts of Hexestrol, a Synthetic Nonsteroidal Carcinogenic Estrogen

Abstract

Hexestrol (HES), a synthetic nonsteroidal estrogen, is carcinogenic in Syrian golden hamsters. The major metabolite of HES is its catechol, 3‘-OH-HES, which can be metabolically converted to the electrophilic catechol quinone, HES-3‘,4‘-Q, by peroxidases and cytochrome P450. Standard adducts were synthesized by reacting HES-3‘,4‘-Q with dG and dA to produce the adducts 3‘-OH-HES-6‘(α,β)-N7Gua and HES-3‘,4‘-Q-6‘-N⁶dA, respectively. When HES-3‘,4‘-Q was reacted with calf thymus DNA, 3‘-OH-HES-6‘(α,β)-N7Gua was identified by HPLC and tandem mass spectrometry as the depurinating adduct, with minor amounts of stable adducts. 3‘-OH-HES was bound to DNA after activation by horseradish peroxidase, lactoperoxidase, or rat liver microsomes. The depurinating adduct 3‘-OH-HES-6‘(α,β)-N7Gua was identified in these systems at levels of 65, 41, and 11 μmol/mol of DNA-P, respectively. Unidentified stable adducts were observed in much lower amounts and were quantified by the ³²P-postlabeling method. Similarly to 3‘-OH-HES, the catechol metabolites of the natural steroidal estrogens estrone (E₁) and estradiol (E₂), namely, 2-OHE₁, 4-OHE₁, 2-OHE₂, and 4-OHE₂, can be oxidized to their corresponding quinones by peroxidases and cytochrome P450. The quinones of the carcinogenic 4-OHE₁ and 4-OHE₂ have chemical and biochemical properties similar to those of HES-3‘,4‘-Q. The results suggest that formation of HES-3‘,4‘-Q may be a critical event in tumor initiation by HES and that HES is an excellent model compound to corroborate the hypothesis that estrogen-3,4-quinones are ultimate carcinogenic metabolites of the natural steroidal estrogens E₁ and E₂.