Identification of Tamoxifen−DNA Adducts Formed by α-Sulfate Tamoxifen and α-Acetoxytamoxifen

Abstract

α-Sulfate trans-tamoxifen and α-sulfate cis-tamoxifen were synthesized as proposed active metabolites of tamoxifen that react with DNA. α-Acetoxytamoxifen was prepared as a model-activated form to produce a reactive carbocation. Calf thymus DNA was reacted with α-hydroxytamoxifen or the activated forms of tamoxifen, and tamoxifen−DNA adducts were analyzed by a ³²P-postlabeling method. The reactivity of α-sulfate trans-tamoxifen to DNA was much higher than that of α-hydroxytamoxifen. The formation of tamoxifen−DNA adducts induced by α-acetoxytamoxifen and α-sulfate cis-tamoxifen was 1100- and 1600-fold, respectively, higher than that of α-hydroxytamoxifen. Both α-sulfate tamoxifens and α-acetoxytamoxifen were highly reactive to 2‘-deoxyguanosine. Four reaction products of dG-tamoxifen were isolated by HPLC and characterized by mass- and proton magnetic resonance spectroscopy. Fractions 1 and 2 that eluted first were identified as the epimers of trans form of dG-N²-tamoxifen. Fractions 3 and 4 were identified as the epimers of cis form of dG-N²-tamoxifen. When DNA was reacted with α-acetoxytamoxifen in vitro, three isomers of dG-N²-tamoxifen were detected: fraction 2 was the major adduct while fractions 1 and 3 were minor adducts.