afimoxifene and cumene-hydroperoxide

One of our laboratories recently showed by 32P-postlabeling that administration of tamoxifen to mice induces two groups of hepatic DNA adducts comprising two major spots, nos. 3 and 5, respectively. 4-Hydroxytamoxifen and alpha-hydroxytamoxifen appear to be the proximate metabolites of groups I and II adducts, respectively. The relative significance of these two adduct groups for tamoxifen carcinogenicity remains to be established. To determine the activation mechanism(s) of tamoxifen and 4-hydroxytamoxifen, in vivo adducts were compared by 32P-postlabeling with adducts generated by microsomal or chemical activation in vitro. Microsomal activation of 4-hydroxytamoxifen and tamoxifen, respectively, in the presence of DNA and cumene hydroperoxide, induced two adducts, which mapped similarly to the corresponding in vivo adduct spots 3 and 5. Chemical oxidation of 4-hydroxytamoxifen with silver(II) oxide, followed by incubation of the product(s) with DNA, elicited the formation of a major spot (Q1), while tamoxifen itself did not react. Rechromatographic analyses revealed that in vitro fractions 3 and Q1 (from 4-hydroxytamoxifen) matched the major in vivo group I adduct fraction 3, consistent with the hypothesis that 4-hydroxytamoxifen is a precursor for adduct fraction 3 in vivo. The in vitro adduct fraction 5 (from tamoxifen) was identical to that formed in vivo, indicating that the metabolic pathway for the formation of group II adducts did not involve 4-hydroxytamoxifen. In conclusion, the results support a model where primary metabolites of tamoxifen undergo secondary metabolism to form DNA adducts, which are detected in vivo after treatment with tamoxifen or 4-hydroxytamoxifen.

Topics: Animals; Antineoplastic Agents, Hormonal; Benzene Derivatives; DNA; DNA Adducts; Female; Mice; Mice, Inbred ICR; Microsomes, Liver; Rats; Tamoxifen

Using rat liver microsomal preparations and peroxidase enzymes, we have investigated the formation of DNA adducts by the antiestrogen compound tamoxifen (TAM) and its metabolite 4-hydroxy-tamoxifen (4-OH-TAM). When reduced nicotinamide-adenine dinucleotide phosphate (NADPH) was used as a cofactor in microsomal activation of either 4-OH-TAM or TAM, one DNA adduct and relative DNA adduct levels of 4.6 and 3.1 x 10(-8), respectively were detected by 32P-postlabeling. The DNA adduct produced by microsomal activation of 4-OH-TAM and TAM was the same. With cumene hydroperoxide (CuOOH) as the cofactor for the microsomal activation of either 4-OH-TAM or TAM, three to six DNA adducts were produced; the relative adduct levels were 8.0 and 20.6 x 10(-8), respectively. Comparison of the DNA adduct patterns produced by 4-OH-TAM and TAM showed that they were distinct. However one of the DNA adducts (a) produced by microsomal activation of 4-OH-TAM using CuOOH was the same as adduct a produced by microsomal activation of 4-OH-TAM with NADPH. Activation of 4-OH-TAM with horseradish peroxidase resulted in the formation of a single DNA adduct and a relative adduct level of 20.7 x 10(-8). Rechromatography analysis of this DNA adduct showed that it was identical to that produced by microsomal activation of 4-OH-TAM with NADPH and one of the adducts produced using CuOOH as the cofactor. Ten DNA adducts and a relative adduct level of 15.3 x 10(-8) were detected in the liver of female Sprague-Dawley rats treated daily with 20 mg/kg of TAM for 7 days. The DNA adduct pattern in the liver of the treated animals was similar to that produced by microsomal activation of TAM using CuOOH as the co-factor. The principal DNA adduct (no. 6) formed in the livers of rats treated with TAM was the same as the principal DNA adduct formed following microsomal activation of TAM using CuOOH as a cofactor. The DNA adduct formed following microsomal activation of either TAM or 4-OH-TAM using NADPH was also present as one of the adducts (1) formed in vivo following TAM treatment. These studies demonstrate that 4-OH-TAM can be activated to form DNA adducts and that it contributes to the formation of DNA adducts in the liver of rats treated with TAM.

Topics: Animals; Benzene Derivatives; Biotransformation; DNA; DNA Adducts; Female; Horseradish Peroxidase; Microsomes, Liver; NADP; Phenobarbital; Rats; Rats, Sprague-Dawley; Tamoxifen