Electrospray ionization-tandem mass spectrometry and 32P-postlabeling analyses of tamoxifen-DNA adducts in humans

Background: Although the nonsteroidal antiestrogen tamoxifen is used as an adjuvant chemotherapeutic agent to treat hormone-dependent breast cancer and as a chemopreventive agent in women with elevated risk of breast cancer, it has also been reported to increase the risk of endometrial cancer. Reports of low levels of tamoxifen-DNA adducts in human endometrial tissue have suggested that tamoxifen induces endometrial cancer by a genotoxic mechanism. However, these findings have been controversial. We used electrospray ionization-tandem mass spectrometry (ES-MS/MS) and (32)p- postlabeling analyses to investigate the presence of tamoxifen-DNA adducts in human endometrial tissue. Methods: Endometrial DNA from eight tamoxifen-treated women and eight untreated women was hydrolyzed to nucleosides and assayed for (E)-alpha-(deoxyguanosin-N-2-yl)-tamoxifen (dG-Tam) and (E)-alpha-(deoxyguanosin-N-2-yl)-N-desmethyltamoxifen (dG-desMeTam), the two major tamoxifen-DNA adducts that have been reported to be present in humans and/or experimental animals treated with tamoxifen, using on-line sample preparation coupled with high-performance liquid chromatography (HPLC) and ES-MS/MS. The same DNA samples were assayed for the presence of dG-Tam and dGdesMeTam by P-32-postlabeling methodology, using two different DNA digestion and labeling protocols, followed by both thin-layer chromatography and HPLC. Results: We did not detect either tamoxifen-DNA adduct by HPLC-ES-MS/MS analyses (limits of detection for dG-Tam and dGdesMeTam were two adducts per 10(9) nucleotides and two adducts per 10(8) nucleotides, respectively) or by P-32- postlabeling analyses (limit of detection for both adducts was one adduct per 109 nucleotides) in any of the endometrial DNA samples. Conclusion: The initiation of endometrial cancer by tamoxifen is probably not due to a genotoxic mechanism involving the formation of dG-Tam or dG-desMeTam.