title: Electrospray ionization-tandem mass spectrometry and 32P-postlabeling analyses of tamoxifen-DNA adducts in humans.
abstract: 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 32P-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-N2-yl)-tamoxifen (dG-Tam) and (E)-alpha-(deoxyguanosin-N2-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 dG-desMeTam by (32)P-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 dG-desMeTam were two adducts per 10(9) nucleotides and two adducts per 10(8) nucleotides, respectively) or by 32P-postlabeling analyses (limit of detection for both adducts was one adduct per 10(9) nucleotides) in any of the endometrial DNA samples.
  
  
  CONCLUSIONS
  The initiation of endometrial cancer by tamoxifen is probably not due to a genotoxic mechanism involving the formation of dG-Tam or dG-desMeTam.
authors.affiliation: Division of Biochemical Toxicology, HFT-110, National Center for Toxicological Research, 3900 NCTR Rd., Jefferson, AR 72079, USA. fbeland@nctr.fda.gov
authors.forename: Frederick A
authors.initials: FA
authors.lastname: Beland
authors.forename: Mona I
authors.initials: MI
authors.lastname: Churchwell
authors.forename: Daniel R
authors.initials: DR
authors.lastname: Doerge
authors.forename: Daniel R
authors.initials: DR
authors.lastname: Parkin
authors.forename: Danuta
authors.initials: D
authors.lastname: Malejka-Giganti
authors.forename: Alan
authors.initials: A
authors.lastname: Hewer
authors.forename: David H
authors.initials: DH
authors.lastname: Phillips
authors.forename: Paul L
authors.initials: PL
authors.lastname: Carmichael
authors.forename: Gonçalo
authors.initials: G
authors.lastname: Gamboa da Costa
authors.forename: M Matilde
authors.initials: MM
authors.lastname: Marques
mesh_terms: D000328:Adult; D000368:Aged; D018931:Antineoplastic Agents, Hormonal / Q000008:administration & dosage / Q000009:adverse effects / Q000378:metabolism*; D002851:Chromatography, High Pressure Liquid; D018736:DNA Adducts / Q000032:analysis* / Q000302:isolation & purification; D016889:Endometrial Neoplasms / Q000139:chemically induced* / Q000000981:diagnostic imaging / Q000235:genetics; D020847:Estrogen Receptor Modulators / Q000008:administration & dosage / Q000009:adverse effects / Q000378:metabolism*; D005260:Female; D006801:Humans; D008875:Middle Aged; D010761:Phosphorus Radioisotopes; D011877:Radionuclide Imaging; D021241:Spectrometry, Mass, Electrospray Ionization; D013629:Tamoxifen / Q000008:administration & dosage / Q000009:adverse effects / Q000378:metabolism*
chemical_list: D018931:Antineoplastic Agents, Hormonal; D018736:DNA Adducts; D020847:Estrogen Receptor Modulators; D010761:Phosphorus Radioisotopes; D013629:Tamoxifen