title: Site-specific tamoxifen-DNA adduct formation: lack of correlation with mutational ability in Escherichia coli.
abstract: We have mapped sites of tamoxifen adduct formation, in the lacI gene using the polymerase STOP assay, following reaction in vitro with alpha-acetoxytamoxifen and horseradish peroxidase (HRP)/H(2)O(2) activated 4-hydroxytamoxifen. For both compounds, most adduct formation occurred on guanines. However, one adenine, within a run of guanines, generated a strong polymerase STOP site with activated 4-hydroxytamoxifen, and a weaker STOP site with alpha-acetoxytamoxifen at the same location. In Escherichia coli the lac I gene reacted with 4-hydroxytamoxifen was more likely to be mutated (2 orders of magnitude) than when reacted with alpha-acetoxytamoxifen, despite the greater DNA adduct formation by alpha-acetoxytamoxifen. This correlates with the greater predicted ability of activated 4-hydroxytamoxifen adducts to disrupt DNA structure than alpha-acetoxytamoxifen adducts. For lac I reacted with activated 4-hydroxytamoxifen, a hot spot of base mutation was located in the region of the only adenosine adduct. No mutational hot spots were observed with alpha-acetoxytamoxifen. Our data clearly shows a lack of correlation between gross adduct number, as assayed by (32)P-postlabeling and mutagenic potential. These data indicate the importance of minor adduct formation in mutagenic potential and further that conclusions regarding the mutagenicity of a chemical may not be reliably derived from the gross determination of adduct formation.
authors.affiliation: Medical Research Council Toxicology Unit, University of Leicester, United Kingdom.
authors.forename: D A
authors.initials: DA
authors.lastname: Lowes
authors.forename: K
authors.initials: K
authors.lastname: Brown
authors.forename: R T
authors.initials: RT
authors.lastname: Heydon
authors.forename: E A
authors.initials: EA
authors.lastname: Martin
authors.forename: T W
authors.initials: TW
authors.lastname: Gant
mesh_terms: D017122:Bacteriophage T4 / Q000201:enzymology; D001483:Base Sequence; D018736:DNA Adducts / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D004252:DNA Mutational Analysis / Q000379:methods; D004256:DNA Polymerase I; D004259:DNA-Directed DNA Polymerase; D004926:Escherichia coli / Q000235:genetics*; D007763:Lac Operon; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D009153:Mutagens / Q000737:chemistry / Q000378:metabolism*; D010761:Phosphorus Radioisotopes / Q000378:metabolism; D013629:Tamoxifen / Q000031:analogs & derivatives / Q000737:chemistry / Q000378:metabolism*; D014764:Viral Proteins
chemical_list: D018736:DNA Adducts; D009153:Mutagens; D010761:Phosphorus Radioisotopes; D014764:Viral Proteins; C105080:alpha-acetoxytamoxifen; C069983:gene 43 protein, Enterobacteria phage T4; D013629:Tamoxifen; C016601:afimoxifene; D004256:DNA Polymerase I; D004259:DNA-Directed DNA Polymerase