Site-specific tamoxifen-DNA adduct formation: lack of correlation with mutational ability in Escherichia coli.
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.
Lowes
D A
DA
Medical Research Council Toxicology Unit, University of Leicester, United Kingdom.
Brown
K
K
Heydon
R T
RT
Martin
E A
EA
Gant
T W
TW
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
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