Copper-mediated oxidative DNA damage induced by eugenol: possible involvement of O-demethylation.
Eugenol used as a flavor has potential carcinogenicity. DNA adduct formation via 2,3-epoxidation pathway has been thought to be a major mechanism of DNA damage by carcinogenic allylbenzene analogs including eugenol. We examined whether eugenol can induce oxidative DNA damage in the presence of cytochrome P450 using [32P]-5'-end-labeled DNA fragments obtained from human genes relevant to cancer. Eugenol induced Cu(II)-mediated DNA damage in the presence of cytochrome P450 (CYP)1A1, 1A2, 2C9, 2D6, or 2E1. CYP2D6 mediated eugenol-dependent DNA damage most efficiently. Piperidine and formamidopyrimidine-DNA glycosylase treatment induced cleavage sites mainly at T and G residues of the 5'-TG-3' sequence, respectively. Interestingly, CYP2D6-treated eugenol strongly damaged C and G of the 5'-ACG-3' sequence complementary to codon 273 of the p53 gene. These results suggest that CYP2D6-treated eugenol can cause double base lesions. DNA damage was inhibited by both catalase and bathocuproine, suggesting that H2O2 and Cu(I) are involved. These results suggest that Cu(I)-hydroperoxo complex is primary reactive species causing DNA damage. Formation of 8-oxo-7,8-dihydro-2'-deoxyguanosine was significantly increased by CYP2D6-treated eugenol in the presence of Cu(II). Time-of-flight-mass spectrometry demonstrated that CYP2D6 catalyzed O-demethylation of eugenol to produce hydroxychavicol, capable of causing DNA damage. Therefore, it is concluded that eugenol may express carcinogenicity through oxidative DNA damage by its metabolite.
Sakano
Katsuhisa
K
Department of Environmental and Molecular Medicine, Mie University School of Medicine, Mie 514-8507, Japan.
Inagaki
Yuji
Y
Oikawa
Shinji
S
Hiraku
Yusuke
Y
Kawanishi
Shosuke
S
D000080242:8-Hydroxy-2'-Deoxyguanosine; D000818:Animals; D002614:Chelating Agents / Q000378:metabolism; D003300:Copper / Q000378:metabolism*; D019941:Cyclin-Dependent Kinase Inhibitor p16 / Q000235:genetics; D019389:Cytochrome P-450 CYP2D6 / Q000378:metabolism*; D004249:DNA Damage; D019175:DNA Methylation; D003849:Deoxyguanosine / Q000031:analogs & derivatives* / Q000378:metabolism; D005054:Eugenol / Q000737:chemistry / Q000378:metabolism*; D016166:Free Radical Scavengers / Q000378:metabolism; D006801:Humans; D007527:Isoenzymes / Q000378:metabolism*; D015394:Molecular Structure; D010084:Oxidation-Reduction; D016159:Tumor Suppressor Protein p53 / Q000235:genetics
D002614:Chelating Agents; D019941:Cyclin-Dependent Kinase Inhibitor p16; D016166:Free Radical Scavengers; D007527:Isoenzymes; D016159:Tumor Suppressor Protein p53; D005054:Eugenol; D003300:Copper; D000080242:8-Hydroxy-2'-Deoxyguanosine; D019389:Cytochrome P-450 CYP2D6; D003849:Deoxyguanosine