Sequence-specific DNA damage induced by carcinogenic danthron and anthraquinone in the presence of Cu(II), cytochrome P450 reductase and NADPH

The mechanism of metal-mediated DNA damage by carcinogenic danthron (1,8-di hydroxyanthraquinone) and anthraquinone was investigated by the DNA sequenc ing technique using P-32-labeled human DNA fragments obtained from the huma n c-Ha-ras-1 protooncogene and the p53 tumor suppressor gene. Danthron caus ed DNA damage particularly at guanines in the 5'-GC-3'., 5'-GGGG-3', 5'-GGG GG-3' sequences (damaged-bases are underlined) in the presence of Cu(II), c ytochrome P450 reductase and the NADPH-generating system. The DNA damage wa s inhibited by catalase and bathocuproine, suggesting the involvement of H2 O2 and Cu(I). The formation of 8-oxo-7,8-dihydro-2'-deoxyguanosine increase d with increasing concentration of danthron. On the other hand, carcinogeni c anthraquinone induced less oxidative DNA damage than danthron. Electron s pin resonance study showed that the semiquinone radical could be produced b y P450 reductase plus NADPH-mediated reduction of danthron, while little si gnal was observed with anthraquinone. These results suggest that danthron i s much more likely to be reduced by P450 reductase and generate reactive ox ygen species through the redox cycle, leading to more extensive Cu(II)-medi ated DNA damage than anthraquinone. In the case of anthraquinone, its hydro xylated metabolites with similar reactivity to danthron may participate in DNA damage in vivo. We conclude that oxidative DNA damage by danthron and a nthraquinone seems to be relevant for the expression of their carcinogenici ty.