Oxidative DNA Damage Induced by Activation of Polychlorinated Biphenyls (PCBs): Implications for PCB-Induced Oxidative Stress in Breast Cancer

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Abstract
We have previously reported that mono- and dichlorinated biphenyls (PCBs) can be metabolized to dihydroxy compounds and further oxidized to reactive metabolites which form adducts with nitrogen and sulfur nucleophiles including DNA [Amaro et al. (1996) Chem. Res. Toxicol. 9, 623−629; Oakley et al. (1996) Carcinogenesis 17, 109−114]. The former studies also demonstrated that during the metabolism of PCBs superoxide may be produced. We have therefore examined the abilities of PCB metabolites to induce free radical-mediated oxidative DNA damage using a newly developed, highly sensitive, 32P-postlabeling assay for 8-oxodeoxyguanosine (8-oxodG) [Devanaboyina, U., and Gupta, R. (1996) Carcinogenesis 17, 917−924]. The incubation of 3,4-dichloro-2‘,5‘-dihydroxybiphenyl (100 μM) with calf thymus DNA (300 μg/mL) in the presence of the breast tissue and milk-associated enzyme, lactoperoxidase (10 mU/mL), and H2O2 (0.5 mM) resulted in a significant increase in free radical-induced DNA damage (253 8-oxodG/106 nucleotides) as compared to vehicle-treated DNA (118 8-oxodG/106 nucleotides). Substituting CuCl2 (100 μM) for lactoperoxidase/H2O2, however, resulted in a substantial increase in 8-oxodG content (2669 8-oxodG/106 nucleotides). FeCl3 was ineffective, suggesting that CuCl2 but not FeCl3 mediates oxidation of PCB dihydroxy metabolites, resulting in oxidative DNA damage. The addition of catalase (100 U/mL) and sodium azide (0.1 M) reduced the effect of CuCl2 (849 and 896 8-oxodG/106 nucleotides, respectively), while superoxide dismutase (600 U/mL) moderately stimulated and glutathione (100 μM) substantially stimulated 8-oxodG formation (3014 and 4415 8-oxodG/106 nucleotides, respectively). The effect of various buffers as well as the effects of PCB structure on Cu(II)-mediated oxidative DNA damage were examined. These results demonstrate that free radicals and oxidative DNA damage are produced during oxidation of lower chlorinated biphenyls. The relevance of the results is discussed in view of the recent report that increased oxidative DNA base damage is detected in the DNA of human breast tumor tissue.