NITRITE-STIMULATED DNA-BINDING OF CARCINOGENIC DIOL EPOXIDES FROM BENZO[A]PYRENE-7,8-DIHYDRODIOL IN HUMAN POLYMORPHONUCLEAR LEUKOCYTES

Human polymorphonuclear leukocytes (PMNs) previously treated with 12-O -tetradecanoyl phorbolmyristate-13-acetate (PMA) to initiate the oxida tive burst activate (-)-trans-7,8-dihydroxy-7,8-dihydrobenzo[a]pyrene [(-)-BP-7,8-diol)] to DNA-binding intermediates. The P-32-postlabellin g technique and HPLC-analysis of enzyme-digested DNA were employed for identification of DNA-adducts following incubation of (-)-BP-7,8-diol in PMNs. The results are consistent with the formation of (+)-anti-BP DE, the ultimate carcinogen of BP, bound via trans-addition of the C-1 0 position in the diol epoxide molecule to the exocyclic nitrogen of d eoxyguanosine (BPDE-N-2-dG adduct). Addition of nitrite, the major aqu eous dissolution product of NO2, stimulated the formation of(+)-anti-B PDE and subsequent binding to both nuclear DNA in PMNs (about twofold) and to DNA present outside the cells (two- to fourfold). Preliminary experiments suggest that nitrite stimulates the metabolism of (-)-BP-7 ,8-diol by direct interaction with myeloperoxidase and hydrogen peroxi de. Consistent with previous work by us and others, the covalent bindi ng of (+)-anti-BPDE to extracellular targets demonstrate that these re active products, expected to be formed intracellularly, can be release d from the leukocytes. Measurement of hydroxyl radical-induced DNA dam age by estimating the formation of 8-hydroxydeoxyguanosine (8-OH-dG) i n resting PMNs revealed low amounts of adducts (1 adduct/10(6) dG-1 ad duct/10(5) dG). Pretreating the cells with PMA or PMA in conjunction w ith nitrite had no significant effect on 8-OH-dG adduct formation.