Formation and excision of covalent deoxyribonucleic acid adducts of benzo[a]pyrene 4,5-epoxide and benzo[a]pyrenediol epoxide I in human lung cells A549.

Abstract

The formation and repair of covalent DNA adducts induced by the two ultimate benzo(a)pyrene (BP) metabolites 4,5-dihydro-4,5-epoxybenzo(a)pyrene (BPE) and 7..beta..,8..cap alpha..-dihydroxy-9..cap alpha..,10..cap alpha..-epoxy-7,8,9,10-tetrahydrobenzo(a)-pyrene (BPDE I) were studied in human epithelioid lung cells A549. The covalent DNA adducts of BPE were characterized by reacting native DNA specifically labeled in a single deoxynucleoside with racemic BPE in vitro. From the chromatographic analysis of enzymatic hydrolysates of BPE-treated DNA, it was concluded that adducts to deoxyguanosine (BPE-dG) and deoxyadenosine (BPE-dA) were formed. No adducts to the pyrimidine nucleotides were detected. DNA adducts with the same chromatographic properties were formed upon treatment of intact A549 cells with BPE. Treatment of A549 cells with racemic BPDE I mostly produced (7R)-N/sup 2/-(7..beta..-8..cap alpha..,9..cap alpha..-trihydroxy-7,8,9,10-tetrahydrobenzo(a)pyren-10-yl)deoxyguanosine (BPDE I-dG). The efficiency of formation of covalent DNA adducts in intact A549 cells was much lower for BPE than for BPDE I. The initial concentration of all adducts was considerably higher in micrococcal nuclease sensitive DNA fractions of A549 chromatin. The time course of the excision from high molecular weight DNA of the covalent BPE and BPDE I adducts was determined from 0- to 35-h posttreatment incubation of A549 cells. The rates of excision decreased according to the sequence BPE-dA >more » BPE-dG > BPDE I-dG. While the removal of BPE-dA was carried to completion, approximately 20% of BPE-dG and BPDE I-dG persisted in the DNA when the excision process had come to a halt or slowed down considerably. The low efficiency of formation and the high excisability of BPE adducts in comparison to BPDE I adducts may explain the lower cytotoxicity of BPE relative to BPDE I for A549 cells.« less