DNA INTERACTIONS OF CARCINOGENIC POLYCYCLIC AROMATIC HYDROCARBONS (BENZO(A)PYRENE, BENZO(C)PHENANTHRENE)
Abstract
The binding of polycyclic aromatic hydrocarbons (PAH) to DNA is one of the initial steps essential in cancer induction by PAH; therefore, it is essential to be able to analyze the specific hydrocarbon adducts present in DNA. The benzo(a)pyrene(BaP)-deoxyribonucleoside adducts formed in rodent embryo cell cultures were analyzed by immobilized boronate chromatography and reverse-phase HPLC to investigate the mechanism of activation of BaP to DNA-binding metabolites. Three BaP-deoxyribonucleoside adducts were present in Wistar rat embryo cells that did not contain cis-vicinal hydroxyl groups (M1, M2, M3) and three adducts were present that did (MS1, MS2, MS3). Cochromatography with known standards suggested that M2 and M3 were syn-BaPDE(7(beta),8(alpha)-dihydroxy-9(beta),10(beta)-epoxy-7,8,9,10-tetrahydroBaP)-deoxycytidine or deoxyguanosine (dG) adducts and MS1 and MS2 were (-)-anti-BaPDE(7(beta),8(alpha)-dihydroxy-9(alpha),10(alpha)-epoxy-7,8,9,10-tetrahydroBaP)-dG and (+)-anti-BaPDE, respectively. Acid hydrolysis of the BaP-DNA adducts containing cis-vicinal hydroxyl groups present in Wistar rat embryo cell cultures to BaP-purine adducts and BaP-tetraols demonstrated that only one adduct was formed from ((+OR-))-anti-BaPDE: MS2 was identified as (+)-anti-BaPDE-dG, MS1 was shown to be formed from a reactive intermediate other than BaPDE, and MS3 was characterized as a syn-BaPDE-deoxyribonucleoside breakdown product. Thus, BaP can undergo multiple pathways of metabolic activation to DNA binding intermediates in rodent embryo cells. The BaP-DNA adducts formed in human mammary epithelial and human mammary carcinoma T 47D cell cultures were analyzed by the same techniques to investigate the activation of BaP in human mammary cell cultures. Both (+)-anti-BaPDE-dG and (-)-anti-BaPDE-dG were present providing the first evidence that (-)-anti-BaPDE is formed in cells. The DNA adducts formed from benzo(c)phenanthrene (BcPh) in rodent embryo cell cultures resulted exclusively from two of the four possible isomers of BcPh-3,4-diol-1,2-epoxide, demonstrating a high degree of stereochemical specificity in the metabolic activation of BcPh. The immobilized boronate chromatography and acid hydrolysis techniques described greatly improved the analysis of PAH-DNA adducts. These studies have helped to elucidate the mechanism of activation of PAH in human and rodent embryo cell cultures.
Degree
Ph.D.
Subject Area
Biochemistry
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