Benzo(a)pyrene:DNA adduct formation in normal human mammary epithelial cell cultures and the human mammary carcinoma T47D cell line.
The benzo(a)pyrene (BaP):DNA adducts formed in normal human mammary epithelial cell cultures and the human mammary carcinoma T47D cell line were analyzed by chromatography and acid hydrolysis of the BaP:deoxyribonucleoside adducts to BaP:purine adducts and BaP:tetraols. Human mammary epithelial cell cultures and human mammary carcinoma T47D cells were exposed to [3H]BaP for 24 h, and the levels of binding were 81 and 182 pmol BaP/mg DNA in normal and T47D cultures, respectively. Analysis of BaP:deoxyribonucleoside adducts resolved by immobilized boronate chromatography and reversephase high-performance liquid chromatography demonstrated the presence of three BaP:deoxyribonucleoside adducts in both cells: M2, MS1, and MS2 in a ratio of 1.6:1:14. Two adducts (MS1 and MS2) bound to the immobilized boronate column indicating the presence of cis-vicinal hydroxyl groups, a configuration which would result from reaction of 7 beta, 8 alpha-dihydroxy-9 alpha, 10 alpha-epoxy-7,8,9,10-tetrahydroBaP (anti-BaPDE) with DNA. MS2 was identified as (+)-anti-BaPDE:deoxyguanosine (dGuo) for it cochromatographed with a [14C]-(+)-anti-BaPDE:dGuo marker, the BaP:purine hydrolysis product of MS2 cochromatographed with [14C]-(+)-anti-BaPDE:guanine, and the tetraol hydrolysis products cochromatographed with (+/-)-anti-BaPDE:tetraols. MS1 was identified as (-)-anti-BaPDE:dGuo for MS1 eluted in the same relative position as a (-)-anti-BaPDE:dGuo marker, the BaP:purine hydrolysis product of MS1 cochromatographed with [14C]-(+)-anti-BaPDE:guanine, and the tetraol hydrolysis products cochromatographed with (+/-)-anti-BaPDE:tetraols. Thus, both adducts that bound to the immobilized boronate column were formed from (+/-)-anti-BaPDE. One major adduct that did not contain cis-vicinal hydroxy groups, M2, was detected in both cell types. M2 was formed from (+/-)-7 beta, 8 alpha-dihydroxy-9 beta, 10 beta-epoxy-7,8,9,10-tetrahydroBaP (syn-BaPDE) as M2 eluted in the same relative position as a syn-BaPDE:dGuo adduct marker and the tetraol hydrolysis products of M2 cochromatographed with tetraols formed from (+/-)-syn-BaPDE. The isolation of the individual BaP:DNA adducts followed by acid hydrolysis allowed the identification of the BaP:DNA adducts formed in human mammary cell cultures and demonstrated the presence of (-)-anti-BaPDE:dGuo. Thus, this work provides the first evidence, other than cochromatography, that (-)-anti-BaPDE is formed in cell systems and reacts with DNA in cells to form (-)-anti-BaPDE:dGuo.(ABSTRACT TRUNCATED AT 400 WORDS)
Pruess-Schwartz
D
D
Baird
W M
WM
Nikbakht
A
A
Merrick
B A
BA
Selkirk
J K
JK
D015123:7,8-Dihydro-7,8-dihydroxybenzo(a)pyrene 9,10-oxide; D001564:Benzo(a)pyrene / Q000378:metabolism*; D001580:Benzopyrenes / Q000378:metabolism; D001940:Breast / Q000378:metabolism*; D001943:Breast Neoplasms / Q000378:metabolism*; D002277:Carcinoma / Q000378:metabolism*; D002460:Cell Line; D002478:Cells, Cultured; D002851:Chromatography, High Pressure Liquid; D004247:DNA / Q000378:metabolism*; D004273:DNA, Neoplasm / Q000378:metabolism; D003849:Deoxyguanosine / Q000378:metabolism; D004848:Epithelium / Q000378:metabolism; D005260:Female; D006801:Humans; D006868:Hydrolysis; D014316:Tritium
D001580:Benzopyrenes; D004273:DNA, Neoplasm; D014316:Tritium; D001564:Benzo(a)pyrene; D015123:7,8-Dihydro-7,8-dihydroxybenzo(a)pyrene 9,10-oxide; D004247:DNA; D003849:Deoxyguanosine