Journal Article

The level of DNA modification by (+)-<i>syn-</i>(11<i>S</i>,12<i>R</i>,13<i>S,</i>14<i>R)-</i> and (–)-<i>anti-(</i>11<i>R</i>,12<i>S</i>,13<i>S</i>,14<i>R</i>)-dihydrodiol epoxides of dibenzo[<i>a</i>,<i>l</i>]pyrene determined the effect on the proteins p53 and p21<sup>WAF1</sup> in the human mammary carcinoma cell line MCF-7

Andreas Luch, Kim Kudla, Albrecht Seidel, Johannes Doehmer, Helmut Greim and William M. Baird

in Carcinogenesis

Volume 20, issue 5, pages 859-865
Published in print May 1999 | ISSN: 0143-3334
Published online May 1999 | e-ISSN: 1460-2180 | DOI: http://dx.doi.org/10.1093/carcin/20.5.859
The level of DNA modification by (+)-syn-(11S,12R,13S,14R)- and (–)-anti-(11R,12S,13S,14R)-dihydrodiol epoxides of dibenzo[a,l]pyrene determined the effect on the proteins p53 and p21WAF1 in the human mammary carcinoma cell line MCF-7

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The polycyclic aromatic hydrocarbon (PAH) dibenzo[a,l]pyrene (DB[a,l]P), the most carcinogenic PAH tested in rodent bioassays, exerts its pathobiological activity via metabolic formation of electrophilically reactive DNAbinding fjord region (+)-syn-(11S,12R,13S,14R)- or (–)-anti-(11R,12S,13S,14R)-DB[a,l]P-dihydrodiol epoxides (DB[a,l]PDEs). DB[a,l]P is metabolized to these DB[a,l]PDEs which bind to DNA in human mammary carcinoma MCF-7 cells. The molecular response of MCF-7 cells to DNA damage caused by DB[a,l]PDEs was investigated by analyzing effects on the expression of the tumor suppressor protein p53 and one of its target gene products, the cyclin-dependent kinase inhibitor p21WAF1. Treatment of MCF-7 cells with (+)-syn- and (–)-anti-DB[a,l]PDE at a concentration range of 0.001–0.1 μM resulted in DB[a,l]PDE–DNA adduct levels between 2 and 30, and 3 and 80 pmol/mg DNA, respectively, 8 h after exposure. (–)-anti-DB[a,l]PDE exhibited a higher binding efficiency that correlated with a significantly stronger p53 response at low concentrations of the dihydrodiol epoxides. The level of p53 increased by 6–8 h after treatment. The p21WAF1 protein amount exceeded control levels by 12 h and remained elevated for 96 h. At a dose of 0.01 μM (+)-syn-DB[a,l]PDE, an increase in p21WAF1 was observed in the absence of a detectable change in p53 levels. The results indicate that the increase in p53 induced by DB[a,l]PDEs in MCF-7 cells requires an adduct level of ~15 pmol/mg DNA and suggest that the level of adducts rather than the specific structure of the DB[a,l]PDE–DNA adduct formed triggers the p53 response. The PAH–DNA adduct level formed may determine whether p53 and p21WAF1 pathways respond, resulting in cell-cycle arrest, or fail to respond and increase the risk of mutation induction by these DNA lesions.

Keywords: (–)-anti-DB[a,l]PDE, dibenzo[a,l]pyrene-11R,12S-dihydrodiol 13S,14R-epoxide; (+)-anti-B[a]PDE, B[a]P-7R,8S-dihydrodiol 9S,10R-epoxide; (+)-syn-DB[a,l]PDE, dibenzo[a,l]pyrene-11S,12R-dihydrodiol 13S,14R-epoxide; B[a]P, benzo[a]pyrene; CDK, cyclin-dependent kinase; DB[a,l]P, dibenzo[a,l]pyrene; DB[a,l]PDE(s), DB[a,l]P-11,12-dihydrodiol 13,14-epoxide(s); DMSO, dimethyl sulfoxide; FCS, fetal calf serum; PAGE, polyacrylamide gel electrophoresis; PAH(s), polycyclic aromatic hydrocarbon(s); PBS, phosphate-buffered saline; PMSF, phenylmethylsulfonyl fluoride; SDS, sodium dodecyl sulfate; TBS, Tris-buffered saline; TCPI, trypsin–chymotrypsin protease inhibitor.

Journal Article.  5428 words.  Illustrated.

Subjects: Clinical Cytogenetics and Molecular Genetics

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