The role of cytochrome P450s in the activation of weak and potent carcinogenic polycyclic aromatic hydrocarbons

Heidi Jane Einolf, Purdue University

Abstract

Cytochrome P450 (CYP) enzymes are required for both the detoxification and activation of carcinogenic polycyclic aromatic hydrocarbons (PAH). The induction of the CYP1A1 isoform results in activation of benzo (a) pyrene (B (a) P) to an ultimate carcinogenic metabolite, B (a) P-7S,8R-diol-9R,10S-epoxide. The environmental PAH, benzo (c) phenanthrene (B (c) Ph) is a weak carcinogen in mice, but synthetic B (c) Ph diol epoxides (B (c) PhDE) are potent carcinogens. The role of CYP1 isoforms in the metabolic activation of B (c) Ph was investigated in mouse skin and in human cell cultures. B (c) Ph is metabolically activated to B (c) PhDE in human mammary MCF-7 cells: the majority of DNA-adducts were from reaction of the most carcinogenic isomer, $(4R,3S)$-dihydroxy-$(2S,1R)$-epoxy-1,2,3,4-tetrahydro-B (c) Ph, with deoxyadenosine. In contrast, low levels of B (c) PhDE-DNA adducts were formed in mouse skin: most of these adducts were from the less carcinogenic isomer, $(4S,3R)$-dihydroxy-$(2S,1R)$-epoxy-1,2,3,4-tetrahydro-B (c) Ph. Pre-treatments with 2,3,7,8-tetrachlorodibenzo-p-dioxin demonstrated that B (c) PhDE-DNA adduct formation in mouse skin and MCF-7 cells was limited by low induction of CYP enzymes by B (c) Ph. Differences in amount and stereoselectivity of B (c) Ph activation in mouse skin and MCF-7 cells provide an explanation for the low carcinogenic activity of B (c) Ph in mouse skin and suggest that human cells may be able to activate this compound to ultimate carcinogenic metabolites. Antisera against B (c) PhDE-DNA adducts was found to detect PAHDE-DNA adducts similar in structure to B (c) PhDE-DNA, including benzo (g) chrysene, chrysene and some methylchrysene derivatives. The antisera should prove useful for detection of DNA adducts formed from this class of environmental PAH. Techniques for CYP1A1 antisense oligonucleotide inhibition were developed to examine the role of CYP1A1 induction on the metabolism of B (a) P in cultured cells. After extensive optimization of mRNA target sequence, cellular uptake, oligonucleotide stability and oligonucleotide treatment time and dose, inhibition of B (a) P metabolism by up to 40% was obtained. These studies demonstrated that induction of specific P450 isoforms by PAH is an important factor in carcinogenic PAH activation, especially in cases of PAH that are weak CYP inducers but potent carcinogens after activation.

Degree

Ph.D.

Advisors

Baird, Purdue University.

Subject Area

Biochemistry

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