Cytochromes P450 in Genetically Engineered Cell Cultures: The Gene Technological Approach

Doehmer, Johannes; Greim, Helmut

doi:10.1007/978-3-642-77763-9_27

Cited by 9 publications

(1 citation statement)

References 64 publications

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“…Understanding the role of each of these isoforms is part of a rationale-based risk assessment. Several cellular systems have been developed to study P450-mediated metabolism and me- tabolite-dependent toxic effects (7). V79 Chinese hamster cells had been selected to serve as host cells for heterologous expression of P450, because these cells possess several biological characteristics which make them a valuable mammalian test system for various toxicological end points (8).…”

Section: Introductionmentioning

confidence: 99%

Stable Expression of Human Cytochrome P450 1B1 in V79 Chinese Hamster Cells and Metabolically Catalyzed DNA Adduct Formation of Dibenzo[a,l]pyrene

Luch

Coffing

Tang

et al. 1998

Chem. Res. Toxicol.

117

111

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Chinese hamster V79 cell lines were constructed for stable expression of human cytochrome P450 1B1 (P450 1B1) in order to study its role in the metabolic activation of chemicals and toxicological consequences. The new V79 cell lines were applied to studies on DNA adduct formation of the polycyclic aromatic hydrocarbon (PAH) dibenzo[a,l]pyrene (DB[a,l]P). This compound has been found to be an environmental pollutant, and in rodent bioassays it is the most carcinogenic PAH yet discovered. Activation of DB[a,l]P in various metabolizing systems occurs via fjord region DB[a,l]P-11, 12-dihydrodiol 13,14-epoxides (DB[a,l]PDE): we found that DB[a,l]P is stereoselectively metabolized in human mammary carcinoma MCF-7 cells to the (-)-anti- and (+)-syn-DB[a,l]PDE which both bind extensively to cellular DNA. To follow up this study and to relate specific DNA adducts to activation by individual P450 isoforms, the newly established V79 cells stably expressing human P450 1B1 were compared with those expressing human P450 1A1. DNA adduct formation in both V79 cell lines differed distinctively after incubation with DB[a,l]P or its enantiomeric 11,12-dihydrodiols. Human P450 1A1 catalyzed the formation of DB[a,l]PDE-DNA adducts as well as several highly polar DNA adducts as yet unidentified. The proportion of these highly polar adducts to DB[a,l]PDE adducts was dependent upon both the concentration of DB[a,l]P and the time of exposure. In contrast, V79 cells stably expressing human P450 1B1 generated exclusively DB[a,l]PDE-DNA adducts. Differences in the total level of DNA binding were also observed. Exposure to 0.1 microM DB[a,l]P for 6 h caused a significantly higher level of DNA adducts in V79 cells stably expressing human P450 1B1 (370 pmol/mg of DNA) compared to those with human P450 1A1 (35 pmol/mg of DNA). A 4-fold higher extent of DNA binding was catalyzed by human P450 1B1 (506 pmol/mg of DNA) compared to human P450 1A1 (130 pmol/mg of DNA) 6 h after treatment with 0.05 microM (-)-(11R,12R)-dihydrodiol. In cells stably expressing human P450 1B1 the DNA adducts were derived exclusively from the (-)-anti-DB[a,l]PDE. These results indicate that human P450 1B1 and P450 1A1 differ in their regio- and stereochemical selectivity of activation of DB[a,l]P with P450 1B1 forming a higher proportion of the highly carcinogenic (-)-anti-(11R, 12S,13S,14R)-DB[a,l]PDE metabolite.

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Section: Introductionmentioning

confidence: 99%

Stable Expression of Human Cytochrome P450 1B1 in V79 Chinese Hamster Cells and Metabolically Catalyzed DNA Adduct Formation of Dibenzo[a,l]pyrene

Luch

Coffing

Tang

et al. 1998

Chem. Res. Toxicol.

117

111

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Cytochromes P450 and drug resistance

Doehmer¹,

Goeptar²,

Vermeulen³

1994

Multiple Drug Resistance in Cancer

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Cytochromes P450 and drug resistance

1993

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Cytochromes P450 are the key enzymes for activating and inactivating many drugs, in particular anticancer drugs. Therefore, individual expression levels of cytochromes P450 may play a crucial role in drug safety and drug efficacy. Overexpression of cytochrome P450 may yield rapid turnover and elimination of drugs before the target site was reached and any pharmacological effect is observed. Therefore, it may be vital to know the individual cytochrome P450 status in order to select the appropriate drug before drug resistance occurs. Expression levels and activity of cytochromes P450 depend on many different factors. These factors include tissue and organ specific expression, sex- and age-dependent expression, genetic differences yielding polymorphic forms, competitive inhibition or induction of cytochromes P450 due to multiple drug interaction, nutrition and diet. Genetically engineered test cells defined for cytochromes P450 are available for studying drugs for metabolic activation and for identifying the metabolically competent cytochrome P450 isoform.

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Cytochromes P450 in Genetically Engineered Cell Cultures: The Gene Technological Approach

Cited by 9 publications

References 64 publications

Stable Expression of Human Cytochrome P450 1B1 in V79 Chinese Hamster Cells and Metabolically Catalyzed DNA Adduct Formation of Dibenzo[a,l]pyrene

Stable Expression of Human Cytochrome P450 1B1 in V79 Chinese Hamster Cells and Metabolically Catalyzed DNA Adduct Formation of Dibenzo[a,l]pyrene

Cytochromes P450 and drug resistance

Cytochromes P450 and drug resistance

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