Cytochrome P450 3A activities and their modulation by α-naphthoflavone in vitro are dictated by the efficiencies of model experimental systems

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Title:Cytochrome P450 3A activities and their modulation by α-naphthoflavone in vitro are dictated by the efficiencies of model experimental systems
Creators:
Bořek-Dohalská, Lucie
Stiborová, Marie
Journal or Publication Title:
Collection of Czechoslovak Chemical Communications, 75, 2, pp. 201-220
Uncontrolled Keywords:α-Naphthoflavone, Cooperativity, Testosterone, Enzymatic oxidation, Metabolism, Cytochrome P450 3A

Abstract

The knowledge on efficiencies of different <i>in vitro</i> systems containing cytochromes P450 (CYP) of a 3A subfamily is crucial to screen potential substrates of these CYPs. We evaluated and compared efficiencies of several <i>in vitro</i> CYP3A enzymatic systems to oxidize the model substrates, α-NF and testosterone, under the standardized experimental conditions. Five CYP3A systems were tested: (i) human hepatic microsomes rich in CYP3A4, (ii) hepatic microsomes of rabbits treated with a CYP3A6 inducer, rifampicine, (iii) microsomes of Baculovirus transfected insect cells containing recombinant human CYP3A4 and NADPH:CYP reductase with or without cytochrome b<sub>5</sub> (Supersomes<sup>TM</sup>), (iv) membranes isolated from <i>Escherichia coli</i>, containing recombinant human CYP3A4, NADPH:CYP reductase and cytochrome b<sub>5</sub>, and (v) human CYP3A4 or rabbit CYP3A6 reconstituted with NADPH:CYP reductase with or without cytochrome b<sub>5</sub> in liposomes. All systems oxidize testosterone to its 6β-hydroxylated metabolite and α-NF to trans-7,8-dihydrodiol and 5,6-epoxide. The most efficient systems oxidizing both compounds were CYP3A4-Supersomes<sup>TM</sup> containing cytochrome b<sub>5</sub>, followed by human hepatic microsomes. This finding suggests these systems to be suitable for general evaluating a variety of compounds as potential substrates of CYP3A4. The lowest efficiencies to oxidize α-NF and testosterone were found for CYP3A4 expressed in membranes of <i>E. coli</i>, and for reconstituted CYP3A4 or CYP3A6. Utilizing the tested enzymatic systems, we also explain here the discrepancies, which showed previously the controversial effects of α-NF on CYP3A-mediated reactions. We demonstrate that inhibition or stimulation of the CYP3A-mediated testosterone hydroxylation by α-NF is dictated by efficiencies of individual enzymatic systems to oxidize the CYP3A substrates.

Title:Cytochrome P450 3A activities and their modulation by α-naphthoflavone in vitro are dictated by the efficiencies of model experimental systems
Creators:
Bořek-Dohalská, Lucie
Stiborová, Marie
Uncontrolled Keywords:α-Naphthoflavone, Cooperativity, Testosterone, Enzymatic oxidation, Metabolism, Cytochrome P450 3A
Divisions:Life and Chemical Sciences > Institute of Organic Chemistry and Biochemistry > Collection of Czechoslovak Chemical Communications
Journal or Publication Title:Collection of Czechoslovak Chemical Communications
Volume:75
Number:2
Page Range:pp. 201-220
E-ISSN:1212-6950
Publisher:Institute of Organic Chemistry and Biochemistry
Related URLs:
URLURL Type
http://dx.doi.org/10.1135/cccc2009525UNSPECIFIED
ID Code:5238
Item Type:Article
Deposited On:08 Mar 2010 12:31
Last Modified:08 Mar 2010 11:32

Citation

Bořek-Dohalská, Lucie; Stiborová, Marie (2010) Cytochrome P450 3A activities and their modulation by α-naphthoflavone in vitro are dictated by the efficiencies of model experimental systems. Collection of Czechoslovak Chemical Communications, 75 (2). pp. 201-220.

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