The xenobiotic inhibitor profile of cytochrome P4502C8.
OBJECTIVE
To investigate inhibition of recombinant CYP2C8 by: (i) prototypic CYP isoform selective inhibitors (ii) imidazole/triazole antifungal agents (known inhibitors of CYP), and (iii) certain CYP3A substrates (given the apparent overlapping substrate specificity of CYP2C8 and CYP3A).


METHODS
CYP2C8 and NADPH-cytochrome P450 oxidoreductase were coexpressed in Spodoptera frugiperda (Sf21) cells using the baculovirus expression system. CYP isoform selective inhibitors, imidazole/triazole antifungal agents and CYP3A substrates were screened for their inhibitory effects on CYP2C8-catalysed torsemide tolylmethylhydroxylation and, where appropriate, the kinetics of inhibition were characterized. The conversion of torsemide to its tolylmethylhydroxy metabolite was measured using an h.p.l.c. procedure.


RESULTS
At concentrations of the CYP inhibitor 'probes' employed for isoform selectivity, only diethyldithiocarbamate and ketoconazole inhibited CYP2C8 by > 10%. Ketoconazole, at an added concentration of 10 microM, inhibited CYP2C8 by 89%. Another imidazole, clotrimazole, also potently inhibited CYP2C8. Ketoconazole and clotrimazole were both noncompetitive inhibitors of CYP2C8 with apparent Ki values of 2.5 microM. The CYP3A substrates amitriptyline, quinine, terfenadine and triazolam caused near complete inhibition (82-91% of control activity) of CYP2C8 at concentrations five-fold higher than the known CYP3A Km. Kinetic studies with selected CYP3A substrates demonstrated that most inhibited CYP2C8 noncompetitively. Apparent Ki values for midazolam, quinine, terfenadine and triazolam ranged from 5 to 25 microM.


CONCLUSIONS
Inhibition of CYP2C8 occurred at concentrations of ketoconazole and diethyldithiocarbamate normally employed for selective inhibition of CYP3A and CYP2E1, respectively. Some CYP3A substrates have the capacity to inhibit CYP2C8 activity and this may have implications for inhibitory drug interactions in vivo.
Ong
C E
CE
Department of Clinical Pharmacology, Flinders Medical Centre and Flinders University, Adelaide, Australia.
Coulter
S
S
Birkett
D J
DJ
Bhasker
C R
CR
Miners
J O
JO
D000818:Animals; D000935:Antifungal Agents / Q000494:pharmacology; D001189:Aryl Hydrocarbon Hydroxylases; D001667:Binding, Competitive; D002478:Cells, Cultured; D065727:Cytochrome P-450 CYP2C8; D051544:Cytochrome P-450 CYP3A; D065607:Cytochrome P-450 Enzyme Inhibitors; D003577:Cytochrome P-450 Enzyme System / Q000378:metabolism; D004050:Ditiocarb / Q000494:pharmacology*; D004347:Drug Interactions; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D006801:Humans; D007654:Ketoconazole / Q000494:pharmacology*; D007700:Kinetics; D008862:Microsomes, Liver / Q000201:enzymology / Q000378:metabolism; D010089:Oxidoreductases, N-Demethylating / Q000037:antagonists & inhibitors / Q000378:metabolism; D011994:Recombinant Proteins / Q000037:antagonists & inhibitors; D018411:Spodoptera; D039181:Steroid 16-alpha-Hydroxylase; D013250:Steroid Hydroxylases / Q000037:antagonists & inhibitors* / Q000378:metabolism; D014162:Transfection
D000935:Antifungal Agents; D065607:Cytochrome P-450 Enzyme Inhibitors; D004791:Enzyme Inhibitors; D011994:Recombinant Proteins; D003577:Cytochrome P-450 Enzyme System; D004050:Ditiocarb; D013250:Steroid Hydroxylases; D001189:Aryl Hydrocarbon Hydroxylases; C450259:CYP2C8 protein, human; D065727:Cytochrome P-450 CYP2C8; D051544:Cytochrome P-450 CYP3A; D039181:Steroid 16-alpha-Hydroxylase; D010089:Oxidoreductases, N-Demethylating; D007654:Ketoconazole