Document Detail

Inhibition of cytochrome P450 3A4 by a pyrimidineimidazole: Evidence for complex heme interactions.
MedLine Citation:
PMID:  17173379     Owner:  NLM     Status:  MEDLINE    
PH-302 inhibits the inducible form of nitric oxide synthase (iNOS) by coordinating with the heme of the monomeric form and preventing formation of the active dimer. Inherent with the mechanism of pharmacology for this compound was the inhibition of cytochrome P450 3A4 (P450 3A4), observed from early ADME screening. Further investigation showed that PH-302 inhibited P450 3A4 competitively with a Ki of approximately 2.0 microM against both midazolam and testosterone hydroxylation in human liver microsomes. As expected, spectral binding analysis demonstrated that inhibition was a result of type II coordination to the P450 heme with the imidazole moiety of PH-302, although only 72% of the maximal absorbance difference was achievable with PH-302 compared to that of the smaller ligand imidazole. Time-dependent inhibition of P450 3A4 by PH-302 was also observed because of metabolite-inhibitory (MI) complex formation via metabolism of the methylenedioxyphenyl group. The profile for time-dependent inhibition in recombinant P450 3A4 was biphasic, and was kinetically characterized by a kinact of 0.08 min-1 and a Ki of 1.2 microM for the first phase (0-1.5 min) and a kinact of 0.06 min-1 and a Ki of 23.8 microM for the second phase (1.5-10 min). Spectral characterization of the PH-302 MI complex demonstrated that formation began to plateau within 3 min, consistent with the kinetic profile of inactivation by PH-302. Meanwhile, spectral evidence for the imidazole-derived type II difference spectrum of PH-302 was captured simultaneously with the formation of the MI complex. The presence of simultaneously operable type II coordination and rapidly saturable MI complex formation with heme by PH-302 indicates the presence of complex heme interactions with this unique molecule. Information from these mechanistic studies adds to our understanding of the nature of P450 3A4 inhibition by PH-302 and provides a potentially useful tool compound for future studies investigating binding interactions in this important drug-metabolizing enzyme.
J Matthew Hutzler; Roger J Melton; Jeanne M Rumsey; Mark E Schnute; Charles W Locuson; Larry C Wienkers
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Publication Detail:
Type:  In Vitro; Journal Article    
Journal Detail:
Title:  Chemical research in toxicology     Volume:  19     ISSN:  0893-228X     ISO Abbreviation:  Chem. Res. Toxicol.     Publication Date:  2006 Dec 
Date Detail:
Created Date:  2006-12-18     Completed Date:  2007-03-09     Revised Date:  2008-11-21    
Medline Journal Info:
Nlm Unique ID:  8807448     Medline TA:  Chem Res Toxicol     Country:  United States    
Other Details:
Languages:  eng     Pagination:  1650-9     Citation Subset:  IM    
Pharmacokinetics, Dynamics and Metabolism (PDM), Department of Medicinal Chemistry, Pfizer Global Research and Development, 700 Chesterfield Parkway West T3A, Chesterfield, Missouri 63017, USA.
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MeSH Terms
Benzodioxoles / chemistry,  metabolism,  pharmacology*
Chromatography, Liquid
Cytochrome P-450 CYP3A
Cytochrome P-450 Enzyme System / antagonists & inhibitors*
Drug Interactions
Enzyme Inhibitors / chemistry,  metabolism,  pharmacology*
Heme / metabolism*
Microsomes, Liver / drug effects*,  enzymology
Molecular Structure
Protein Binding
Pyrimidines / chemistry,  metabolism,  pharmacology*
Tandem Mass Spectrometry
Reg. No./Substance:
0/Benzodioxoles; 0/Enzyme Inhibitors; 0/PH 302; 0/Pyrimidines; 14875-96-8/Heme; 9035-51-2/Cytochrome P-450 Enzyme System; EC protein, human; EC P-450 CYP3A

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