title: Interaction of sulfaphenazole derivatives with human liver cytochromes P450 2C: molecular origin of the specific inhibitory effects of sulfaphenazole on CYP 2C9 and consequences for the substrate binding site topology of CYP 2C9.
abstract: The effects of sulfaphenazole, 1, on typical activities catalyzed by human cytochromes P450 of the 1A, 3A, and 2C subfamilies expressed in yeast were studied. 1 acts as a strong, competitive inhibitor of CYP 2C9 (K(i) = 0.3 +/- 0.1 microM); it is much less potent toward CYP 2C8 and 2C18 (K(i) = 63 and 29 microM, respectively) and fails to inhibit CYP 1A1, 1A2, 3A4, and 2C19. From difference visible spectroscopy experiments using microsomes of yeast expressing various human P450s, 1 selectively interacts only with CYP 2C9 with the appearance of a peak at 429 nm as expected for the formation of a P450 Fe(III)-nitrogenous ligand complex (Ks = 0.4 +/- 0.1 microM). Comparative studies of the spectral interaction and inhibitory effects of twelve compounds related to 1 with CYP 2C9 showed that the aniline function of 1 is responsible for the formation of the iron-nitrogen bond of the 429 nm-absorbing complex and is necessary for the inhibitory effects of 1. The study of two new compounds synthesized during this work, in which the N-phenyl group of 1 was replaced with either an ethyl group or a 3,4-dichlorophenyl group, showed that the presence of an hydrophobic substituent at position 1 of the pyrazole function of 1 is required for a strong interaction with CYP 2C9. A model for the binding of 1 in the CYP 2C9 active site is proposed; that takes into account three major interactions that should be at the origin of the high-affinity and specific inhibitory effects of 1 toward CYP 2C9: (i) the binding of its nitrogen atom to CYP 2C9 iron, (ii) an ionic interaction of its SO2N- anionic site with a cationic residue of CYP 2C9, and (iii) an interaction of its N-phenyl group with an hydrophobic part of the protein active site.
authors.affiliation: Laboratoire de Chimie et Biochimie Pharmacologiques et Toxicologiques, URA 400 CNRS, Université Paris V, France.
authors.forename: A
authors.initials: A
authors.lastname: Mancy
authors.forename: S
authors.initials: S
authors.lastname: Dijols
authors.forename: S
authors.initials: S
authors.lastname: Poli
authors.forename: P
authors.initials: P
authors.lastname: Guengerich
authors.forename: D
authors.initials: D
authors.lastname: Mansuy
mesh_terms: D001189:Aryl Hydrocarbon Hydroxylases; D001665:Binding Sites; D003001:Cloning, Molecular; D065607:Cytochrome P-450 Enzyme Inhibitors; D003577:Cytochrome P-450 Enzyme System / Q000737:chemistry* / Q000378:metabolism*; D006801:Humans; D007700:Kinetics; D008099:Liver / Q000201:enzymology*; D008861:Microsomes / Q000201:enzymology; D008958:Models, Molecular; D015394:Molecular Structure; D011487:Protein Conformation; D011994:Recombinant Proteins / Q000037:antagonists & inhibitors / Q000737:chemistry / Q000378:metabolism; D012441:Saccharomyces cerevisiae; D013053:Spectrophotometry; D039181:Steroid 16-alpha-Hydroxylase; D013250:Steroid Hydroxylases / Q000037:antagonists & inhibitors / Q000737:chemistry* / Q000378:metabolism*; D013329:Structure-Activity Relationship; D013379:Substrate Specificity; D013426:Sulfaphenazole / Q000031:analogs & derivatives* / Q000138:chemical synthesis / Q000378:metabolism / Q000494:pharmacology*
chemical_list: D065607:Cytochrome P-450 Enzyme Inhibitors; D011994:Recombinant Proteins; D013426:Sulfaphenazole; D003577:Cytochrome P-450 Enzyme System; D013250:Steroid Hydroxylases; D001189:Aryl Hydrocarbon Hydroxylases; D039181:Steroid 16-alpha-Hydroxylase