title: Competitive inhibition of coumarin 7-hydroxylation by pilocarpine and its interaction with mouse CYP 2A5 and human CYP 2A6.
abstract: 1. We have shown earlier that pilocarpine strongly inhibits mouse and human liver coumarin 7-hydroxylase activity of CYP 2A and pentoxyresorufin O-deethylase activity of CYP 2B in vitro. Since pilocarpine, like coumarin, contains a lactone structure we have studied in more detail its inhibitory potency on mouse and human liver coumarin 7-hydroxylation. 2. Pilocarpine was a competitive inhibitor of coumarin 7-hydroxylase in vitro both in mouse and human liver microsomes although it was not a substrate for CYP 2A5. Ki values were similar, 0.52 +/- 0.22 microM in mice and 1.21 +/- 0.51 microM in human liver microsomes. 3. Pilocarpine induced a type II difference spectrum in mouse, human and recombinant CYP 2A5 yeast cell microsomes, with Ka values of 3.7 +/- 1.6, 1.6 +/- 1.1 and 1.5 +/- 0.1 microM, respectively. 4. Increase in pH of the incubation medium from pH 6 to 7.5 increased the potency of inhibition of coumarin 7-hydroxylation by pilocarpine. 5. Superimposition of pilocarpine and coumarin in such a way that their carbonyls, ring oxygens and the H-7' of coumarin and N-3 of pilocarpine overlap yielded a common molecular volume of 82%. 6. The results indicate that pilocarpine is a competitive inhibitor and has a high affinity for mouse CYP 2A5 and human CYP 2A6. In addition the immunotype nitrogen of pilocarpine is coordinated towards the haem iron in these P450s.
authors.affiliation: Department of Pharmacology and Toxicology, University of Kuopio, Finland.
authors.forename: T
authors.initials: T
authors.lastname: Kinonen
authors.forename: M
authors.initials: M
authors.lastname: Pasanen
authors.forename: J
authors.initials: J
authors.lastname: Gynther
authors.forename: A
authors.initials: A
authors.lastname: Poso
authors.forename: T
authors.initials: T
authors.lastname: Järvinen
authors.forename: E
authors.initials: E
authors.lastname: Alhava
authors.forename: R O
authors.initials: RO
authors.lastname: Juvonen
mesh_terms: D000818:Animals; D001189:Aryl Hydrocarbon Hydroxylases; D001665:Binding Sites; D001667:Binding, Competitive; D003374:Coumarins / Q000378:metabolism*; D065637:Cytochrome P-450 CYP2A6; D065607:Cytochrome P-450 Enzyme Inhibitors; D003577:Cytochrome P-450 Enzyme System / Q000378:metabolism; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D005260:Female; D006801:Humans; D006900:Hydroxylation / Q000187:drug effects; D007700:Kinetics; D008297:Male; D051379:Mice; D008811:Mice, Inbred DBA; D008862:Microsomes, Liver / Q000201:enzymology; D006899:Mixed Function Oxygenases / Q000037:antagonists & inhibitors* / Q000378:metabolism; D008956:Models, Chemical; D010277:Parasympathomimetics / Q000378:metabolism / Q000494:pharmacology*; D010862:Pilocarpine / Q000378:metabolism / Q000494:pharmacology*
chemical_list: D003374:Coumarins; D065607:Cytochrome P-450 Enzyme Inhibitors; D004791:Enzyme Inhibitors; D010277:Parasympathomimetics; D010862:Pilocarpine; D003577:Cytochrome P-450 Enzyme System; C030123:coumarin; D006899:Mixed Function Oxygenases; D001189:Aryl Hydrocarbon Hydroxylases; D065637:Cytochrome P-450 CYP2A6