title: Effect of inhibitor time-dependency on selectivity towards cyclooxygenase isoforms.
abstract: Cyclooxygenase (Cox) is a key enzyme in the biosynthesis of prostaglandins and, as such, is the target of non-steroidal anti-inflammatory drugs (NSAIDs). Two isoforms exist, being expressed constitutively (Cox-1), or inducibly in response to inflammatory mediators (Cox-2). Currently available NSAIDs inhibit both isoforms somewhat equipotently but selective Cox-2 inhibition may eliminate unwanted side effects. We have characterized the kinetic mechanisms of the interactions of purified recombinant human cyclooxygenase-1 and -2 (hCox-1, hCox-2) with the selective Cox-2 inhibitor N-(2-cyclohexyloxy-4-nitrophenyl)methanesulphonamide (NS-398) and some classical non-selective NSAIDs. NS-398, flurbiprofen, meclofenamic acid and indomethacin are time-dependent, irreversible inhibitors of hCox-2. The inhibition is consistent with a two-step process, involving an initial rapid equilibrium binding of enzyme and inhibitor, characterized by Ki, followed by the slow formation of a tightly bound enzyme-inhibitor complex, characterized by a first-order rate constant kon. NS-398 is a time-independent inhibitor of hCox-1, consistent with the formation of a reversible enzyme-inhibitor complex. Flurbiprofen, meclofenamic acid and indomethacin are also time-dependent inhibitors of hCox-1 and hence show little selectivity for one isoform over the other. Flufenamic acid is time independent towards both isoforms and is also non-selective. The high degree of selectivity of NS-398 towards Cox-2 results therefore from the difference in the nature of the time-dependency of inhibition of the two isoforms.
authors.affiliation: Department of Biochemistry and Molecular Biology, Merck Frosst Centre for Therapeutic Research, Pointe-Claire-Dorval, Quebec, Canada.
authors.forename: M
authors.initials: M
authors.lastname: Ouellet
authors.forename: M D
authors.initials: MD
authors.lastname: Percival
mesh_terms: D016861:Cyclooxygenase Inhibitors / Q000378:metabolism / Q000494:pharmacology*; D004305:Dose-Response Relationship, Drug; D005480:Flurbiprofen / Q000378:metabolism / Q000494:pharmacology; D006801:Humans; D007213:Indomethacin / Q000378:metabolism / Q000494:pharmacology; D007527:Isoenzymes / Q000378:metabolism*; D007700:Kinetics; D008469:Meclofenamic Acid / Q000378:metabolism / Q000494:pharmacology; D009578:Nitrobenzenes / Q000008:administration & dosage / Q000378:metabolism / Q000494:pharmacology; D011451:Prostaglandin-Endoperoxide Synthases / Q000378:metabolism*; D011994:Recombinant Proteins; D013449:Sulfonamides / Q000008:administration & dosage / Q000378:metabolism / Q000494:pharmacology
chemical_list: D016861:Cyclooxygenase Inhibitors; D007527:Isoenzymes; D009578:Nitrobenzenes; D011994:Recombinant Proteins; D013449:Sulfonamides; C080955:N-(2-cyclohexyloxy-4-nitrophenyl)methanesulfonamide; D008469:Meclofenamic Acid; D005480:Flurbiprofen; D011451:Prostaglandin-Endoperoxide Synthases; D007213:Indomethacin