Gierse J K, Koboldt C M, Walker M C, Seibert K, Isakson P C
Monsanto/Searle Discovery Research, 700 Chesterfield Parkway North, Chesterfield, MO 63198, USA.
Biochem J. 1999 May 1;339 ( Pt 3)(Pt 3):607-14.
Non-steroidal anti-inflammatory drugs (NSAIDs) inhibit the formation of prostaglandins by cyclo-oxygenases (COX). The discovery of a second COX isoform (COX-2) associated with inflammation led to agents that selectively inhibit COX-2, e.g. celecoxib. We evaluated the kinetics of inhibition of celecoxib and several NSAIDs. Celecoxib displays classic competitive kinetics on COX-1 (Ki=10-16 microM). An initial competitive interaction with COX-2 can also be discerned with celecoxib (Ki=11-15 microM), followed by a time-dependent interaction leading to potent inhibition, characterized as inactivation (Kinact=0.03-0.5 s-1). Half-maximal inhibition (IC50) using end-point assays reflects the competitive component on COX-1 (IC50=4-19 microM) and the inactivation component on COX-2 (IC50=0.003-0.006 microM). NSAIDs exhibit four distinct modes of COX inhibition based on kinetic behaviour: (1) competitive, e.g. ibuprofen; (2) weak binding, time-dependent, e.g. naproxen, oxicams; (3) tight binding, time-dependent, e.g. indomethacin; (4) covalent, e.g. aspirin. In addition, most NSAIDs display different kinetic behaviour for each isoform. Weakly binding inhibitors show variable behaviour in enzyme assays, with apparent inhibitory activity being markedly influenced by experimental conditions; determination of kinetic constants with this class is unreliable and IC50 values are strongly dependent on assay conditions. Although IC50 determinations are useful for structure/activity analyses, the complex and distinct mechanisms of enzyme inhibition of each COX isoform by the NSAIDs renders comparison of inhibitory activity on COX-1 and COX-2 using IC50 ratios of questionable validity.
非甾体抗炎药（NSAIDs）通过环氧化酶（COX）抑制前列腺素的形成。与炎症相关的第二种COX同工型（COX-2）的发现催生了选择性抑制COX-2的药物，如塞来昔布。我们评估了塞来昔布和几种NSAIDs的抑制动力学。塞来昔布对COX-1表现出典型的竞争性动力学（Ki = 10 - 16 microM）。塞来昔布与COX-2最初也可识别出竞争性相互作用（Ki = 11 - 15 microM），随后是时间依赖性相互作用，导致强效抑制，其特征为失活（Kinact = 0.03 - 0.5 s-1）。使用终点测定法的半数最大抑制（IC50）反映了COX-1上的竞争性成分（IC50 = 4 - 19 microM）和COX-2上的失活成分（IC50 = 0.003 - 0.006 microM）。基于动力学行为，NSAIDs表现出四种不同的COX抑制模式：（1）竞争性，如布洛芬；（2）弱结合、时间依赖性，如萘普生、昔康类；（3）紧密结合、时间依赖性，如吲哚美辛；（4）共价性，如阿司匹林。此外，大多数NSAIDs对每种同工型表现出不同的动力学行为。弱结合抑制剂在酶测定中表现出可变行为，表观抑制活性受实验条件显著影响；用这类抑制剂测定动力学常数不可靠，IC50值强烈依赖于测定条件。尽管IC50测定对结构/活性分析有用，但NSAIDs对每种COX同工型的酶抑制机制复杂且不同，使得使用IC50比值比较对COX-1和COX-2的抑制活性的有效性存疑。
