Department of Clinical Pharmacology, School of Medicine, Flinders University, Adelaide, Australia.
Drug Metab Dispos. 2011 Apr;39(4):644-52. doi: 10.1124/dmd.110.037036. Epub 2011 Jan 18.
Enzyme selective inhibitors represent the most valuable experimental tool for reaction phenotyping. However, only a limited number of UDP-glucuronosyltransferase (UGT) enzyme-selective inhibitors have been identified to date. This study characterized the UGT enzyme selectivity of niflumic acid (NFA). It was demonstrated that 2.5 μM NFA is a highly selective inhibitor of recombinant and human liver microsomal UGT1A9 activity. Higher NFA concentrations (50-100 μM) inhibited UGT1A1 and UGT2B15 but had little effect on the activities of UGT1A3, UGT1A4, UGT1A6, UGT2B4, UGT2B7, and UGT2B17. NFA inhibited 4-methylumbelliferone and propofol (PRO) glucuronidation by recombinant UGT1A9 and PRO glucuronidation by human liver microsomes (HLM) according to a mixed (competitive-noncompetitive) mechanism, with K(i) values ranging from 0.10 to 0.40 μM. Likewise, NFA was a mixed or noncompetitive inhibitor of recombinant and human liver microsomal UGT1A1 (K(i) range 14-18 μM), whereas competitive inhibition (K(i) 62 μM) was observed with UGT2B15. NFA was subsequently applied to the reaction phenotyping of human liver microsomal acetaminophen (APAP) glucuronidation. Consistent with previous reports, APAP was glucuronidated by recombinant UGT1A1, UGT1A6, UGT1A9, and UGT2B15. NFA concentrations in the range of 2.5 to 100 μM inhibited APAP glucuronidation by UGT1A1, UGT1A9, and UGT2B15 but not by UGT1A6. The mean V(max) for APAP glucuronidation by HLM was reduced by 20, 35, and 40%, respectively, in the presence of 2.5, 50, and 100 μM NFA. Mean K(m) values decreased in parallel with V(max), although the magnitude of the decrease was smaller. Taken together, the NFA inhibition data suggest that UGT1A6 is the major enzyme involved in APAP glucuronidation.
酶选择性抑制剂是反应表型鉴定最有价值的实验工具。然而，迄今为止，仅鉴定出有限数量的 UDP-葡糖醛酸基转移酶 (UGT) 酶选择性抑制剂。本研究对非那西丁（NFA）的 UGT 酶选择性进行了表征。结果表明，2.5 μM NFA 是重组和人肝微粒体 UGT1A9 活性的高度选择性抑制剂。较高浓度的 NFA（50-100 μM）抑制 UGT1A1 和 UGT2B15，但对 UGT1A3、UGT1A4、UGT1A6、UGT2B4、UGT2B7 和 UGT2B17 的活性影响较小。NFA 以混合（竞争性-非竞争性）机制抑制重组 UGT1A9 的 4-甲基伞形酮和丙泊酚（PRO）葡糖醛酸化以及人肝微粒体（HLM）的 PRO 葡糖醛酸化，K(i) 值范围为 0.10 至 0.40 μM。同样，NFA 是重组和人肝微粒体 UGT1A1 的混合或非竞争性抑制剂（K(i) 范围为 14-18 μM），而与 UGT2B15 观察到竞争性抑制（K(i) 62 μM）。随后，将 NFA 应用于人肝微粒体对乙酰氨基酚（APAP）葡糖醛酸化的反应表型鉴定。与先前的报道一致，APAP 由重组 UGT1A1、UGT1A6、UGT1A9 和 UGT2B15 葡糖醛酸化。浓度为 2.5 至 100 μM 的 NFA 抑制 UGT1A1、UGT1A9 和 UGT2B15 但不抑制 UGT1A6 的 APAP 葡糖醛酸化。在存在 2.5、50 和 100 μM NFA 的情况下，APAP 葡糖醛酸化的 HLM 的平均 V(max) 分别降低了 20%、35%和 40%。平均 K(m) 值与 V(max) 平行下降，尽管下降幅度较小。总的来说，NFA 抑制数据表明 UGT1A6 是 APAP 葡糖醛酸化的主要酶。
