In vitro characterisation of human renal and hepatic frusemide glucuronidation and identification of the UDP-glucuronosyltransferase enzymes involved in this pathway
In vitro characterisation of human renal and hepatic frusemide glucuronidation and identification of the UDP-glucuronosyltransferase enzymes involved in this pathway
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DOI：
10.1016/j.bcp.2008.04.014
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发表时间：
2008-07-15
影响因子：
5.8
通讯作者：
Miners, John O.
中科院分区：
文献类型：
--
作者：
Kerdpin, Oranun;Knights, Kathleen M.;Miners, John O.
为了深入了解速尿（FSM）的肾脏和肝脏葡萄糖醛酸化，本研究：（i）通过人肝微粒体（HLM）和人肾皮质-（HKCM）和髓质-（HKMM）微粒体表征FSM葡萄糖醛酸化的动力学，（ii）鉴别参与该途径的人UDP-葡萄糖醛酸基转移酶。HLM、HKCM和HLMM可有效葡萄糖醛酸化FSM。在所有组织中，FSM葡糖苷酸（FSMG）形成遵循Michaelis-Menten动力学。虽然HKMM形成FSMG的平均Km（386 +/- 68 mu M）低于HLM（988 +/- 271 mu M）和HKCM（704 +/- 278 mu M）的K（m）值，但三种组织的平均V（max）/K（m）值相当。筛选一组重组UGT酶的葡萄糖醛酸FSM能力。UGT 1A 1、1A 3、1A 6、1A 7、1A 9、1A 10和2B 7代谢FSM。在肾脏和肝脏表达的酶中，动力学参数的比较表明UGT 1A 9在FSM葡萄糖醛酸化中起主要作用，尽管UGT 1A 1也可能促进HLM形成FSMG。与这些观察结果一致，UGT 1A选择性抑制剂保泰松和磺酰草胺使HIM、HKCM和HKMM的FSMG形成减少60- 80%，而UGT 2B 7选择性抑制剂氟康唑使FSM葡萄糖醛酸化减少60- 80%。
In order to gain insights into the renal and hepatic glucuronidation of frusemide (FSM), this study: (i) characterised the kinetics of FSM glucuronidation by human liver microsomes (HLM) and human kidney cortical- (HKCM) and medullary- (HKMM) microsomes, and (ii) identified the human UDP-glucuronosyltransferase enzyme(s) involved in this pathway. HLM, HKCM and HLMM efficiently glucuronidated FSM. FSM glucuronide (FSMG) formation followed Michaelis-Menten kinetics in all tissues. While the mean Km for FSMG formation by HKMM (386 +/- 68 mu M) was lower than the K(m) values for HLM (988 +/- 271 mu M) and HKCM (704 +/- 278 mu M), mean V(max)/K(m) values were comparable for the three tissues. A panel of recombinant UGT enzymes was screened for the capacity to glucuronidate FSM. UGT 1A1, 1A3, 1A6, 1A7, 1A9, 1A10 and 2B7 metabolised FSM. Of the renally and hepatically expressed enzymes, comparison of kinetic parameters suggests a predominant role of UGT1A9 in FSM glucuronidation, although UGT1A1 may also contribute to FSMG formation by HLM. Consistent with these observations, the UGT1A selective inhibitors phenylbutazone and sulfinpyrazone decreased FSMG formation by HIM, HKCM and HKMM by 60-80%, whereas the UGT2B7 selective inhibitor fluconazole reduced FSM glucuronidation by