Carbonyl reduction of warfarin: Identification and characterization of human warfarin reductases
Carbonyl reduction of warfarin: Identification and characterization of human warfarin reductases
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DOI：
10.1016/j.bcp.2016.03.025
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发表时间：
2016-06-01
影响因子：
5.8
通讯作者：
Wsol, Vladimir
中科院分区：
文献类型：
--
作者：
Malatkova, Petra;Sokolova, Simona;Wsol, Vladimir
华法林是一种广泛使用的抗凝剂，不幸的是，这种药物通常会导致严重的不良事件，包括死亡。尽管据报道，包括华法林通过 CYP450 代谢的多种因素会影响华法林治疗的安全性和有效性，但患者中华法林剂量的巨大差异尚未完全阐明。除了CYP450介导的华法林氧化代谢外，还原代谢也参与华法林生物转化。然而，华法林的还原代谢在很大程度上尚未被探索，值得进一步研究。我们研究了人肝脏组分对华法林的还原作用，发现细胞质中华法林的还原速度比微粒体中的华法林还原速度高 9 倍（V-max = 77.2 vs. 8.7 pmol/mg 蛋白质分钟）。此外，在测试了九种重组胞质羰基还原酶还原华法林的能力时，AKR1C3和CBR1被鉴定为华法林还原酶，并确定了它们的动力学参数。 AKR1C3 的华法林内部清除率比 CBR1 高 3 个数量级（分别为 CLint = 65.922 与 0.070 μl/mg 蛋白质分钟）。这是首次在人肝亚细胞部分中鉴定出华法林还原酶。此外，我们使用 HPLC 方法描述了华法林还原的手性方面，该方法能够检测单个华法林醇立体异构体。 Cytosol 和 AKR1C3 表现出 (R)-华法林的立体选择性代谢，优先形成 (SR)-华法林醇作为华法林的主要体内代谢物。另一方面，微粒体和CBR1优先还原(S)-华法林，分别形成(RS)-华法林醇和(SS)-华法林醇。 (C) 2016 Elsevier Inc. 保留所有权利。
Warfarin is a widely used anticoagulant and, unfortunately, is a drug that is commonly implicated in serious adverse events including fatalities. Although several factors, including the metabolism of warfarin via CYP450, have been reported to affect the safety and efficacy of warfarin therapy, the wide variance in the warfarin dosage in patients has not been completely clarified. In addition to the oxidative metabolism of warfarin mediated by CYP450, reductive metabolism is involved in warfarin biotransformation. However, the reductive metabolism of warfarin has been largely unexplored and deserves further investigation. We studied warfarin reduction by human liver fractions and found a 9-fold higher velocity of warfarin reduction in the cytosol than in microsomes (V-max = 77.2 vs. 8.7 pmol/mg protein min, respectively). Furthermore, of nine recombinant cytosolic carbonyl reducing enzymes tested for their ability to reduce warfarin, AKR1C3 and CBR1 were identified as warfarin reductases and their kinetic parameters were determined. The internal clearance of warfarin was 3 orders of magnitude higher with AKR1C3 than with CBR1 (CLint = 65.922 vs. 0.070 mu l/mg protein min, respectively). This is the first time that warfarin reducing enzymes in human liver subcellular fraction have been identified. Moreover, we have described the chiral aspects of warfarin reduction using an HPLC method that enabled the detection of individual warfarin alcohol stereoisomers. Cytosol and AKR1C3 exhibit the stereoselective metabolism of (R)-warfarin to preferentially form (SR)-warfarin alcohol as the primary in vivo metabolite of warfarin. On the other hand, microsomes and CBR1 preferentially reduce (S)-warfarin to form (RS)-warfarin alcohol and (SS)-warfarin alcohol, respectively. (C) 2016 Elsevier Inc. All rights reserved.