Simplified assays of lipolysis enzymes for drug discovery and specificity assessment of known inhibitors
Simplified assays of lipolysis enzymes for drug discovery and specificity assessment of known inhibitors
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DOI：
10.1194/jlr.d058438
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发表时间：
2016-01-01
影响因子：
6.5
通讯作者：
Prentki, Marc
中科院分区：
文献类型：
--
作者：
Iglesias, Jose;Lamontagne, Julien;Prentki, Marc
脂类被用作细胞积木和浓缩的能量储存，也充当信号分子。甘油/脂肪酸循环，包括脂肪分解和脂肪生成，产生许多脂质信号。为了药物筛选的目的，目前还没有可靠的方法来测量几种脂解酶的活性，这导致了各种已知脂肪酶抑制剂的选择性受到质疑。我们现在描述脂肪分解酶的简单分析，包括脂肪甘油三酯脂肪酶(ATGL)、激素敏感脂肪酶(HSL)、sn-1-二酰甘油脂肪酶(DAGL)、单甘油脂肪酶、α/β-水解酶结构域6和羧酸酯酶1(CES1)。我们观察到，许多已报道的抑制剂缺乏特异性。因此，Cay10499(HSL抑制剂)和RHC20867(DAGL抑制剂)也抑制其他脂肪酶。与相应的小鼠酶相比，人ATGL和HSL对抑制剂的敏感性有显著差异。因此，ATGListatin抑制小鼠ATGL，但不抑制人ATGL，HSL抑制剂WWL11和化合物13f对小鼠酶有效，但对人酶的抑制作用较弱。其中许多脂肪酶抑制剂也抑制人类CES1。研究结果描述了可用于药物筛选的脂肪酶活性测定的可靠方法，并对许多早期描述的脂肪酶抑制剂的特异性提出了警告。
Lipids are used as cellular building blocks and condensed energy stores and also act as signaling molecules. The glycerolipid/ fatty acid cycle, encompassing lipolysis and lipogenesis, generates many lipid signals. Reliable procedures are not available for measuring activities of several lipolytic enzymes for the purposes of drug screening, and this resulted in questionable selectivity of various known lipase inhibitors. We now describe simple assays for lipolytic enzymes, including adipose triglyceride lipase (ATGL), hormone sensitive lipase (HSL), sn-1-diacylglycerol lipase (DAGL), monoacylglycerol lipase, alpha/beta-hydrolase domain 6, and carboxylesterase 1 (CES1) using recombinant human and mouse enzymes either in cell extracts or using purified enzymes. We observed that many of the reported inhibitors lack specificity. Thus, Cay10499 (HSL inhibitor) and RHC20867 (DAGL inhibitor) also inhibit other lipases. Marked differences in the inhibitor sensitivities of human ATGL and HSL compared with the corresponding mouse enzymes was noticed. Thus, ATGListatin inhibited mouse ATGL but not human ATGL, and the HSL inhibitors WWL11 and Compound 13f were effective against mouse enzyme but much less potent against human enzyme. Many of these lipase inhibitors also inhibited human CES1. Results describe reliable assays for measuring lipase activities that are amenable for drug screening and also caution about the specificity of the many earlier described lipase inhibitors.