Human Arylacetamide Deacetylase Is a Principal Enzyme in Flutamide Hydrolysis
Human Arylacetamide Deacetylase Is a Principal Enzyme in Flutamide Hydrolysis
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DOI：
10.1124/dmd.109.026567
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发表时间：
2009-07-01
影响因子：
3.9
通讯作者：
Yokoi, Tsuyoshi
中科院分区：
文献类型：
--
作者：
Watanabe, Akinobu;Fukami, Tatsuki;Yokoi, Tsuyoshi
氟替卡松是一种抗雄激素药物，广泛用于治疗前列腺癌。氟替卡松的初始代谢途径是羟基化和水解。最近报道，水解产物4-硝基-3-（三氟甲基）苯胺（FLU-1）进一步代谢为N-羟基FLU-1，一种假定的肝毒性物质。然而，还没有表征负责芴醇水解的酯酶。在本研究中，我们发现，人芳基乙酰胺脱乙酰酶（AADAC）有效地水解芴醇使用重组AADAC在COS 7细胞中表达。与此相反，负责许多药物水解的羧酸酯酶1（CES 1）和CES 2不能水解氟西汀。AADAC在内质网中特异性表达。在人肝微粒体中检测到较高的氟苷水解酶活性（K-m，794 +/- 83 μ M; V-max，1.1 +/- 0.0 nmol/min/mg蛋白），而在人肝胞质溶胶中活性极低。双-（壬基苯基）-苯基磷酸盐、二异丙基氟磷酸盐和硫酸毒扁豆碱（依地平）强烈抑制人肝微粒体中的氟替卡松水解酶活性，但氟化钠、苯甲基磺酰氟和双硫仑中度抑制。用重组AADAC获得了相同的抑制模式。此外，显示AADAC表达的人肝和空肠微粒体可水解氟替卡松，但不表达AADAC的人肺和肾微粒体显示轻微活性。在人肝微粒体（n = 50）中，AADAC蛋白的表达水平与荧光素水解酶活性呈显著正相关（r = 0.66，p < 0.001）。总之，这些结果清楚地表明，氟草胺仅被AADAC水解。AADAC可能是引起氟他胺肝毒性的重要酶。
Flutamide, an antiandrogen drug, is widely used for the treatment of prostate cancer. The initial metabolic pathways of flutamide are hydroxylation and hydrolysis. It was recently reported that the hydrolyzed product, 4-nitro-3-(trifluoromethyl)phenylamine (FLU-1), is further metabolized to N-hydroxy FLU-1, an assumed hepatotoxicant. However, the esterase responsible for the flutamide hydrolysis has not been characterized. In the present study, we found that human arylacetamide deacetylase (AADAC) efficiently hydrolyzed flutamide using recombinant AADAC expressed in COS7 cells. In contrast, carboxylesterase1 (CES1) and CES2, which are responsible for the hydrolysis of many drugs, could not hydrolyze flutamide. AADAC is specifically expressed in the endoplasmic reticulum. Flutamide hydrolase activity was highly detected in human liver microsomes (K-m, 794 +/- 83 mu M; V-max, 1.1 +/- 0.0 nmol/min/mg protein), whereas the activity was extremely low in human liver cytosol. The flutamide hydrolase activity in human liver microsomes was strongly inhibited by bis-(nonylphenyl)-phenylphosphate, diisopropylphosphorofluoride, and physostigmine sulfate (eserine) but moderately inhibited by sodium fluoride, phenylmethylsulfonyl fluoride, and disulfiram. The same inhibition pattern was obtained with the recombinant AADAC. Moreover, human liver and jejunum microsomes showing AADAC expression could hydrolyze flutamide, but human pulmonary and renal microsomes, which do not express AADAC, showed slight activity. In human liver microsomal samples (n = 50), the flutamide hydrolase activities were significantly correlated with the expression levels of AADAC protein (r = 0.66, p < 0.001). In conclusion, these results clearly showed that flutamide is exclusively hydrolyzed by AADAC. AADAC would be an important enzyme responsible for flutamide-induced hepatotoxicity.