Cytochrome P450 3A Enzymes Are Key Contributors for Hepatic Metabolism of Bufotalin, a Natural Constitute in Chinese Medicine Chansu
Cytochrome P450 3A Enzymes Are Key Contributors for Hepatic Metabolism of Bufotalin, a Natural Constitute in Chinese Medicine Chansu
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细胞色素 P450 3A 酶是中药天然成分蟾蜍他林肝脏代谢的关键贡献者
DOI：
10.3389/fphar.2019.00052
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发表时间：
2019-02-04
影响因子：
5.6
通讯作者：
Yang, Ling
中科院分区：
文献类型：
--
作者：
Dai, Zi-Ru;Ning, Jing;Yang, Ling
蟾毒他林（Bufotalin，BFT）是一种天然存在的蟾蜍二烯内酯类化合物，具有抗肿瘤和心脏毒性等多种药理和毒理作用。本研究的目的是描述BFT在人体内的代谢途径，确定BFT在人体内肝脏代谢的关键酶，并探讨酶选择性的相关分子机制。经LC-MS/MS和NMR技术鉴定，BFT在人肝微粒体中的主要代谢产物为5β-羟基蟾毒他林。反应表型分析和化学抑制试验表明，CYP 3A 4和CYP 3A 5是BFT 5β-羟基化的关键酶。动力学分析表明，HLM和人CYP 3A 4中的BFT 5β-羟基化遵循双相动力学，而CYP 3A 5中的BFT 5β-羟基化遵循底物抑制动力学。分子对接模拟结果表明，BFT可与CYP 3A 4和CYP 3A 5上两个不同的配体结合位点结合，这部分解释了BFT在CYP 3A 4和CYP 3A 5中不同的动力学行为。这些发现对于阐明BFT在人体内的I相代谢、深入了解CYP 3A酶与蟾蜍二烯内酯之间的关键相互作用以及开发具有改善药代动力学和安全性特征的蟾蜍二烯内酯类药物非常有帮助。
Bufotalin (BFT), one of the naturally occurring bufodienolides, has multiple pharmacological and toxicological effects including antitumor activity and cardiotoxicity. This study aimed to character the metabolic pathway(s) of BFT and to identify the key drug metabolizing enzyme(s) responsible for hepatic metabolism of BFT in human, as well as to explore the related molecular mechanism of enzymatic selectivity. The major metabolite of BFT in human liver microsomes (HLMs) was fully identified as 5β-hydroxylbufotalin by LC-MS/MS and NMR techniques. Reaction phenotyping and chemical inhibition assays showed that CYP3A4 and CYP3A5 were key enzymes responsible for BFT 5β-hydroxylation. Kinetic analyses demonstrated that BFT 5β-hydroxylation in both HLMs and human CYP3A4 followed the biphasic kinetics, while BFT 5β-hydroxylation in CYP3A5 followed substrate inhibition kinetics. Furthermore, molecular docking simulations showed that BFT could bind on two different ligand-binding sites on both CYP3A4 and CYP3A5, which partially explained the different kinetic behaviors of BFT in CYP3A4 and CYP3A5. These findings are very helpful for elucidating the phase I metabolism of BFT in human and for deeper understanding the key interactions between CYP3A enzymes and bufadienolides, as well as for the development of bufadienolide-type drugs with improved pharmacokinetic and safety profiles.