基于细胞色素P450酶代谢研究中药钩吻的毒性及配伍减毒机制

Gelsemium elegans, namely Duan changcao, a toxic traditional Chinese medicine, has significant analgesia effect and no addiction. It is used to treat rheumatism pain and neuralgia. However, its side effects greatly limit its clinical application. Therefore, how to attenuate toxicity is the key problem for security application of Gelsemium elegans. Our previous study firstly found that the cytochrome P450 enzyme CYP3A4/3A5 and CYP2E1 were mainly responsible for the metabolism of gelsemine. The metabolites included demethylation, hydroxylation and oxidation of metabolites and their toxicity was significantly decreased compared with parent drug. Thus, CYP enzyme metabolism is an important factor in regulating Gelsemium elegans toxicity. Based on explication of the CYP enzyme metabolism mechanism, our project will introduce CYP transfected cells and gene knockout mice models, with the observation on the changed toxicology effect and analgesic effect, to further study of CYP enzyme metabolism in Gelsemium elegans toxicity and efficacy. And then, from the perspective of detoxication by compatibility, we will determine the Gelsemium elegans toxicity and efficacy after compatibility of radix glycyrrhizae (CYP3A inducer). Our project illustrates the mechanism of CYP enzyme metabolic effects on Gelsemium elegans toxicity-effect and reveal the scientific connotation of Gelsemium elegans detoxication, so as to provide a new basis for its safe application.

有毒中药钩吻又名断肠草，其镇痛效果显著且无依赖性，用于治疗风湿痹痛和神经痛，但是其毒副作用大大限制了它的临床应用。因此，如何减毒是钩吻安全应用需解决的关键问题。我们研究首次发现钩吻吲哚型生物碱毒效成分在细胞色素P450 (CYP) 酶CYP3A4/3A5、CYP2E1的主要作用下，形成脱甲基、羟基化和氧化的代谢产物；而且，这些代谢产物的细胞毒性显著降低。由此，我们认为CYP酶代谢作用是调控钩吻毒性的重要因素。本项目将在明确钩吻生物碱CYP酶代谢机理的基础上，首先采用CYP转基因细胞和基因敲除小鼠模型，以毒理效应和镇痛效果的变化为观察指标，深入研究CYP酶代谢在钩吻毒性及药效中的作用；进而从配伍减毒的角度，配合药效评价，研究甘草（CYP3A诱导剂）与钩吻配伍减毒的作用。本项目通过阐明CYP酶代谢影响钩吻毒-效的作用机制，揭示钩吻减毒的科学内涵，从而为其安全应用提供新依据。

有毒中药钩吻又名断肠草，其镇痛效果显著且无依赖性，用于治疗风湿痹痛和神经痛，但是其毒副作用大大限制了它的临床应用。因此，如何减毒是钩吻安全应用需解决的关键问题。在国家基金青年项目的资助下，本课题研究发现钩吻有效成分钩吻素子、钩吻素甲和胡蔓藤碱甲在P450酶的作用下发生显著的代谢，经NMS和高分辨率质谱的鉴定，其代谢包括羟基化和去甲基等代谢反应，该代谢反应主要经同工酶CYP3A4/5介导。小鼠毒性实验发现抑制CYP3A4活性后，钩吻生物碱的毒性显著地下降。斑马鱼毒性实验发现钩吻生物碱的毒性显著地大于其CYP介导形成的代谢产物。此外，药效实验初步表明抑制CYP3A4活性后，其镇痛效果没有发生显著变化。钩吻与甘草配伍后可以显著降低其毒性。因此，CYP酶代谢在钩吻的毒性作用中是解毒作用，揭示了钩吻减毒的科学内涵，为其安全应用提供了新依据，已发表SCI论文4篇。