基于体内外模型的新苯二氮卓类衍生物代谢与药动学机制研究

The continual emergence and rapid spread of novel psychoactive substances (NPS) is a serious social problem in worldwide. Current tools of their safety and efficacy assessments can not catch up with NPS appeared. NPS problems present an ongoing challenge for law enforcement authorities to effectively combat due to the limited toxicological data available. Study on effects of NPS on metabolism and toxicity is a difficult and hot research in forensic medicine. New designer benzodiazepines have become a rapidly growing class of drugs of abuse. In China, new designer benzodiazepines are the most frequently observed compounds in cases of drug-facilitated sexual assaults. There is very limited data on metabolism and pharmacokinetic mechanisms of new designer benzodiazepines, thus making the toxicological interpretation in cases problematic. Based on the previous study, the project is to create in vivo and in vitro models exposed with new designer benzodiazepines first. The processes of metabolism and pharmacokinetics are studied. The phase I metabolites and phase II metabolites are identified by high resolution mass spectrometry in vivo model. The metabolic pathways in human and rat are mapped. The same targets and reliable biomarkers are explored. The pharmacokinetic characteristics are investigated to clear relationships between structure activity and metabolism, clearance. The metabolic phenotypings of new designer benzodiazepines are identified in vitro hepatocytes. The mechanisms of metabolism and elimination are elucidated to reveal the relationships between NPS abuse kinetics, their mechanisms and health risk. The studies aim to develop full-scale, effective models to study metabolism and pharmacokinetics of NPS and to explore their drug efficacy and toxicity mechanisms. Such data provides new technical support to control NPS in our country.

新精神活性物质(NPS)的全球快速蔓延引发严重的社会问题，而其体内代谢和安全评价数据缺乏是目前法医学研究及鉴定的热点和难点。本项目拟在前期研究基础上，以当前药物辅助性犯罪案件频现的新苯二氮卓类衍生物为目标物，构建体内外实验模型进行体内处置和动力学过程研究。通过体内实验模型和高分辨质谱等技术，确认不同种属的体内代谢过程及其I相、Ⅱ相代谢物，探索药物代谢规律以及共性、特征标志物；查明药代动力学特征，揭示药物取代基与代谢、消除及生物转化的关系；利用体外基因重组CYP450s酶系模型，鉴别参与生物转化的CYP酶代谢表型，进一步阐明代谢消除机制，为揭示人体滥用NPS后的生物学和物理化学过程及机制、评估潜在毒性风险提供科学依据。本研究旨在以新苯二氮卓类衍生物为代表，建立适用于NPS代谢与药动学特征研究的科学、有效的实验模型，并探讨NPS药效及毒性的相关机制，以期为我国NPS滥用的管控提供新的技术保障。

本研究基于体内外模型，研究当前药物辅助性犯罪案件频现的新苯二氮卓类衍生物的代谢与药动学机制。优化前处理、色谱条件及质谱参数，构建高分辨质谱数据库，形成前处理简便、快速，灵敏度高，筛选范围宽泛的高通量鉴识方法；率先将热辅助碳纤维离子化技术应用于生物基质中目标物鉴别；利用pHLM孵育和斑马鱼孵育两个代谢模型，研究科纳唑仑的代谢途径；研究氟阿普唑仑药动学及药效与毒性作用的机制研究，考察CYP450重组酶对氟阿普唑仑在人肝微粒体中孵育代谢的影响，发现介导氟阿普唑仑代谢的酶型为CYP2B6 及CYP3A4；构建大鼠给药模型，利用LC-QE考察氟阿普唑仑在大鼠血样、尿样、粪便及胆汁中的代谢物；研究二氯西泮与其同分异构体4'-氯地西泮的代谢特征差异，通过色谱、质谱可有效区分二氯西泮及其同分异构体4'-氯地西泮的体内代谢物；在此基础上，通过一系列大鼠肝微粒体温孵体系实验，发现新型1,4-苯二氮䓬类物质的代谢反应类型主要包括脱烷基化、羟基化、乙酰化、还原反应、葡萄糖醛酸结合等代谢反应，明确了新型1,4-苯二氮䓬类物质的代谢规律和质谱特征。. 本项目率先将热辅助碳纤维离子化技术应用于生物基质中目标物鉴别，利用质谱技术研究科纳唑仑、氟阿普唑仑、二氯西泮和4’-氯地西泮等新苯二氮卓类衍生物的药物代谢机制和药代动力学规律，补充了新苯二氮卓类衍生物的基础数据，其研究具有原始的创新性。本研究共发表15篇文章，其中SCI文章12篇；参加国际会议2次；形成标准1个；获得质谱学报优秀论文奖。. 本项目研究成果明确新苯二氮卓类衍生物的代谢规律、质谱特征和体内标志物，不仅提升新精神活性物质法医毒物学的鉴别能力，而且为新精神活性物质的发现、认识与管控提供新路径。