羧酸酯酶调控剂的高通量高内涵筛选与评价体系及其应用

Human carboxylesterase (CEs) plays an important role in the homeostasis of lipid metabolism, and becomes the treatment target of lipid metabolism disorders related to hyperlipidemia and obesity. Due to the lack of efficient screening and evaluation methods of CEs regulators for complex biological systems, which great limits the development and application of CEs regulators. This project is to construct an effective screening and evaluation method of CEs regulator suitable for living cell system with the help of high content imaging system based on the CEs subtype fluorescent probes with in-situ imaging function. Meanwhile, by combing with subcellular localization fluorescence probes, we can synchronously evaluate the effectiveness and safety of CEs regulators at the living cell level in high conformation imaging system. The above evaluation system is applied to large-scale screening of natural products on the regulation of human two kinds of CEs, the effective discovery of CEs regulators and the synchronous evaluation of their effectiveness in living cells. Finally, we carried out animal level pharmacodynamic evaluation, optimized and corrected the whole system to form a standardized CEs regulators efficient screening and evaluation system. The project is expected to solve the key technical problems in the field of efficient screening and evaluation of CEs regulators, and also provide follow-up drugs and technical support for the treatment of lipid metabolism disorders.

人羧酸酯酶（CEs）在脂质代谢稳态中发挥着重要作用，是高血脂、肥胖等代谢性疾病的治疗靶点。长期以来，由于缺乏适于复杂生物体系的CEs调控剂高效筛选与评价方法，极大限制了CEs特异性调控剂的发现与应用。本项目拟以前期设计研发的具有原位成像功能的CEs亚型特异性荧光探针为底物，借助高内涵成像系统构建适于活细胞体系的CEs调控剂高效筛选与评价方法。同时，结合亚细胞器定位荧光探针，在高内涵成像系统上实现活细胞层面的CEs调控剂的有效性和安全性的同步评价。并将上述评价体系用于天然产物对人两种CEs调控作用的规模化筛选，CEs调控剂的高效发现以及其药效的精细评价等研究。在此基础上，结合动物实验进一步验证该评价体系最终形成标准化的CEs调控剂的高效筛选与评价方法。该项目有望解决CEs调控剂高效筛选与评价领域面临的关键技术难题，同时还可为寻找脂质代谢紊乱相关疾病的治疗药物提供后续药物和技术支撑。

羧酸酯酶（CEs）不仅参与多种酯类前药的代谢，同时也是高血脂、肥胖等代谢性疾病的治疗靶点。构建适于复杂生物体系的CEs调控剂高效筛选与评价方法，从天然产物资源中发现安全强效的 CEs 特异性调控剂，不仅有助于开发脂类代谢相关疾病的候选药物，还有助于深入研究 CEs 内源性功能。本项目拟以前期设计研发的CEs亚型特异性荧光探针和商品化细胞器探针为底物，借助高内涵成像系统构建适于活细胞体系的CEs调控剂高通量高内涵筛选与评价体系。随后利用此体系开展CEs抑制剂/诱导剂的药效和安全性的高通量筛选及作用机制的精细表征。最后选择高品质药物开展动物层面药效评价。项目的开展有望解决CEs调控剂高效筛选与评价领域面临的关键技术难题，同时还可为寻找脂质代谢紊乱相关疾病的治疗药物提供后续药物和技术支撑。主要研究进展包括1）通过优化反应条件，构建了适于细胞水平的CEs调控剂有效性与安全性同步筛选与评价体系；2）完成了＞150种天然产物对CEs的诱导/抑制作用研究，3）完成了CEs的调控剂的抑制/诱导作用的精细表征和作用机制研究；4）开展了＞50种天然产物对其他丝氨酸水解酶抑制作用研究；5）开展了生物碱和黄酮类天然产物及降糖降脂药物的安全性高内涵评价；6）开展了动物模型构建及动物层面CEs调控剂药效评价。截至目前，本课题已圆满完成课题任务书中各项内容，其科研任务量已大大超出原有任务计划书。本项目共发表SCI论文14篇，申报国家发明专利1项，协助培养博士后1名，博士1名，硕士3名，本科生1名.