基于肿瘤异常代谢酶NQO1 还原活化的前药系统的构建并设计合成NQO1 靶向抗肺癌前药

NAD(P)H:quinone oxidoreductase-1 (NQO1) is an abnormal metabolic reductase highly elevated in cancer cells, especially in lung cancer. NQO1 can catalyze the two-electron reduction of a wide variety of quinones to hydroquinones. Based on this bioreductive activation, a novel type of NQO1-targeted prodrug has innovatively been designed. The prodrug consists of three components including a NQO1 trigger group by reduction, a self-cleavable linker and a coupled anti-cancer drug. Our research group has recently revealed that the reported trigger group showed poor selectivity towards NQO1, which can also be triggered by CPR, a one-electron reductive metabolic reductase widely exists in the liver tissues. This could lead to potential hepatotoxicity. By analyzing the structural feature of the catalytic pocket of both NQO1 and CPR, in this project we are promoted to improve the selectivity of the trigger group for NQO1 based on the structural factors and redox potentials. Firstly, in order to rapid evaluation of the selectivity of trigger group at enzyme-based and cell-based levels, we have designed NQO1-triggered small-molecule fluorescent probes by replacing the drug fragment of the prodrug with a fluorescent reporter group. In the light of the structure properties for NQO1 selectivity, we ought to obtain highly selective NQO1 trigger group and NQO1-targeted prodrug system which is expected to be promising approaches for anti-lung cancer drug discovery. In this project, we mainly focus on combining three kinds of drugs including the current first-line anti-lung cancer drug Gefitinib, the anti-metabolite drug Fluorouracil, and a novel broad-spectrum antitumor candidate with the NQO1-target prodrug system to discover novel NQO1-targeted prodrugs for lung cancer treatment with high potency and low toxicity.

NQO1是肿瘤(特别是肺癌)中的异常还原代谢酶，其可将醌双电子还原为氢醌。本项目创新性利用该还原激活设计新型NQO1靶向前药，其由NQO1还原触发基团、自剪切连接链及抗肿瘤药物三部分组成。本项目组前期探索发现，现有还原触发基团不具NQO1选择性，其还可被肝脏中单电子还原酶CPR激活而脱靶释药，产生潜在肝毒性。在分析NQO1/CPR催化口袋特征的基础上，本项目拟从结构因素及氧化还原电位两大方面优化NQO1触发基团的活性及选择性。首先将前药中药物片段替换为荧光报告基团，获得NQO1还原触发型荧光探针，用于酶及细胞水平快速、可视化评价触发基团的选择性，探明选择性规律，并获得高选择性NQO1还原活化前药系统。该前药系统有望为肺癌靶向治疗开辟一条新途径。本项目着重将抗肺癌一线药吉非替尼、抗代谢药氟尿嘧啶及本课题组开发的广谱抗肿瘤候选药物与前药系统相连，以期获得高效低毒的新型抗肺癌NQO1靶向前药。

NQO1是肿瘤(特别是肺癌)中的异常还原代谢酶，其可将醌双电子还原为氢醌。本项目创新性利用该还原激活设计新型NQO1靶向前药，其由NQO1还原触发基团、自剪切连接链及抗肿瘤药物三部分组成。NQO1还原触发基团的选择性很关键，现有的NQO1底物往往缺乏选择性，可快速被肝脏中单电子还原酶CPR激活而脱靶，产生潜在肝毒性。本项目在分析NQO1/CPR催化口袋特征的基础上，设计合成了结构多样的邻醌/对醌触发基团50个，从结构因素及氧化还原电位两大方面优化NQO1触发基团的活性及选择性。通过基于荧光探针及辅酶NADPH的评价与筛选方法，发现邻醌/对醌母体上具有邻二甲基、邻二甲氧基取代、侧链中含有苯环时具有较优NQO1还原触发活性及选择性，并且获得了NQO1/CPR选择性大幅提升的触发基团及连接链体系。本项目研究进一步将抗肺癌一线药吉非替尼、抗代谢药氟尿嘧啶及本课题组开发的广谱桥环呫吨酮类抗肿瘤候选药物的药效骨架部分与前药系统相连，设计合成了50个NQO1触发型前药分子，并进行了酶、细胞水平的活性评价，发现若需要维持原药的体外抗肿瘤活性，前药分子体系整体分子量不宜过大。本项目研究最终获得了具有良好肿瘤细胞/正常细胞选择性的NQO1还原活化靶向前药5个，并从分子水平合理阐释了NQO1还原触发、自剪切的释药机制。其中1个优选前药分子在体内表现出了良好的抗非小细胞肺癌活性及选择性，体内安全性显著优于原药5-FU，可作为高效低毒的新型抗肿瘤候选药物进一步研究。此外，本项目还发现了首个基于亲和力的NQO1小分子荧光探针，可用于细胞内NQO1酶的可视化成像研究。本项目研究为NQO1还原活化靶向前药体系及相关前药设计研究奠定了基础，为NQO1高表达的肺癌靶向治疗开辟了一条新途径。在本项目资助下，共发表标注本项目的SCI论文共计6篇，单篇最高影响因子6.429；在药物化学相关会议上发表会议论文5篇；申请中国发明专利1项；培养博士研究生1名，硕士研究生1名。