Cyclo-Ru-IQ的设计、合成及其抑制Nrf2逆转非小细胞肺癌顺铂耐药的分子机制

The latest findings show that Nrf2/ARE signaling pathway is closely related to the occurrence, proliferation and drug resistance of lung cancer, and has become a new target for lung cancer chemotherapy. Inhibition of this pathway can reverse lung cancer mutidrug resistance, thereby enhancing the efficacy of chemotherapy. Our previous studies suggested that cyclometallated ruthenium compounds equiped with isoquinoline derivatives (Cyclo-Ru-IQ) possessed obvious antitumor properties, could reverse A549/DDP cisplatin resistance and down regulate the level of Nrf2. Therefore, we speculate that “Cyclo-Ru-IQ could reverse non-small cell lung cancer(NSCLC) cisplatin resistance by inhibiting the Nrf2/ARE signaling pathway”. To confirm this hypothesis, in this project, four series of Cyclo-Ru-IQ will be synthesized firstly, and the inhibitory activity toward NSCLC cisplatin resistance cancer cells will be detected in vitro and in vivo through cell experiments and animal experiments. The structure-activity relationship which will be obtained through theoretical calculation further guide the optimization of molecular design. The effects of Cyclo-Ru-IQ on the upstream and downstream signaling pathways of Nrf2/ARE will be investigated, and the molecular mechanism of reversing NSCLC cisplatin resistance and inhibiting Nrf2 expression will be explored and elucidated. The results of these studies will provide a theoretical basis for the development of novel Nrf2 signaling pathway inhibitors and provide new ideas for the development of cyclometallated ruthenium antitumor drugs.

Nrf2/ARE信号通路与肺癌的发生、增殖及其耐药均密切相关，已成为肺癌化疗的新靶点。抑制该通路能够逆转肺癌耐药，从而增强化疗效果。本课题组研究发现，异喹啉类环金属化钌配合物（Cyclo-Ru-IQ）具有优异的抗肿瘤活性，能够逆转A549/DDP顺铂耐药，且可以下调Nrf2的表达。我们推测“Cyclo-Ru-IQ主要通过抑制Nrf2/ARE信号通路，逆转非小细胞肺癌顺铂耐药”。为证实此假说，本项目拟合成4个系列的Cyclo-Ru-IQ配合物，通过细胞实验及动物实验，检测其在体外、体内逆转非小细胞肺癌顺铂耐药的效果，并通过理论计算获取配合物的构效关系，指导优化分子设计；研究配合物对Nrf2/ARE上下游信号通路的调控，探讨并阐明其逆转非小细胞肺的顺铂耐药和抑制Nrf2表达的分子机制。项目的研究结果将为开发新型的Nrf2信号通路抑制剂提供理论依据，也为环金属化钌类抗肿瘤药物的开发提供新思路。

肺癌的耐药与Nrf2/ARE信号通路的异常活化密切相关。为阐明金属钌配合物逆转NSCLC的顺铂耐药的相关机制，本项目设计合成了系列环金属化异喹啉类钌配合物（Cyclo-Ru-IQ），从细胞实验和动物实验两个层面上详细研究了其抗肿瘤活性及对Nrf2/ARE上下游信号通路的调控，探讨其逆转A549/DDP顺铂耐药的分子机制。研究结果表明，与非环金属化的异喹啉类钌配合物相比，Cyclo-Ru-IQ抑制肿瘤细胞的活性提高了30多倍，这源于其结构的改变引起配合物的疏水性增大，细胞吸收增多。另外，构效关系研究表明主配体含有给电子基团比相应的含有吸电子基团的配合物的肿瘤细胞抑制活性更好。重要的是，Cyclo-Ru-IQ对耐药细胞A549/DDP具有良好的抑制作用，且可以通过多个细胞通路协同促进耐药细胞的凋亡。1、通过抑制CDK1/Cyclin B和Cyclin A的表达，诱导细胞S，G2/M双周期阻滞，进而通过诱导线粒体膜电位降低，细胞内ROS水平升高，DNA损伤等途径诱导细胞凋亡。2、通过下调p-AKT蛋白表达水平，上调p-GSK3β和p-Fyn的水平，促进Nrf2降解，进而逆转A549/DDP的耐药。同时，免疫荧光实验结果表明Cyclo-Ru-IQ可以阻止Nrf2在A549/DDP细胞核中的表达。3、通过抑制PI3K，p-Akt，p-mTOR及Nrf2和MRP1的表达，逆转A549/DDP的耐药。值得注意的是，本课题所设计合成的系列Cyclo-Ru-IQ配合物对人正常HK-2细胞以及斑马鱼胚胎均表现出较低的毒性。总之，本项目研究结果表明系列Cyclo-Ru-IQ配合物既可以诱导肿瘤细胞凋亡，又可以通过调控Nrf2信号通路的上、下游相关蛋白的表达，逆转A549/DDP的耐药。项目研究结果表明该类配合物具有潜在的开发价值，为临床上NSCLC耐药的治疗提供了新思路。