新型NLRP3非ATP竞争性抑制剂的设计及其抗肺癌活性探索研究

Studies have shown that NLRP3 inflammasome could promote the occurrence and metastasis of lung cancer, and therefore NLRP3 inflammasome was expected to be a potential effective target for the treatment of lung cancer and its metastasis. Most of the currently developed NLRP3 inhibitors are ATP competitive inhibitors, while the development of drugs against ATP pockets was likely to cause drug resistance or off-target. Therefore, it is of great scientific significance to explore non-ATP competitive inhibitors. Very recently, a study shown that oridonin was a non-ATP competitive inhibitor of NLRP3, but its activity was not strong enough and the structure-activity relationship was not clear. In this study, we will address those issues: on the one hand, using homology modeling and docking techniques to rationally tranform oridonin, in order to obtain new non-ATP competitive NLRP3 inhibitors with the scaffold of oridonin. On the other hand, based on the above research results, we will establish quantitative structure-activity relationship models for virtual screening to discover non-ATP competitive NLRP3 inhibitors with new skeleton structure. Meanwhile, we will evaluate those compounds for anti-lung cancer and its metastatic activities to obtain candidate compounds with relevant activities. The successful implementation of this study will provide new ideas for the discovery of NLRP3 inhibitors with novel binding mode, and provide a good molecular tool for the study of treatment of NLRP3-related diseases, thus it has important theoretical significance and application value.

研究显示NLRP3炎症小体会促进肺癌的发生与转移，有望成为抗肺癌及转移的有效靶点。目前发展的NLRP3抑制剂多为ATP竞争性抑制剂，而单一针对ATP口袋研发药物易引起耐药或脱靶，因此探索非ATP竞争性抑制剂具有重要的科学意义。最新研究发现冬凌草甲素是NLRP3非ATP竞争性抑制剂，但其活性不够强，构效关系尚不明确。本项目拟针对这些问题，一方面利用同源模建与对接技术，对冬凌草甲素进行合理的结构改造，以期获得具有冬凌草甲素骨架特征的新型非ATP竞争性NLRP3蛋白抑制剂；进一步基于上述研究结果建立定量构效关系模型以期作为虚拟筛选的模型，发掘具有新骨架结构的此类新型抑制剂。通过对新化合物进行抗肺癌与转移活性评价及作用机制探索，寻找新药候选分子。本项目的成功实施将为新型结合模式的NLRP3抑制剂的发现提供新思路，同时为研究NLRP3相关疾病的治疗提供良好的工具分子，具有重要的理论意义和应用价值。

炎症是人体维持组织稳态的一种基础性先天免疫反应，促使机体能快速响应各种有害因子引起的感染或损伤。然而，过度的炎症反应对机体组织的正常功能是有害的，并且在极端情况下会导致明显的组织损伤。核苷酸结合寡聚化结构域样受体蛋白3（NLRP3）炎症小体是机体固有免疫系统的重要调节因子，NLRP3炎症小体的过度激活涉及到多种慢性炎症性疾病的发生与发展，如痛风、结肠炎、和癌症等。开发特异性靶向NLRP3炎症小体的抑制剂有望为NLRP3驱动的炎症性疾病提供新颖的治疗策略。天然产物冬凌草甲素能够选择性地抑制NLRP3炎症小体的活化，但其抗炎活性和成药性仍然有待进一步提高。我们基于课题组前期构建的冬凌草甲素类似物库，通过筛选获得对NLRP3炎症小体具有较优抑制活性的先导化合物，再对其结构衍生化，共获得72个新化合物。其中最优化合物对NLRP3显示出两位数纳摩尔抑制活性，不会影响NLRP3的ATP酶活性，并且具有比冬凌草甲素更优的安全指数（IC50 = 77.2 nM vs 780.4 nM, SI = 40.52 vs 8.54）。作用机制显示该化合物可以通过抑制ASC寡聚化并影响NLRP3-ASC/NEK7的相互作用来阻止NLRP3的组装与激活。该化合物对炎症小体家族具有良好的选择性，可以在不影响NLRC4或AIM2炎性小体的情况下抑制多种激动剂诱导的NLRP3激活。优选化合物在尿酸钠(MSU)诱导的痛风性关节炎模型中具有耐受性和显著的抗炎活性，并且部分化合物在肿瘤模型中也具有一定抗肿瘤活性。本项目获得了一种具有新型骨架的抑制NLRP3炎性小体的先导化合物，这为进一步研究NLRP3相关疾病的治疗提供了重要的工具分子。