靶向结肠炎症部位巨噬细胞的口服给药系统的构建及评价

The activation of NF-κB signaling pathway in colonic macrophages plays a key role in the pathogenesis of ulcerative colitis, therefore, the inhibition of NF-κB could become an effective target for ulcerative colitis treatment. In this study, we aimed to develop an oral colon-specific drug delivery system for targeting and co-delivering drugs and siRNA to macrophages at the inflammatory site, in order to inhibit the activation of NF-κB signaling pathway and block the inflammatory cascade. Briefly, a mannose-modified nanoparticle (NP) that co-delivery NF-κBp65 siRNA and budesonide is developed and loaded in a redox-sensitive hydrogel microfiber (GEL-NP), which is further coated with hydroxypropyl methylcellulose acetate succinate to form a pH-sensitive microsphere (MP-GEL-NP). In the colon environment, the MP-GEL-NP is degraded at alkaline pH and releases the GEL-NP, which could adhere to the inflammatory site by electrostatic adsorption, meanwhile, the diselenide bonds contained in the GEL-NP would be specifically cleaved in the inflammatory environment for further releasing the NP. Subsequently, the NP could actively target the macrophages via mannose receptor-mediated endocytosis. Finally, the polyethyleneimine unit contained in the NP would achieve the intracellular release of NF-κBp65 siRNA and budesonide through the ability of endosomal escape. Consequently, the RNA interference and the glucocorticoid might provide a synergistic effect on the treatment of ulcerative colitis. Finally, based on the investigation of the preparation process and the multi bio-responsiveness of the system, the therapeutic effect of MP-GEL-NP is evaluated by animal experiments. This study could also provide new research idea and methods for the treatment of ulcerative colitis through targeting the macrophages at the inflammatory site.

肠道巨噬细胞NF-κB信号通路的活化，在溃疡性结肠炎（UC）发病中起关键作用，抑制其活化成为治疗UC的有效靶点。本项目通过构建靶向结肠炎症部位巨噬细胞的口服给药系统递送siRNA和药物，抑制NF-κB信号通路活化，阻断炎症级联反应。首先制备甘露糖修饰共载siRNA和布地奈德的纳米粒（NP），并包载于氧化敏感的炎症靶向水凝胶纤维（GEL-NP）中，再以HPMCAS包衣制备pH敏感微球。在结肠环境中，该pH敏感微球溶解释放GEL-NP，其借静电作用黏附于炎症部位，在炎症过氧化环境中其所含二硒键断裂并释放NP；NP通过表面甘露糖主动靶向巨噬细胞，并借助聚乙烯亚胺的溶酶体逃逸作用，完成NF-κBp65 siRNA和布地奈德的传递，实现RNA干扰与药物发挥协同作用。在考察给药系统制备工艺和各级生物响应性的基础上，通过动物实验评价其有效性。本项目可为靶向炎症部位巨噬细胞治疗UC提供新的研究思路和方法。

肠道巨噬细胞NF-κB信号通路的活化，在溃疡性结肠炎（UC）发病中起关键作用。NF-κB活化可促进TNF-α等促炎因子的释放，而TNF-α等释放反过来可正反馈调节NF-κB的表达，如此炎症不断被放大，形成炎症的级联反应；抑制巨噬细胞中NF-κB的活化，削弱其多途径致炎损伤作用，阻断炎症的级联效应成为治疗UC的有效靶点。基于此本项目采用RNA干扰联合药物，通过构建靶向结肠炎症部位巨噬细胞递药系统递送siRNA和药物，抑制NF-κB信号通路活化，阻断炎症级联反应治疗UC。为此本研究首先合成了PCL-PEG和PCL-PEI作为载体材料，通过调节二者比例可有效负载布地奈德（Bud）和NF-κBp65 siRNA，成功制备了共载Bud和NF-κBp65 siRNA的聚合物混合胶束（Bud/siRNA-PMM）；细胞及动物实验显示，Bud/siRNA-PMM可以有效地将NF-κBp65 siRNA和Bud递送至结肠炎症部位巨噬细胞中，实现RNA干扰与药物协同作用，抑制巨噬细胞NF-κB的活化，阻断TNF-α等炎性因子的释放，并可以诱导巨噬细胞向M2型极化，发挥治疗UC的作用；同时实验显示，对UC并发关节炎该胶束亦显示出良好的靶向治疗作用；而且该递药系统具有良好的生物安全性。在此基础上，通过静电吸附层层自组装和溶剂挥散法制备了pH敏感的、结肠炎症黏膜黏附的口服结肠靶向纳微递药系统。体内外实验显示，以壳聚糖、透明质酸及Eudrgit S100或以PEI和PLGA为载体材料，构建的靶向结肠炎症部位巨噬细胞的纳微递药系统，借助于pH敏感性及炎症部位黏膜的生物黏附性，可有效的将药物递送并富集于结肠炎症部位，并靶向递送至炎症巨噬细胞中，发挥治疗UC的作用。本项目的实施为靶向结肠炎症部位巨噬细胞递药系统的构建和药物/siRNA协同治疗UC提供了新的研究思路和方法。项目执行期间发表相关SCI论文8篇。