针对免疫复合物肾小球肾炎的siRNA靶向递释系统研究

In China, there is an increasing trend of morbidity of end-stage renal disease (ESRD) which is mainly caused by glomerulonephritis (GN). Here, GN itself is often initiated by immune complex (IC) deposits in the glomeruli, which will activate humoral mediated and/or cellular mediated immune response to damage the glomerulus. Numerous researches have already established that IC mainly deposits in subcutaneous and mesangial areas of glomeruli. Currently, the clinical treatment of GN mainly relies on the combined use of immunosuppressants such as glucocorticoids and cytotoxic drugs such as cyclophosphamide. Unfortunately, these treatments have serious adverse effects. This project aims to use phospholipid, which could be used in intravenous injection, to construct p38MAPK-siRNA loaded solid lipid nanoparticles (SLN) which will target to glomerular basement membrane, endothelial cells (EC), mesangial cells (MC) and podocyte (PC). We plan to explore the relationship between targeting efficiency and various characteristics of these particles, in order to screen out the most preferable electric potential and particle size range for targeting. Consequently, p38MAPK-siRNA will be loaded onto the particles so that the siRNA can be delivered into PC cells and cells located in glomerular basement membrane, EC, and MC. Hence, p38MAPK could be silenced at post-transcriptional level in these regions, reducing the expression level of inflammatory factors, which in turn reduces tissue damage and glomerular cell lesion. In this way, our high efficiency siRNA delivery system can be used to treat GN. Also, the glomerular targeting capability, therapeutic effect and working mechanism of this delivery system would be further investigated as part of our research. All together, this project could lay the cornerstone for developing effective and low-toxic targeted drugs against GN.

我国终末期肾病（ESRD）患病率呈升高趋势。肾小球肾炎（GN）是ESRD的主因。GN常由免疫复合物于肾小球沉积引起，其主要沉积区在肾小球内皮下和系膜区。临床治疗GN主要以糖皮质激素和细胞毒剂联用，毒副作用严重。申请人所在课题组已实现肾小球系膜细胞靶向递药。本项目拟在此基础上，以基底膜和内皮细胞（EC）、系膜细胞（MC）和足细胞（PC）为靶，以可用于静脉注射的磷脂为载体，构建载p38MAPK-siRNA的固体脂质纳米粒（SLN）。探索其粒径、电性与靶向效率的关系，优化能靶向基底膜、EC、MC和PC细胞的SLN的最适电性和粒径范围。将siRNA靶向递送至肾小球基底膜，进而进入EC、MC和PC细胞内，诱导炎症基因p38MAPK转录后水平的基因沉默，减少炎症因子表达，从而减少组织损伤及肾小球细胞病变，达到治疗GN的目的，并评价其靶向性、治疗效果及作用机制。为研发高效低毒的GN治疗靶向新药提供基础。

一、原计划完成的工作.本课题依托项目为针对免疫复合物肾小球肾炎的siRNA靶向递释系统研究，计划以基底膜和内皮细胞（EC）、系膜细胞（MC）和足细胞（PC）为靶，以可用于静脉注射的磷脂为载体，构建载p38MAPK-siRNA的固体脂质纳米粒（SLN）。探索其粒径、电性与靶向效率的关系，优化能靶向基底膜、EC、MC和PC细胞的SLN的最适电性和粒径范围。将siRNA靶向递送至肾小球基底膜，进而进入EC、MC和PC细胞内，诱导炎症基因p38MAPK转录后水平的基因沉默，减少炎症因子表达，从而减少组织损伤及肾小球细胞病变，达到治疗GN的目的，并评价其靶向性、治疗效果及作用机制。为研发高效低毒的GN治疗靶向新药提供基础。..二、实际完成情况.截至2022年12月31日，课题组顺利完成了原申请书中所有研究任务。首先，成功优化配方制备了载有p38MAPK和p65的对应siRNA的脂质纳米粒。其次，在体外和动物模型中测试验证了该脂质纳米系统的靶向能力、循环时间、治疗效果、安全性等。研究结果证明该纳米粒可以有效地靶向至肾小球基底膜区域的重要细胞，并在胞内释放siRNA，从而有效抑制炎症，减轻肾小球肾炎的损伤。该递药系统安全性高，效果优于常用的肾小球肾炎治疗药物，证明了siRNA的靶向递送可以为炎症性疾病治疗提供新型治疗方案。为未来相关研究提供了一定的基础。.此外，课题组还完成了数个相关研究内容。具体包括：.1）.设计构建了几类递药系统：肿瘤靶向胶原酶递送系统；利用OX40L阻断外泌体的自身免疫病治疗系统；肝素酶抑制靶向系统；多肽配体和小分子药物的靶向共递送系统；利用内源性蛋白的脑靶向系统；克服口服胃肠道粘膜屏障的生物大分子药物递释系统等。.2）.制备并探索了丁苯酞一种前药的治疗效果。.3）.探索研究了多孔二氧化硅纳米粒的孔径在免疫刺激方面的作用。.4）.课题一共发表标注基金编号（81872824）SCI论文12篇，包括JCR、Sci Adv、Biomaterials等领域权威期刊。其中11篇论文课题组成员为第一作者、通讯作者或共同通讯作者，依托单位四川大学为第一通讯作者单位。转让发明专利1项。培养硕士5人，博士1人。