近红外光触发的ROS响应的阳离子脂质基因载体的研究

Gene therapy is a new biomedical technology for treating cancer. However, how to achieve efficient and low-toxic gene vectors is an urgent problem to be solved. In the previous study, the constructed lipid gene vectors of different structures can achieve effective cell uptake, but there are still insufficient escape of endosomes, the stimuli response bond is difficult to break, and the performance of gene expression is not ideal. This project intends to utilize the special environment of cancer cells and combine the properties of light-emitting molecules to release ROS and fluoresce in near-infrared light. We designed a cationic lipid gene vector based on ROS to respond to thioketal linkages to construct light-controlled release and imaging. On the one hand, the production of ROS by photoactive molecules promotes the escape of endosomes and breaks the ROS-sensitive thioketal linkage to release DNA. On the other hand, the light-sensitive molecules in the nanoparticles emit fluorescence under near-infrared light, trace the transport path of the nanoparticles in the cells or within the tissues, and also investigate the escape ability of the endosomes and the release of DNA according to the distribution of fluorescence and DNA. The situation, in turn, clarifies the process and mechanism of gene delivery, and obtains the structure-activity relationship, providing a research basis for exploring new and efficient, low-toxicity and intelligent non-viral gene vectors.

基因治疗是治疗癌症的新型生物医学技术。然而，如何实现高效低毒的基因载体是目前亟待解决的问题。在前期的研究中，通过构建不同结构的脂质基因载体，可实现有效的细胞摄取，但是仍存在内涵体逃逸不足，刺激响应连接键难以断裂，以及基因表达的性能不够理想。本项目拟利用癌细胞的特殊环境，并结合光敏分子在近红外光照释放ROS和发出荧光的特性，设计基于ROS响应硫缩酮连接键构建光控释放及成像的阳离子脂质基因载体。一方面，释放的ROS能促进内涵体逃逸并且ROS响应硫缩酮连接键断裂释放DNA。另一方面，纳米颗粒中的光敏分子在近红外光照下发出荧光，示踪纳米颗粒在细胞内或组织内的运输路径，也能根据荧光与DNA的分布情况考察内涵体逃逸能力及DNA的释放情况，进而阐明递送基因过程和机理，获得构效关系，为探索新型的高效、低毒及智能型的非病毒基因载体提供研究基础。

癌症治疗是人类面临的重大健康难题之一，设计与制备新型高效低毒且肿瘤微环境响应性的纳米药物成为癌症治疗的研究热点。然而大多数纳米药物因为难以在作用位点高效累积，无法及时有效地释放药物和肿瘤微环境的影响限制其治疗效果。本项目构建了兼具近红外光响应、线粒体靶向和GSH消耗的纳米药物体系。首先采用ROS响应的硫缩酮作为连接键，靶向分子三苯基膦（TPP）作为亲水头部，光敏剂PPa作为疏水尾部的构建线粒体靶向脂质前药，同时阳离子脂质前药作为药物载体包裹GSH消耗剂马来酸二乙酯(Diethylmaleate, DEM)，构建ROS响应性纳米药物体系。此纳米药物具有存储稳定性好、靶向癌细胞的线粒体、ROS和近红外光触释放DEM。纳米药物有效增加线粒体的用药浓度，同时最大程度增加近红外光照产生ROS的杀伤力，导致线粒体功能紊乱，从而促进癌细胞凋亡。通过本项目的实施，有效改善癌症的治疗效果，为响应型脂质体纳米药物设计制备和临床治疗提供理论以及实验基础，具有十分重要的科学意义。