双模态可视化自组装基因载体用于干细胞治疗的研究

Cell-based therapies show great potential for cancer treatment. With specific tumor-oriented migration and incorporation property, mesenchymal stem cells (MSCs) have been used as efficient targeted delivery vehicles in gene therapy. However, for clinical applications, the therapeutic effects are far from satisfactory. One of the impediments to successful gene therapy is the inefficient delivery of genes to cells and the inability to monitor therapeutic response at the targeted site. Thus, in this project, we aim to develop dual-modal molecular imaging functionalized self-assemble transfection agents for stem cell therapy. A serial of hydrophobically modified peptide dendrons will be synthesized and used to construct nanoparticles combining with magnetic resonance imaging (MRI) agents and optical imaging (OI) agents. We will synthesize amphiphilic dendrons with the different degree of hydrophobic and cationic units' modification to form nanoparticles with high surface-area-to-volume ratio and high charge density via self-organisation progress, thereby these vectors can be used for efficient gene condensation and delivery. Furthermore, the superparamagnetic iron oxide (SPIO) nanocrystals, MRI agents, are self-assembled into the controlled clusters in the hydrophobic core and small molecular Cy5.5, OI agents, can be conjugated on the hydrophilic corona. Herein, the nanoparticles with different size and various compositions will be constructed, and then the nanoparticles' transfection efficiency and imaging sensitivity will be evaluated. We seek to determine and delineate the physical and chemical properties of the novel dual-modality molecular imaging transfect agents and develop a structure-activity relationship understanding of the way in which amphiphilic dendrons with hydrophilic cationic groups, OI agents and a variety of lipophilic units at their focal points can self-assemble with SPIO nanocrystals with elevated stability and effective imaging, and subsequently bind to DNA with high transfection efficiency. A comprehensive understanding of mesenchymal stem cell based cancer therapy mechanisms will be used in laboratory modules for the development of stem cell based anti-cancer therapy. Project from this proposed research holds great promise for clinical translation.

应用间充质干细胞作为肿瘤靶向性基因药物递送系统是当前干细胞发展领域的研究热点。本项目将设计、开发具有自主知识产权的可视化基因载体，旨在解决间充质干细胞基因转染的安全性问题，同时实现干细胞体内抗癌治疗的无创示踪。设计合成两亲性肽类树突状大分子，应用自组装方式同时引入磁共振（MRI）和光学成像（OI）显影成分，并通过结合阳离子小分子自组装形成高密度正电荷纳米粒子完成治疗基因的携带、保护及干细胞转染，实现基因修饰的干细胞体内MRI/OI双模式成像。通过考察荧光基团/SPIO纳米粒子组建模式和阳离子小分子修饰密度对干细胞的转染效率、体内双模式成像效果及其安全性能的影响，系统研究纳米自组装结构与成像功能和细胞转染效率之间的关系，揭示纳米材料与生物学响应之间的构效规律；为间充质干细胞抗癌治疗提供可靠的数据分析，为其临床推广打下坚实基础。

由于干细胞具备低免疫原性和分化性能，使得干细胞回输治疗成为多种疾病的备选治疗方案。但是，生物体内存在复杂生理/病理环境，加之现有临床方法无法对输入回体内的干细胞实现有效监测和功能评价，造成临床工作者难以获知具体细胞信息、不能做出治疗的预期判定。而影像学为观测干细胞在体内的行踪提供了一个新的解决途径。在本课题具体实施中，项目组设计并制备了系列可视化载体系统兼备基因传输功能，并完成了系统的优化与功能评价。研究中，我们构建了两亲性超支化大分子以及两亲性肽类大分子，借助亲疏水自组装功能包裹超顺磁性材料和光学探针，完成了双模态可视化载体的构建。我们分析了载体结构与影像效果之间关系，发现调节载体结构的可控团聚能够提高磁共振显影性能。并且，我们利用影像设备，动态、无创观察了干细胞在体内的行踪，获得了细胞迁移路径等重要信息。其次，借助两亲大分子化学结构上的的正电性氨基基团，可以有效结合微环DNA质粒，形成复合纳米粒子；并且依靠影像探针将转染与成像一体化。通过低分子量两亲性超支化大分子自组装构建的纳米/基因可视化复合载体，可以促进细胞内吞并高效转染细胞，并且转染效果可以借助磁共振、光学成像观察，实现了基因转染的多模态成像分析。在项目执行期间，本课题组邀请了几位影像学、药物递送相关领域的知名学者访问，此外，负责人及项目组成员多次参加国内外本领域专业学术会议并做报告。通过本项目的开展，发表 SCI 收录论文7篇，申请中国发明专利2项。上述研究为我们在后续课题中进一步开展更加深入系统的细胞治疗和纳米材料相互作用原理相关研究奠定了坚实的基础。