干细胞来源的新型囊泡（exosome）靶向递送siRNA治疗脑胶质瘤的研究

In the previous studies, we successfully constructed the non-viral delivery system and tranfected the tumor therapeutic cytokines to the mesenchymal stem cells (MSCs). Our studies showed the ability of mesenchymal stem cells (MSCs) to migrate towards lung tumor sites, which makes them a great hope for efficient targeted-delivery vehicles in cancer gene therapy. However, the entrapment of MSCs by lung tissue after i.v. injection and their potential in vivo toxicity limited its further applications. Therefore, based on the previous studies and the project supported by National Nature Science Foundation of China, we would like to develop a novel delivery system composed of exosomes extracted from MSCs, combined with the non-viral transfection technology. General speaking, to enhance the tumor homing ability of exosomes, the MSCs were firstly transduced with CXCR4, a cytokine which tend to migrate towards tumor site, followed by the extraction of exosomes from MSCs (MSC-exosomes). SiRNA was loaded to the MSC-exosomes by electroporation. And then, the MSC-exosomes were injected to the glioma bearing mice by tail vein.The development of MSC-exosomes as a novel targeted delivery vehicle will not only provide a novel method for the glioma treatment, but also has important significance both for the theory and practice of the targeting gene delivery systems. In the future, these strategies could be extended by using various genes to achieve a general exosome-based gene therapy platform for the treatment of different disease.

在前期研究中，我们成功实现了干细胞作为靶向载体对肺转移瘤的治疗作用。然而，研究发现，干细胞作为细胞载体，存在其内在的缺陷，如静脉注射后大多被肺截留，体内潜在的毒副作用及难以储存等问题。因此，本研究在青年科学基金工作的研究基础上，基于exosome能透过血脑屏障的特性，对干细胞中提取的exosomes进行研究，并结合非病毒载体转染技术及RNA干扰技术，提出了使用基因修饰的干细胞分泌的exosomes（MSC-exosomes）作为载体靶向递送siRNA治疗脑胶质瘤的研究思路，国内外未见报道。研究首先用非病毒载体将趋化因子受体CXCR4导入干细胞并提取exosomes，该MSC-exosomes用于携载siRNA，并将其静脉注射入荷瘤鼠体内用以治疗脑胶质瘤。该研究的开展，不仅扩展了肿瘤靶向治疗的研究领域，也为MSC-exosomes作为一种新型靶向载体的临床应用奠定了基础。

在前期研究中，我们成功实现了干细胞作为靶向载体对肺转移瘤的治疗作用。然而，研究发现，干细胞作为细胞载体，存在其内在的缺陷，如静脉注射后大多被肺截留，体内潜在的毒副作用及难以储存等问题。因此，本研究在青年科学基金工作的研究基础上，基于exosome能透过血脑屏障的特性，对干细胞中提取的exosomes进行研究。本项目制备合成普鲁兰多糖-精胺阳离子聚合物、screenfect、低分子量壳聚糖-聚乙烯亚胺等作为非病毒基因转染载体，并考察了干细胞上不同非病毒载体的基因导入效率和细胞毒性。随之，也构建了病毒载体，并比较了非病毒载体和慢病毒载体在干细胞上对CXCR4基因的转染效率。经过对比，在后期实验中，将病毒载体作为转染载体，构建稳定表达CXCR4的干细胞株，并将肿瘤坏死因子相关的凋亡诱导配体TRAIL导入稳定表达CXCR4的干细胞株，制备行CXCR4＋TRAIL＋exosomes，并对乳腺癌细胞脑转移小鼠模型进行治疗，证明了治疗组具有较好的肿瘤杀伤效果。对exosomes作为新型靶向载体的用于脑转移瘤模型的治疗研究进一步拓展了肿瘤靶向治疗的研究领域，使MSC-exosomes能够成为广泛使用的基因靶向载体，携载不同的治疗基因，实现对肿瘤的基因靶向治疗，为其在临床的普遍应用奠定基础。