基于脑胶质瘤骨髓间充质干细胞靶向介导双基因治疗的可视化研究

Glioblastoma (GBM) is the most common and most aggressive primary brain tumor. Despite aggressive surgery, radiotherapy and chemotherapy, the prognosis of GBM patients remains poor. Gene therapy is increasingly being explored as a novel therapeutic option. It has been reported that bone marrow mesenchymal stem cells (BMSC), with the immunological compatibility, can target GBM cells by penetrating blood-brain barrier and blood-tumor barrier. In this study, a dual-regulatory element lentiviral vector (Lv-TIE-2-K5-Egr1-hNIS) will be constructed, which contains: 1) kringle 5 (K5) of human plasminogen, an angiogenesis inhibitor, driven by the promoter of TIE-2 gene that overexpressed in the tumor endothelial cells; 2) human sodium iodide symporter body (hNIS) gene, driven by a radiation-activated gene promoter, Egr1. The Lv-TIE-2-K5-Egr1-hNIS transduced BMSCs will be transplanted into the GBM-bearing nude mice and further be recruited to the tumor microenvironment, inducing the K5 overexpression. 188Re will then be injected intravenously, leading to the hNIS expression in BMSCs within the tumor by activation of the Egr1 promoter through β and γ rays. More expression of hNIS protein in tumor tissue will result in more tumor-spefic uptake of 188Re, forming a postive feedback. We expect that this dual-regulatory element lentivirus could incorparate the anti-angiogenesis therapy by K5 protein overexpression and the radiohtherapy by tumor-spefic 188Re-uptake via one single vector. Noninvasive 188Re micro-SPECT imaging will be used to assess the treatment response dynamically. If successful, this lentivirus-based gene therapy will provide a new adjuvant therapeutical option for GBM patients.

脑胶质瘤发病率高，预后差。临床常规手术及放化疗均不能显著提高患者生存率；而尽管存在自身免疫、血脑及血肿瘤屏障等问题，基因治疗仍是改善其预后的潜在有效新方法。骨髓间充质干细胞（BMSC）无免疫原性，可越过上述屏障向肿瘤迁移。本研究拟在人钠碘同向转运体（hNIS）基因的上游插入可被射线激活的Egr1启动子，同时加上由肿瘤新生血管特异性启动子（TIE-2）启动的抗肿瘤新生血管基因人纤溶酶原kringle 5（K5），构建一个同时表达hNIS和K5的重组慢病毒载体；以该病毒转染BMSC，尾静脉注入脑胶质瘤裸鼠模型，然后实时显像监测体内双基因治疗作用。转染BMSC表达K5，而由静脉给予发射β及γ射线的188Re激活Egr1启动hNIS表达，进而促进188Re摄取，形成hNIS-188Re摄取正反馈，获得188Re和K5蛋白的双重靶向抑瘤功能。项目研究获得结果将为脑胶质瘤辅助治疗及疗效评估提供新方法。

细胞介导的抗肿瘤基因治疗是实体肿瘤靶向治疗的新方法。骨髓间充质干细胞（BMSC）在脑胶质瘤靶向性上有独特的优势。本研究利用对辐射敏感的Egr1启动子可被射线诱导并激活启动其下游基因转录的特性，在人钠碘同向转运体（hNIS）基因上游连接Egr1启动子，同时应用TIE-2肿瘤新生血管特异性启动子启动抗肿瘤新生血管基因人纤溶酶原kringle 5（K5），构建慢病毒基因治疗载体，感染BMSC；将携带两种治疗基因的干细胞经静脉注入脑胶质瘤荷瘤裸鼠体内，干细胞靶向性聚集到肿瘤内后TIE-2启动子启动K5蛋白表达，达到抗肿瘤血管生成作用。同时，静脉给予同时发射β及γ射线的188Re，188Re激活Egr1启动子，启动下游的hNIS蛋白表达，hNIS蛋白表达的增高又进一步促进188Re摄取，形成正反馈效应，保证肿瘤组织摄取足量188Re，达到188Re和K5蛋白的双重靶向抑瘤作用。同时实时显像监测基因治疗疗效，为脑胶质瘤辅助治疗及疗效评估提供新方法。