CRISPR/Cas系统定位修复血友病A外周血来源的iPSCs基因缺陷的研究

项目名称： CRISPR/Cas系统定位修复血友病A外周血来源的iPSCs基因缺陷的研究

项目编号： No.81470302

项目类型： 面上项目

立项/批准年度： 2015

项目学科： 医药、卫生

项目作者： 张磊

作者单位： 中国医学科学院

项目金额： 70万元

中文摘要： 血友病A是X染色体连锁的凝血因子VIII (FVIII)基因缺陷所致的遗传性出血性疾病，最有希望治愈血友病的途径是基因治疗。诱导多能干细胞（iPSCs）技术可以获得具有自身遗传背景的多潜能干细胞，而CRISPR/Cas9系统可以进行高效的定点基因缺陷修复，因此iPSCs技术联合CRISPR/Cas9技术进行血友病A基因治疗的研究具有良好的可行性和临床应用前景。本研究首先建立血友病A患者外周血来源的iPSCs，拟用CRISPR/Cas9技术对患者iPSCs的缺陷FVIII基因进行定点修复，将修复后的iPSCs定向分化为肝内皮细胞，检测其生成FVIII的能力和活性，并移植到免疫缺陷小鼠肝脏中检测产生人源性FVIII的能力。本研究使用iPSCs技术联合CRISPR/Cas9技术来研究血友病A基因定点修复治疗，为遗传性基因缺陷性疾病的临床基因治疗提供理论依据和技术支持。

中文关键词： 血友病A；基因治疗；诱导性多潜能干细胞

英文摘要： Hemophilia A is an X-linked hereditary hemorrhagic disease which is caused by defective synthesis of factor FVIII.Genetic therapy is the most hopeful way to cure hemophilia. Induced pluripotent stem cells (iPSCs) technique can obtain patients' stem cells of their own genetic background and the CRISPR/Cas9 system is a good tool for genetic repair in fixed positions in an efficient way. So the genetic therapy of hemophilia A by the combination of the iPSCs technique and the CRISPR/Cas9 technique has good feasibility and prospect for clinical application. Our research will firstly establish the iPSCs lines from the peripheral blood of hemophilia A and then use the CRISPR/Cas9 technique to repair the defected FVIII gene of the iPSCs in fixed position, and then differentiate the repaired iPSCs to hepatic endothelial cells. We will detect their ability of generating FVIII and the activity of FVIII. Then transplant the hepatic endothelial cells to the liver of immunodeficiency mice and detect its ability of generating human FVIII. This study use the CRISPR/Cas9 technique and the iPSCs technique to research the genetic therapy of hemophilia A, which will provide theoretical basis and technical support to the clinical genetic therapy of inherited genetic defective diseases.

英文关键词： hemophilia A;gene therapy;induced pluripotent stem cells