基于iPSC和CRISPR技术研究进行性骨化性纤维发育不良的基因治疗

Fibrodysplasia ossificans progressiva (FOP) is a seriously disabling even mortiferous rare inherited bone disease, mostly in an autosomal dominant pattern of pathogenesis. Two clinical features characterized classic FOP patient are skeletal malformations and progressive heterotopic ossification. The heterotopic ossification in connective tissue can lead to immobility of joints, and then developed into a lifelong disability even the death of FOP patients. In our previous studies, we clinically collected 20 more FOP patients in chinese population, and the gene test results showed that most of them were identified ACVR1 gene mutation c.617G>A (p.R206H). Although, the molecular mechanisms underlying FOP pathogenesis achieved some progress, but so far there is no effecitve the treatment of FOP. In this research project, we will use induced pluripotent stem cell (iPSC) technology to construct FOP disease model (ACVR1-p.R206H), and clustered regularly interspaced short palindromic repeat (CRISPR-Cas9) technology was utlized to precisely repair the ACVR1 gene mutation c.617G>A (p.R206H) in FOP iPSC. By comparing the differentiation efficiency and behavior of FOP-iPSC and repaired FOP-iPSC derived bone-forming progenitor cells as endothelial cells and mesenchymal cells, we attempt to restore the function of the progenitor cells in FOP. This project will provide experimental basis for the in vivo gene therapy of FOP, and accumulate experience for the gene therapy of genetic diseases.

进行性骨化性纤维发育不良(FOP)是一种致残致死的罕见遗传性骨病，多呈常染色体显性方式发病。典型FOP患者的临床特征主要表现为骨骼畸形和进行性异位骨化。结缔组织的异位骨化可导致关节功能丧失，进而发展成终生残疾甚至死亡。课题组前期临床收集20多例中国人群FOP患者，基因检测结果显示大部分病例为基因ACVR1点突变c.617G>A (p.R206H)。目前尽管对该基因突变导致FOP的分子机制有所研究，但尚无有效的治疗方法。本研究拟利用诱导多能干细胞(iPSC)技术构建FOP疾病模型(ACVR1-p.R206H)，并利用CRISPR-Cas9技术精确修复FOP-iPSC中的突变。比较FOP-iPSC和修复后FOP-iPSC分化所得内皮细胞、间质细胞等骨形成前体细胞的分化效率和行为变化，以期恢复FOP中骨形成前体细胞的功能。本项目将为FOP体内基因治疗提供实验依据，为遗传病的基因治疗积累研究经验。

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