Chattong S, Ruangwattanasuk O, Yindeedej W, Setpakdee A, Manotham K
Molecular and Cell biology Unit, Department of Medicine, Lerdsin General Hospital, Bang-Rak Bangkok, Thailand.
S.S. Manufacturing Co., Ltd 114/7 Moo2 Mahasawadi, Nonthaburi, Thailand.
Gene Ther. 2017 Jul;24(7):425-432. doi: 10.1038/gt.2017.42. Epub 2017 May 22.
In humans, mutations in the β-globin gene (HBB) have two important clinical manifestations: β-thalassemia and sickle cell disease. The progress in genome editing and stem cell research may be relevant to the treatment of β-globin-related diseases. In this work, we employed zinc-finger nuclease (ZFN)-mediated gene integration of synthetic β-globin cDNA into HBB loci, thus correcting almost all β-globin mutations. The integration was achieved in both HEK 293 cells and isolated dental pulp stem cell (DPSCs). We also showed that DPSCs with an artificial gene knock-in were capable of generating stable six-cell clones and were expandable at least 10-fold; therefore, they may serve as a personalized stem cell factory. In addition, transfection with non-integrated pCAG-hOct4 and culturing in a conditioned medium converted the genome-edited DPSCs to CD34 HSC-like cells. We believe that this approach may be useful for the treatment of β-globin-related diseases, especially the severe form of β-thalassemia.
在人类中，β-珠蛋白基因（HBB）的突变有两种重要的临床表现：β-地中海贫血和镰状细胞病。基因组编辑和干细胞研究的进展可能与β-珠蛋白相关疾病的治疗有关。在这项工作中，我们采用锌指核酸酶（ZFN）介导的合成β-珠蛋白cDNA基因整合到HBB基因座中，从而纠正了几乎所有的β-珠蛋白突变。这种整合在HEK 293细胞和分离的牙髓干细胞（DPSC）中均得以实现。我们还表明，人工基因敲入的DPSC能够产生稳定的六细胞克隆，并且至少可以扩增10倍；因此，它们可作为个性化的干细胞工厂。此外，用未整合的pCAG-hOct4转染并在条件培养基中培养可将基因组编辑的DPSC转化为CD34造血干细胞样细胞。我们认为这种方法可能对β-珠蛋白相关疾病的治疗有用，尤其是严重形式的β-地中海贫血。
