Lillehei Heart Institute, Department of Medicine, University of Minnesota, Minneapolis, MN 55455, USA.
Neurosciences Department, Biodonostia Research Institute-University of the Basque Country UPV-EHU, San Sebastián 20014, Spain; CIBERNED, Institute Carlos III, Madrid 28029, Spain.
Mol Ther. 2019 Dec 4;27(12):2147-2157. doi: 10.1016/j.ymthe.2019.08.011. Epub 2019 Aug 28.
Limb girdle muscular dystrophy type 2A (LGMD2A), caused by mutations in the Calpain 3 (CAPN3) gene, is an incurable autosomal recessive disorder that results in muscle wasting and loss of ambulation. To test the feasibility of an autologous induced pluripotent stem cell (iPSC)-based therapy for LGMD2A, here we applied CRISPR-Cas9-mediated genome editing to iPSCs from three LGMD2A patients to enable correction of mutations in the CAPN3 gene. Using a gene knockin approach, we genome edited iPSCs carrying three different CAPN3 mutations, and we demonstrated the rescue of CAPN3 protein in myotube derivatives in vitro. Transplantation of gene-corrected LGMD2A myogenic progenitors in a novel mouse model combining immunodeficiency and a lack of CAPN3 resulted in muscle engraftment and rescue of the CAPN3 mRNA. Thus, we provide here proof of concept for the integration of genome editing and iPSC technologies to develop a novel autologous cell therapy for LGMD2A.
肢带型肌营养不良 2A 型（LGMD2A）是一种由钙蛋白酶 3（CAPN3）基因突变引起的不可治愈的常染色体隐性疾病，会导致肌肉萎缩和行走能力丧失。为了测试基于自体诱导多能干细胞（iPSC）的治疗 LGMD2A 的可行性，我们在这里应用 CRISPR-Cas9 介导的基因组编辑技术对来自 3 名 LGMD2A 患者的 iPSC 进行了基因修正，以纠正 CAPN3 基因中的突变。我们使用基因敲入方法对携带三种不同 CAPN3 突变的 iPSC 进行了基因组编辑，并在体外证明了肌管衍生物中 CAPN3 蛋白的恢复。在一种将免疫缺陷和 CAPN3 缺乏相结合的新型小鼠模型中移植基因修正的 LGMD2A 成肌祖细胞，可导致肌肉植入和 CAPN3 mRNA 的恢复。因此，我们在这里提供了概念验证，证明了基因组编辑和 iPSC 技术的整合可以开发用于治疗 LGMD2A 的新型自体细胞疗法。
