In vivo genome editing in mouse restores dystrophin expression in Duchenne muscular dystrophy patient muscle fibers.
In vivo genome editing in mouse restores dystrophin expression in Duchenne muscular dystrophy patient muscle fibers.
复制标题
小鼠体内基因组编辑恢复杜氏肌营养不良症患者肌纤维中肌营养不良蛋白的表达
DOI：
10.1186/s13073-021-00876-0
复制
发表时间：
2021-04-12
期刊：
影响因子：
12.3
通讯作者：
Li X
中科院分区：
文献类型：
--
作者：
Chen M;Shi H;Gou S;Wang X;Li L;Jin Q;Wu H;Zhang H;Li Y;Wang L;Li H;Lin J;Guo W;Jiang Z;Yang X;Xu A;Zhu Y;Zhang C;Lai L;Li X
编码肌营养不良蛋白的DMD基因突变会导致Duchenne肌营养不良症(DMD)，这是最常见的致命遗传病。簇状规则间隔短回文重复(CRISPR)介导的基因编辑是一种有望永久治愈DMD的策略。在这项研究中，我们开发了一种新的策略，通过CRISPR介导的46-54外显子的大规模切除来重组DMD突变。通过使用DMD患者来源的原始肌肉来源干细胞(DMD-MDSCs)，我们将该方法与其他DMD抢救策略进行了比较。此外，通过将DMD-MDSCs移植到免疫缺陷小鼠体内，建立了患者来源的异种移植(PDX)DMD小鼠模型。通过腺相关病毒载体将CRISPR基因编辑组件肌肉注射到小鼠模型中。结果表明，大规模切除突变的DMD外显子可有效恢复Dystrophin蛋白的表达。我们还证实了来自Prevoella和Francisella 1(Cas12a)的CRISPR能够以与CRISPR相关蛋白9(Cas9)相同的效率纠正DMD突变。此外，在经大规模切除策略处理后的PDX DMD小鼠模型中，超过10%的人DMD肌纤维表达dystrophin。体内修复的dystrophin是功能性的，通过dystrophin糖蛋白复合体成员β-dystrolycan的表达证明了这一点。我们在PDX DMD小鼠模型中证明了临床相关的CRISPR/Cas9可以在活体内恢复人肌肉细胞中的dystrophin。这项研究为基因治疗在其他遗传性疾病中的应用提供了一种方法。网上版载有补充材料，可在10.1186/s13073-021-00876-0查阅。
Mutations in the DMD gene encoding dystrophin—a critical structural element in muscle cells—cause Duchenne muscular dystrophy (DMD), which is the most common fatal genetic disease. Clustered regularly interspaced short palindromic repeat (CRISPR)-mediated gene editing is a promising strategy for permanently curing DMD. In this study, we developed a novel strategy for reframing DMD mutations via CRISPR-mediated large-scale excision of exons 46–54. We compared this approach with other DMD rescue strategies by using DMD patient-derived primary muscle-derived stem cells (DMD-MDSCs). Furthermore, a patient-derived xenograft (PDX) DMD mouse model was established by transplanting DMD-MDSCs into immunodeficient mice. CRISPR gene editing components were intramuscularly delivered into the mouse model by adeno-associated virus vectors. Results demonstrated that the large-scale excision of mutant DMD exons showed high efficiency in restoring dystrophin protein expression. We also confirmed that CRISPR from Prevotella and Francisella 1(Cas12a)-mediated genome editing could correct DMD mutation with the same efficiency as CRISPR-associated protein 9 (Cas9). In addition, more than 10% human DMD muscle fibers expressed dystrophin in the PDX DMD mouse model after treated by the large-scale excision strategies. The restored dystrophin in vivo was functional as demonstrated by the expression of the dystrophin glycoprotein complex member β-dystroglycan. We demonstrated that the clinically relevant CRISPR/Cas9 could restore dystrophin in human muscle cells in vivo in the PDX DMD mouse model. This study demonstrated an approach for the application of gene therapy to other genetic diseases. The online version contains supplementary material available at 10.1186/s13073-021-00876-0.
登录
查看更多内容
DOI：
10.1038/mt.2015.220
发表时间：
2016-03
期刊：
Molecular therapy : the journal of the American Society of Gene Therapy
影响因子：
--
作者：
Bakondi B;Lv W;Lu B;Jones MK;Tsai Y;Kim KJ;Levy R;Akhtar AA;Breunig JJ;Svendsen CN;Wang S
通讯作者：
Wang S
影响因子：
2.4
作者：
Flanigan, Kevin M.
通讯作者：
Flanigan, Kevin M.
影响因子：
82.9
作者：
Charlesworth, Carsten T.;Deshpande, Priyanka S.;Porteus, Matthew H.
通讯作者：
Porteus, Matthew H.
影响因子：
3.5
作者：
Dunckley, MG;Manoharan, M;Dickson, G
通讯作者：
Dickson, G
DOI：
10.1038/mtna.2014.64
发表时间：
2014-12-02
期刊：
Molecular therapy. Nucleic acids
影响因子：
--
作者：
通讯作者：
--