Department of Clinical Neurosciences, Laboratory of Cellular and Molecular Neurotherapies (LCMN), Lausanne University Hospital, 1011 Lausanne, Switzerland; Neurosciences Research Center (CRN), LCMN, Lausanne University Hospital, 1011 Lausanne, Switzerland.
CEA, DRF, Institut François Jacob, Molecular Imaging Research Center (MIRCen), F-92265 Fontenay-aux-Roses, France; Neurodegenerative Diseases Laboratory, CNRS, CEA, Université Paris-Sud, Université Paris-Saclay (UMR9199), F-92265 Fontenay-aux-Roses, France.
Cell Rep. 2017 Sep 19;20(12):2980-2991. doi: 10.1016/j.celrep.2017.08.075.
Neurodegenerative disorders are a major public health problem because of the high frequency of these diseases. Genome editing with the CRISPR/Cas9 system is making it possible to modify the sequence of genes linked to these disorders. We designed the KamiCas9 self-inactivating editing system to achieve transient expression of the Cas9 protein and high editing efficiency. In the first application, the gene responsible for Huntington's disease (HD) was targeted in adult mouse neuronal and glial cells. Mutant huntingtin (HTT) was efficiently inactivated in mouse models of HD, leading to an improvement in key markers of the disease. Sequencing of potential off-targets with the constitutive Cas9 system in differentiated human iPSC revealed a very low incidence with only one site above background level. This off-target frequency was significantly reduced with the KamiCas9 system. These results demonstrate the potential of the self-inactivating CRISPR/Cas9 editing for applications in the context of neurodegenerative diseases.
神经退行性疾病是一个主要的公共卫生问题，因为这些疾病的发病率很高。CRISPR/Cas9 系统的基因编辑使得修饰与这些疾病相关的基因序列成为可能。我们设计了 KamiCas9 自我失活编辑系统，以实现 Cas9 蛋白的瞬时表达和高效率的编辑。在第一个应用中，我们靶向了成人小鼠神经元和神经胶质细胞中的亨廷顿病（HD）相关基因。在 HD 的小鼠模型中，突变型亨廷顿蛋白（HTT）被有效地失活，导致疾病的关键标志物得到改善。用组成型 Cas9 系统对分化的人 iPSC 中的潜在脱靶位点进行测序，结果显示只有一个位点的脱靶频率高于背景水平，发生率非常低。而使用 KamiCas9 系统则显著降低了脱靶频率。这些结果表明，自我失活的 CRISPR/Cas9 编辑在神经退行性疾病的应用中具有很大的潜力。
