Disruption of splicing-regulatory elements using CRISPR/Cas9 to rescue spinal muscular atrophy in human iPSCs and mice

• 来源期刊: 国家科学评论（英文版）
• 关键词: spinal muscular atrophy SMN2 splicing-regulatory elements CRISPR/Cas9 germline correction
• 年，卷（期）: 2020,（1）
• 页码范围: 92-101
• 页数: 10页
• 语种: 英文
• DOI: 10.1093/nsr/nwz131

摘要

We here report a genome-editing strategy to correct spinal muscular atrophy (SMA).Rather than directly targeting the pathogenic exonic mutations,our strategy employed Cas9 and guide-sgRNA for the targeted disruption ofintronic splicing-regulatory elements.We disrupted intronic splicing silencers (ISSs,including ISS-N1 and ISS + 100) of survival motor neuron (SMN) 2,a key modifier gene of SMA,to enhance exon 7 inclusion and full-length SMN expression in SMA iPSCs.Survival of splicing-corrected iPSC-derived motor neurons was rescued with SMN restoration.Furthermore,co-injection of Cas9 mRNA from Streptococcus pyogenes (SpCas9) or Cas9 from Staphylococcus aureus (SaCas9) alongside their corresponding sgRNAs targeting ISS-N1 into zygotes rescued 56％ and 100％ of severe SMA transgenic mice (Smn-/-,SM-N2tg/-).The median survival of the resulting mice was extended to ＞400 days.Collectively,our study provides proof-of-principle for a new strategy to therapeutically intervene in SMA and other RNA-splicing-related diseases.