Therapeutic Genome Editing for Myotonic Dystrophy Type 1 Using CRISPR/Cas9

Yanlin Wang, Lei Hao, Hongcai Wang, Katherine Santostefano, Arjun Thapa, John Cleary, Hui Li, Xiuming Guo, Naohiro Terada, Tetsuo Ashizawa, Guangbin Xia

Research output: Contribution to journalArticle

15 Scopus citations

Abstract

Myotonic dystrophy type 1 (DM1) is caused by a CTG nucleotide repeat expansion within the 3′ UTR of the Dystrophia Myotonica protein kinase gene. In this study, we explored therapeutic genome editing using CRISPR/Cas9 via targeted deletion of expanded CTG repeats and targeted insertion of polyadenylation signals in the 3′ UTR upstream of the CTG repeats to eliminate toxic RNA CUG repeats. We found paired SpCas9 or SaCas9 guide RNA induced deletion of expanded CTG repeats. However, this approach incurred frequent inversion in both the mutant and normal alleles. In contrast, the insertion of polyadenylation signals in the 3′ UTR upstream of the CTG repeats eliminated toxic RNA CUG repeats, which led to phenotype reversal in differentiated neural stem cells, forebrain neurons, cardiomyocytes, and skeletal muscle myofibers. We concluded that targeted insertion of polyadenylation signals in the 3′ UTR is a viable approach to develop therapeutic genome editing for DM1. Myotonic dystrophy type 1 is caused by toxic RNAs with expanded CUG repeats in the 3′ UTR of the DMPK gene. Wang et al. developed a strategy to eliminate the toxic repeats by insertion of polyadenylation signals upstream of the expanded repeats for personalized cell-based therapy and in vivo therapeutic genome editing.

Original languageEnglish (US)
Pages (from-to)2617-2630
Number of pages14
JournalMolecular Therapy
Volume26
Issue number11
DOIs
StatePublished - Nov 7 2018

Keywords

  • CRISPR/Cas9
  • DMPK
  • SaCas9
  • SpCas9 nickase
  • genome editing
  • induced pluripotent stem cell
  • myotonic dystrophy
  • nucleotide repeat expansion
  • polyadenylation
  • stem cell

ASJC Scopus subject areas

  • Molecular Medicine
  • Molecular Biology
  • Genetics
  • Pharmacology
  • Drug Discovery

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