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

Mol Ther. 2018 Nov 7;26(11):2617-2630. doi: 10.1016/j.ymthe.2018.09.003. Epub 2018 Sep 11.


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.

Keywords: CRISPR/Cas9; DMPK; SaCas9; SpCas9 nickase; genome editing; induced pluripotent stem cell; myotonic dystrophy; nucleotide repeat expansion; polyadenylation; stem cell.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't

MeSH terms

  • 3' Untranslated Regions
  • CRISPR-Cas Systems / genetics
  • Cell Differentiation / genetics
  • Gene Editing / methods
  • Genetic Therapy / methods
  • HEK293 Cells
  • Humans
  • Muscle, Skeletal / growth & development
  • Myocytes, Cardiac / physiology
  • Myotonic Dystrophy / genetics*
  • Myotonic Dystrophy / pathology
  • Myotonic Dystrophy / therapy
  • Myotonin-Protein Kinase / genetics*
  • Neural Stem Cells / physiology*
  • Neurons / physiology
  • RNA 3' Polyadenylation Signals / genetics
  • RNA, Guide, CRISPR-Cas Systems
  • Transfection
  • Trinucleotide Repeat Expansion / genetics*


  • 3' Untranslated Regions
  • DMPK protein, human
  • RNA, Guide, CRISPR-Cas Systems
  • Myotonin-Protein Kinase