Published January 5, 2022 | Version v1
Journal article Open

Comprehensive transcriptome-wide analysis of spliceopathy correction of myotonic dystrophy using CRISPR-Cas9 in iPSCs-derived cardiomyocytes

  • 1. Department of Gene Therapy & Regenerative Medicine, Vrije Universiteit Brussel, 1090 Brussels, Belgium
  • 2. Sorbonne Universit , Inserm, Institut de Myologie, Centre de Recherche en Myologie, F-75013 Paris, France
  • 3. Department of Gene Therapy & Regenerative Medicine, Vrije Universiteit Brussel, 1090 Brussels, Belgium - Center for Molecular & Vascular Biology, Department of Cardiovascular Sciences, University of Leuven, 3000 Leuven, Belgium



CTG repeat expansion (CTGexp) is associated with aberrant alternate splicing that contributes to cardiac dysfunction in myotonic dystrophy type 1 (DM1). Excision of this CTGexp repeat using CRISPR-Cas resulted in the disappearance of punctate ribonuclear foci in cardiomyocyte-like cells derived from DM1-induced pluripotent stem cells (iPSCs). This was associated with correction of the underlying spliceopathy as determined by RNA sequencing and alternate splicing analysis. Certain genes were of particular interest due to their role in cardiac development, maturation, and function (TPM4CYP2J2DMDMBNL3CACNA1HROCK2ACTB) or their association with splicing (SMN2GCFC2MBNL3). Moreover, while comparing isogenic CRISPR-Cas9-corrected versus non-corrected DM1 cardiomyocytes, a prominent difference in the splicing pattern for a number of candidate genes was apparent pertaining to genes that are associated with cardiac function (TNNTTNNT2TTNTPM1SYNE1CACNA1AMTMR1NEBLTPM1), cellular signaling (NCOR2CLIP1LRRFIP2CLASP1CAMK2G), and other DM1-related genes (i.e., NUMA1MBNL2LDB3) in addition to the disease-causing DMPK gene itself. Subsequent validation using a selected gene subset, including MBNL1MBNL2INSRADD3, and CRTC2, further confirmed correction of the spliceopathy following CTGexp repeat excision. To our knowledge, the present study provides the first comprehensive unbiased transcriptome-wide analysis of the differential splicing landscape in DM1 patient-derived cardiac cells after excision of the CTGexprepeat using CRISPR-Cas9, showing reversal of the abnormal cardiac spliceopathy in DM1.


Dastidar et al. 2021.pdf

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UPGRADE – Unlocking Precision Gene Therapy 825825
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