A universal gene correction approach for FKRP-associated dystroglycanopathies to enable autologous cell therapy

Neha R. Dhoke, Hyunkee Kim, Sridhar Selvaraj, Karim Azzag, Haowen Zhou, Nelio A.J. Oliveira, Sudheer Tungtur, Carolina Ortiz-Cordero, James Kiley, Qi Long Lu, Anne G. Bang, Rita C.R. Perlingeiro

Research output: Contribution to journalArticlepeer-review

Abstract

Mutations in the fukutin-related protein (FKRP) gene result in a broad spectrum of muscular dystrophy (MD) phenotypes, including the severe Walker-Warburg syndrome (WWS). Here, we develop a gene-editing approach that replaces the entire mutant open reading frame with the wild-type sequence to universally correct all FKRP mutations. We apply this approach to correct FKRP mutations in induced pluripotent stem (iPS) cells derived from patients displaying broad clinical severity. Our findings show rescue of functional α-dystroglycan (α-DG) glycosylation in gene-edited WWS iPS cell-derived myotubes. Transplantation of gene-corrected myogenic progenitors in the FKRPP448L-NSG mouse model gives rise to myofiber and satellite cell engraftment and, importantly, restoration of α-DG functional glycosylation in vivo. These findings suggest the potential feasibility of using CRISPR-Cas9 technology in combination with patient-specific iPS cells for the future development of autologous cell transplantation for FKRP-associated MDs.

Original languageEnglish (US)
Article number109360
JournalCell reports
Volume36
Issue number2
DOIs
StatePublished - Jul 13 2021

Bibliographical note

Funding Information:
This project was supported by NIH grants R01 AR071439 and AR055299 (to R.C.R.P.) and the LGMD2I Research Funds (to R.C.R.P. and A.G.B.). We are grateful to Jiri Vajsar for providing the WWS patient sample. We thank Lila Habib for her contribution in the generation of WWS iPS cells. The monoclonal antibody to MHC and the IIH6 antibody were obtained from the Developmental Studies Hybridoma Bank, developed under the auspices of the NICHD and maintained by the University of Iowa. The authors are grateful to Cynthia Faraday for graphic design.

Funding Information:
This project was supported by NIH grants R01 AR071439 and AR055299 (to R.C.R.P.) and the LGMD2I Research Funds (to R.C.R.P. and A.G.B.). We are grateful to Jiri Vajsar for providing the WWS patient sample. We thank Lila Habib for her contribution in the generation of WWS iPS cells. The monoclonal antibody to MHC and the IIH6 antibody were obtained from the Developmental Studies Hybridoma Bank, developed under the auspices of the NICHD and maintained by the University of Iowa. The authors are grateful to Cynthia Faraday for graphic design. N.R.D. designed and performed the experiments, analyzed the data, and wrote the manuscript. H.K. S.S. and K.A. designed and performed the experiments and analyzed the data. H.Z. N.A.J.O. S.T. and J.K. performed the experiments and analyzed the data. C.O.-C. contributed to design and data analysis. Q.L.L. and A.G.B. provided the reagents and contributed to the interpretation of the data and the writing of the manuscript. R.C.R.P. contributed to the experimental design, interpretation of the data, and writing of the manuscript. The authors declare no competing interests.

Publisher Copyright:
© 2021 The Author(s)

Keywords

  • CRISPR-Cas9
  • FKRP mutations
  • gene editing
  • muscle engraftment
  • patient-specific iPS cells
  • phenotype rescue
  • satellite cell engraftment
  • universal correction
  • WWS
  • α-dystroglycan

PubMed: MeSH publication types

  • Journal Article

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