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<p>Dystrophy-associated fer-1-like protein (dysferlin) conducts plasma membrane repair. Mutations in the <i>DYSF</i> gene cause a panoply of genetic muscular dystrophies. We targeted a frequent loss-of-function, <i>DYSF</i> exon 44, founder frameshift mutation with mRNA-mediated delivery of SpCas9 in combination with a mutation-specific sgRNA to primary muscle stem cells from two homozygous patients. We observed a consistent &gt;60% exon 44 re-framing, rescuing a full-length and functional dysferlin protein. A new mouse model harboring a humanized <i>Dysf</i> exon 44 with the founder mutation, hEx44mut, recapitulates the patients’ phenotype and an identical re-framing outcome in primary muscle stem cells. Finally, gene-edited murine primary muscle stem-cells are able to regenerate muscle and rescue dysferlin when transplanted back into hEx44mut hosts. These findings are the first to show that a CRISPR-mediated therapy can ameliorate dysferlin deficiency. We suggest that gene-edited primary muscle stem cells could exhibit utility, not only in treating dysferlin deficiency syndromes, but also perhaps other forms of muscular dystrophy.</p>

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Gene-editing in patient and humanized-mice primary muscle stem cells rescues dysferlin expression in dysferlin-deficient muscular dystrophy

  • 期刊论文

  • 出版模式：

    开放获取

  • 发表日期：

    2025年1月2日

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  • Helena Escobar,
  • Silvia Di Francescantonio,
  • Julia Smirnova,
  • Robin Graf,
  • Stefanie Müthel,
  • Andreas Marg,
  • Alexej Zhogov,
  • Supriya Krishna,
  • Eric Metzler,
  • Mina Petkova,
  • Oliver Daumke,
  • Ralf Kühn,
  • Simone Spuler

摘要

Dystrophy-associated fer-1-like protein (dysferlin) conducts plasma membrane repair. Mutations in the DYSF gene cause a panoply of genetic muscular dystrophies. We targeted a frequent loss-of-function, DYSF exon 44, founder frameshift mutation with mRNA-mediated delivery of SpCas9 in combination with a mutation-specific sgRNA to primary muscle stem cells from two homozygous patients. We observed a consistent >60% exon 44 re-framing, rescuing a full-length and functional dysferlin protein. A new mouse model harboring a humanized Dysf exon 44 with the founder mutation, hEx44mut, recapitulates the patients’ phenotype and an identical re-framing outcome in primary muscle stem cells. Finally, gene-edited murine primary muscle stem-cells are able to regenerate muscle and rescue dysferlin when transplanted back into hEx44mut hosts. These findings are the first to show that a CRISPR-mediated therapy can ameliorate dysferlin deficiency. We suggest that gene-edited primary muscle stem cells could exhibit utility, not only in treating dysferlin deficiency syndromes, but also perhaps other forms of muscular dystrophy.