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A Single CRISPR-Cas9 Deletion Strategy that Targets the Majority of DMD Patients
Restores Dystrophin Function in hiPSC-Derived Muscle Cells
#MMPMID26877224
Young CS
; Hicks MR
; Ermolova NV
; Nakano H
; Jan M
; Younesi S
; Karumbayaram S
; Kumagai-Cresse C
; Wang D
; Zack JA
; Kohn DB
; Nakano A
; Nelson SF
; Miceli MC
; Spencer MJ
; Pyle AD
Cell Stem Cell
2016[Apr]; 18
(4
): 533-40
PMID26877224
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Mutations in DMD disrupt the reading frame, prevent dystrophin translation, and
cause Duchenne muscular dystrophy (DMD). Here we describe a CRISPR/Cas9 platform
applicable to 60% of DMD patient mutations. We applied the platform to
DMD-derived hiPSCs where successful deletion and non-homologous end joining of up
to 725 kb reframed the DMD gene. This is the largest CRISPR/Cas9-mediated
deletion shown to date in DMD. Use of hiPSCs allowed evaluation of dystrophin in
disease-relevant cell types. Cardiomyocytes and skeletal muscle myotubes derived
from reframed hiPSC clonal lines had restored dystrophin protein. The internally
deleted dystrophin was functional as demonstrated by improved membrane integrity
and restoration of the dystrophin glycoprotein complex in vitro and in vivo.
Furthermore, miR31 was reduced upon reframing, similar to observations in Becker
muscular dystrophy. This work demonstrates the feasibility of using a single
CRISPR pair to correct the reading frame for the majority of DMD patients.
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