CRISPR-Cas9 Gene Therapy for Duchenne Muscular Dystrophy

Abstract

Discovery of the CRISPR-Cas (clustered regularly interspaced short palindromic repeat, CRISPR-associated) system a decade ago has opened new possibilities in the field of precision medicine. CRISPR-Cas was initially identified in bacteria and archaea to play a protective role against foreign genetic elements during viral infections. The application of this technique for the correction of different mutations found in the Duchenne muscular dystrophy (DMD) gene led to the development of several potential therapeutic approaches for DMD patients. The mutations responsible for Duchenne muscular dystrophy mainly include exon deletions (70% of patients) and point mutations (about 30% of patients). The CRISPR-Cas 9 technology is becoming increasingly precise and is acquiring diverse functions through novel innovations such as base editing and prime editing. However, questions remain about its translation to the clinic. Current research addressing off-target editing, efficient muscle-specific delivery, immune response to nucleases, and vector challenges may eventually lead to the clinical use of the CRISPR-Cas9 technology. In this review, we present recent CRISPR-Cas9 strategies to restore dystrophin expression in vitro and in animal models of DMD.

Guanidinium-Rich Lipopeptide-Based Nanoparticle Enables Efficient Gene Editing in Skeletal Muscles

“…40 The mdx mice were designed to carry a nonsense point mutation (C-to-T transition) in exon 23 that aborted full-length dystrophin expression. 41 Delivery of RNP together with a repair template targeting this region could disrupt the point mutation or correct the gene via nonhomologous end joining (NHEJ) or HDR 2,42,43 processes. Therefore, the dystrophin expression can be recovered to restore the muscle strength.…”

Therapeutic Strategies for Dystrophin Replacement in Duchenne Muscular Dystrophy

“…These modifications include the correction of nonsense mutations, the insertion of one or multiple nucleotides to reframe a frameshift mutation, the modification of splicing sites to favor exon skipping and reframing, etc. ( 58 ).…”

“… Clustered Regularly Interspaced Short Palindromic Repeats approaches. This figure adapted from Happi Mbakam et al ( 58 ) represents the principle of CRISPR-Cas9 gene modification, which uses a guide RNA and a Cas9 to mediate the nuclease activity resulting in insertions, deletions, or INDELs (A) . Panel (B) represents the general mechanism for base editing, which uses a sgRNA (spacer sequence + scaffold) and a Cas9 nickase fused with a cytosine or an adenine deaminase.…”

“…Koo et al also reported up to 8 ± 0.7% INDELs. Happi Mbakam et al recently presented an update of the CRISPR-Cas9 gene therapy for dystrophin replacement in DMD ( 58 ).…”

Prime editing optimized RTT permits the correction of the c.8713C>T mutation in DMD gene

“… 6 , 7 Different types of mutations leading to DMD have been identified in the DMD gene, which is one of the biggest human genes, 8 and different therapeutic strategies have been developed. 9 , 10 These mutations include exonic and intronic duplications accounting for 10%–15% of DMD mutations, small insertions and deletions (3%), point mutations (nonsense and missense mutations, splice site mutations, and mid intronic mutations, 26%) and single- or multi-exon deletions (60%–70%). 7 , 11 Many research groups have been using CRISPR-Cas9 genome editing to modify the DMD gene to restore the dystrophin expression.…”