Therapeutic approaches to muscular dystrophy

Abstract

Muscular dystrophies are a heterogeneous group of genetic disorders characterized by muscle weakness and wasting. Duchenne muscular dystrophy (DMD) is the most common and severe form of muscular dystrophy, and although the molecular mechanisms of the disease have been extensively investigated since the discovery of the gene in 1986, there is currently no effective treatment. However, new gene-based therapies have recently emerged with particular noted advances in using conventional gene replacement strategies, RNA-based technology and pharmacological approaches. While the proof of principle has been demonstrated in animal models, several clinical trials have recently been undertaken to investigate the feasibility of these strategies in patients. In particular, antisense-mediated exon skipping has shown encouraging results and holds promise for the treatment of dystrophic muscle. Here, we summarize the recent progress in therapeutic approaches to muscular dystrophies, with an emphasis on gene therapy and exon skipping for DMD.

Figure 1.

Schematic representation of micro-dystrophin constructs. Full-length dystrophin consists of an actin binding domain (ABD1) at the N-terminus, four hinge domains (H1-4), 24 spectrin-like repeats (R1–R24) (within which lies a second ABD (ABD2)), a cysteine rich domain (CR) and the carboxy-terminal domain (CT). The CR itself consists of a WW domain, two EF-hand regions and a zinc finger domain (ZZ) that together form the dystroglycan binding site (122). Dystrophin also binds to α1-syntrophin and β1-syntrophin at the CT domain (123). Expression of the construct generated by deletion of exons 17–48 (Δ17–48), which was originally observed in a mildly affected Becker's muscular dystrophy patient, corrected 95% of the morphology in transgenic mdx mice and supports near normal force development (39,124). Later modification of this construct (ΔH2-R19) and one carrying deletions of R4–R23 (ΔR4–R23) have also been shown to reverse dystrophic pathology in transgenic mdx mice (41). Further truncation of dystrophin with deletion of the CT domain (ΔR4–R23/ΔCT) resulted in significant improvement in force in older dystrophic mice (45), confirming that the CT domain is not critical for dystrophin function. Recent work has now shown that replacing hinge 2 with hinge 3 in micro-dystrophin (ΔH2-R23 + H3/ΔCT) significantly improves the functional capacity of truncated dystrophins as demonstrated by prevention of muscle degeneration in mdx mice injected with rAAV6-ΔH2-R23 + H3/ΔCT (125). The size of a construct dictates which vectors can be used for delivery. The full-length cDNA only fits into gutted or high-capacity adenoviral vectors, the larger mini-dystrophins can be delivered with lentiviral vectors, while the smallest micro-dystrophins can be systemically delivered using rAAV.

Figure 2.

Antisense-mediated exon skipping rationale for DMD. (A) Patients with DMD have mutations which disrupt the open reading frame of the dystrophin pre-mRNA. In this example, exon 50 is deleted, creating an out-of-frame mRNA and leading to the synthesis of a truncated non-functional or unstable dystrophin (left panel). An antisense oligonucleotide directed against exon 51 can induce effective skipping of exon 51 and restore the open reading frame, therefore generating an internally deleted but partly functional dystrophin (right panel). (B) Multi exon-skipping rationale for DMD. The optimal skipping of exons 45–55 leading to the del45–55 artificial dystrophin could transform the DMD phenotype into the asymptomatic or mild BMD phenotype. This multiple exon skipping could theoretically rescue up to 63% of DMD patients with a deletion (126).