Deep Parikh

Background: Duchenne Muscular Dystrophy (DMD) is a lethal neuromuscular disorder that affects almost 1 in 3500 males and is seen in boys around the ages 3-4. The disease symptoms progress very quickly following this.¹ DMD is an X-linked disorder that occurs due to mutations of the dystrophin gene on chromosome XP21.² In 70% of cases, DMD is inherited. In the other 30%, however, a de novo mutation occurs.¹ DMD presents with many clinical symptoms in the muscular, cardiac, and respiratory systems.² The incidence of DMD is 15.9 cases per 100,000 births in the United States. Deletions or duplications account for about 70-80% of gene mutations in DMD.² Dysfunctional dystrophin leads to the breakdown of muscle and rapid tissue degeneration. Muscle necrosis leads to reduced myofibers that are replaced with connective and adipose tissue. All current medications are targeted at being palliative to ease the symptoms of those with DMD. Glucocorticoids are one of the most widely used medications for DMD which slow the degeneration of myofibers.²

Objective: The use of exon skipping to restore dystrophin production in DMD patients with specific exon mutations was explored.

Search Methods: The PubMed database was utilized to conduct a search on DMD and gene therapy from 2017 to 2023 using keywords such as “DMD gene therapy”and “exon skipping”.

Results: Exon skipping is a technique in which the mutated part of a gene is skipped in order to restore the reading frame and allow the production of a partially functional protein. Limbe-girdle muscular dystrophy 2C (LGMD2C) is a muscular dystrophy very similar to DMD but has a mutation in the SGCG gene instead which produces sarcoglycan gamma protein. A mouse model of LGMD2C was engineered with CRISPR/CAS9 by deleting a single thymine from SGCG exon 6 (521DT).³ Phosphorodiamidate morpholino oligomers (PMO) were used for exon skipping in the 521DT mice. This therapy specifically targeted exon 6 in both cell cultures and in mice. Results showed that there was the production of a partially functional sarcoglycan gamma protein.⁴ These experiments can be translated to treat DMD as well as displayed by the drug Casimersen. Casimersen skips exon 45 during RNA splicing, therefore, can treat DMD patients with a mutation in exon 44. A phase III clinical trial was conducted on 43 patients with a confirmed DMD exon 44 mutation. All the patients were male aged 7-21. It was found that Casimersen led to an increase in dystrophin production in 88% of the patients. There was also a significant increase seen in the distance walked by patients in 6 minutes, and no serious adverse events were noted.⁵ Exon skipping can also be enhanced in combination with other techniques such as FORCE conjugation in which a target molecule such as TfR1 is attached to the PMO to increase uptake by the cells. It was found that FORCE conjugation with TfR1 increased cellular uptake and enhanced exon skipping, therefore, increasing the dystrophin production and also led to prolonged effects.⁶

Conclusions: Experimental studies and clinical trials have displayed exon skipping can be an effective therapy to treat specific DMD mutations. Studies have also shown how exon skipping can be enhanced to further treatment and create prolonged effects.

Works Cited

1. Venugopal V, Pavlakis S. Duchenne Muscular Dystrophy. [Updated 2022 Jul 11]. In: StatPearls [Internet]. Treasure Island (FL): StatPearls Publishing; 2022 Jan-. Available from: https://www.ncbi.nlm.nih.gov/books/NBK482346/
2. Basta M, Pandya AM. Genetics, X-Linked Inheritance. In: StatPearls. Treasure Island (FL): StatPearls Publishing; May 8, 2022.
3. Demonbreun AR, Wyatt EJ, Fallon KS, et al. A gene-edited mouse model of limbgirdle muscular dystrophy 2C for testing exon skipping. Dis Model Mech. 2019;13(2):dmm040832. Published 2019 Nov 4. doi:10.1242/dmm.040832
4. Wyatt EJ, Demonbreun AR, Kim EY, et al. Efficient exon skipping of SGCG mutations mediated by phosphorodiamidate morpholino oligomers. JCI Insight. 2018;3(9):e99357. Published 2018 May 3. doi:10.1172/jci.insight.99357
5. Wilton-Clark H, Yokota T. Casimersen for Duchenne muscular dystrophy. Drugs Today (Barc). 2021;57(12):707-717. doi:10.1358/dot.2021.57.12.3352740
6. Desjardins CA, Yao M, Hall J, et al. Enhanced exon skipping and prolonged dystrophin restoration achieved by TfR1-targeted delivery of antisense oligonucleotide using FORCE conjugation in mdx mice. Nucleic Acids Res. 2022;50(20):11401-11414. doi:10.1093/nar/gkac641