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#apaperaday: Biological and genetic therapies for the treatment of Duchenne muscular dystrophy

In today’s #apaperaday, Prof. Aartsma-Rus reads and comments on the paper titled: Biological and genetic therapies for the treatment of Duchenne muscular dystrophy

Today’s pick is an expert review from Wilton-Clark and Toshifumi Yokota on dystrophin restoring approaches from Expert Opinion on Biological Therapy  DOI: 10.1080/14712598.2022.2150543

Duchenne is caused by lack of dystrophin. This leads to a plethora of pathological pathways in skeletal muscle & heart. Restoring dystrophin makes sense as a therapeutic approach. These approaches are based on the fact that internally deleted dystrophins are partially functional.

So none of the dystrophin restoring approaches currently in (pre)clinical stages will restore the normal dystrophin (aside from exon skipping for single exon duplications). The authors discuss exon skipping, micro-dystrophin AAV, stop codon readthrough, genome editing…

But also cell therapy and utrophin upregulation. I will not discuss the latter 2 because: utrophin upregulation is back to square 1, and clinical trials never achieved to reach compound serum levels to increase utrophin levels in patients.

For cell therapy: current clinical trials with cell therapies from Capricor do not aim to restore dystrophin. Rather it is based on growth factors produced by transplanted heart stem cells, which have a positive effect and heart and skeletal muscle. Repeated treatment required.

So back to the real dystrophin restoring approaches:


1. Exon skipping aims to make the gene code readable for Duchenne patients so a partially functional dystrophin – as produced in Becker – can be produced. Exon skipping is achieved by antisense oligonucleotides (AONs)

4 AONs are approved in the USA (targeting exon 51, 45 & 53 (2) and 1 in Japan (targeting exon 53). These AONs all are of the PMO chemistry. Exon skipping is a mutation specific approach which is why AONs targeting different exons is required.

Each AON applies to only a subset of patients and the currently approved AONs apply to only ~30% of patients. Authors argue that using a cocktail of AONs would allow treatment of larger groups. e.g. skipping exons 45-55 would apply to over 60% of patients.

However (not discussed by authors), using such a cocktail would mean that part of the AONs are not effective. E.g. for a deletion of exon 48-50, the AONs targeting exon 48, 49 and 50 are not effective, as the patient does not have these exons. This is not allowed by regulators.

You cannot give compounds to patients with potential side effects when you know they will be ineffective. So that means you would have to customize the cocktail for each patient – making it again mutation specific, with the added burden of having to do extensive safety studies.

Authors mention that currently approved AONs show safety and efficacy of this approach. So far treatment indeed seems safe – despite the high doses used. However, I disagree about efficacy: we do not know yet that AONs slow down disease progression.

Even if the currently approved AONs slow down disease progression, there is room for improvement: AONs that are delivered better to muscle and lead to larger amounts of dystrophin. These are not discussed by the authors here (maybe due to space constraints)


2. AAV microdystrophin: AAV is the only viral vector delivering to muscle. However, it has limited capacity: only a microdystrophin – containing the bare minimum of functional domains – fits. Sarepta, Pfizer, Solid and (not mentioned in the review) Genethon are conducting trials.

The trials have revealed that microdystrophin can be restored in muscle after systemic treatment with AAV. However, it also leads to serious side effects and even death as authors rightly point out. So far it is not known whether microdystrophin will slow down disease progression.

Authors indicate AAV microdystrophin only applies to patients without preexisting immunity to AAV (agreed), & it is mutation independent (not true!) It has become clear that patients with N-terminal deletions of dystrophin are at risk of an anti-microdystrophin immune response.

This is because these patients will never have produced the actin-binding (N-terminal) part of dystrophin and therefore it acts as a neo-antigen. Patients with mutations in the center of the gene, will have made this region – dystrophin is not stable but immune cells have seen it.

Currently patients with deletions at the beginning of the dystrophin gene are excluded from micro-dystrophin AAV gene therapy trials by Pfizer and Sarepta as the risk of an auto-immune response is too severe.


3. Stop codon readthrough: this applies only to patients who have a substitution mutation where the code for an amino acid is changed into a stop signal. Read through suppression allows production of an almost normal dystrophin – albeit at low levels.

Stop codon readthrough compound ataluren (translarna) has conditional approval from the EMA in Europe since 2014. Confirmatory studies are needed. However, long term data from the STRIDE registry suggests that patients treated have up to 4 years later loss of ambulation. Authors discuss that this is an oral drug, while other approaches all are more invasive.


4. Genome editing (CRISPR Cas9). There are different ways this can be used

  1. to restore mutations: this does not work for muscle as it is post-mitotic (cells do not divide).
  2. creating errors: this can work to permanently reframe mutations. This works in cultured cells and animals. However, there are risks, i.e. cutting also at an untargeted location. Furthermore, delivery is still an issue. It requires AAV, which currently has safety concerns

We cannot increase the doses used for micro-dystrophin, while for genome editing likely we have to to deliver enough to all the muscles. Other challenges are the fact that Cas9 is a protein from bacteria and many people have a preexisting immune response to it.

Authors point out the less risky base editors that do not cut through the DNA. However, for now the efficiency is low. Towards the future authors indicate that AAV needs to be further improved so lower doses can be used that are more safe (agreed). CRISPR delivery with lipid nanoparticles would be more safe as it allows shorter expression of Cas9, so less risks of side effects. As authors correctly point out however, currently the delivery to muscle with these nanoparticles is very inefficient.


Authors do not discuss how delivery to muscle also needs to be approved for exon skipping AONs – I think this is also something that needs to be done. The challenge there will be to find a safe way to do this efficiently.

The paper gives a nice overview. I may have different views on things than the authors, but that is what happens with expert opinions: experts may have slightly different opinions 🙂