Myostatin is a negative regulator of muscle mass, and several strategies are being developed to knockdown its expression to improve muscle-wasting conditions. Strategies using antimyostatin-blocking antibodies, inhibitory-binding partners, signal transduction blockers, and RNA interference system (RNAi)-based knockdown have yielded promising results and increased muscle mass in experimental animals. These approaches have, however, a number of disadvantages such as transient effects or adverse immune complications. We report here the use of antisense oligonucleotides (AOs) to manipulate myostatin pre-mRNA splicing and knockdown myostatin expression. Both 2'O-methyl phosphorothioate RNA (2'OMePS) and phosphorodiamidate morpholino oligomers (PMO) led to efficient exon skipping in vitro and in vivo and knockdown of myostatin at the transcript level. The substantial myostatin exon skipping observed after systemic injection of Vivo-PMO into normal mice led to a significant increase in soleus muscle mass as compared to the controls injected with normal saline suggesting that this approach could be feasible to ameliorate muscle-wasting pathologies.
The knockdown of myostatin, a negative regulator of skeletal muscle mass may have
important implications in disease conditions accompanied by muscle mass loss like cancer,
HIV/AIDS, sarcopenia, muscle atrophy, and Duchenne muscular dystrophy (DMD). In DMD
patients, where major muscle loss has occurred due to a lack of dystrophin, the
therapeutic restoration of dystrophin expression alone in older patients may not be
sufficient to restore the functionality of the muscles. We recently demonstrated that
phosphorodiamidate morpholino oligomers (PMOs) can be used to re-direct myostatin splicing
and promote the expression of an out-of-frame transcript so reducing the amount of the
synthesized myostatin protein. Furthermore, the systemic administration of the same PMO
conjugated to an octaguanidine moiety (Vivo-PMO) led to a significant increase in the mass
of soleus muscle of treated mice. Here, we have further optimized the use of Vivo-PMO in
normal mice and also tested the efficacy of the same PMO conjugated to an arginine-rich
cell-penetrating peptide (B-PMO). Similar experiments conducted in mdx dystrophic mice
showed that B-PMO targeting myostatin is able to significantly increase the tibialis
anterior (TA) muscle weight and when coadministered with a B-PMO targeting the dystrophin
exon 23, it does not have a detrimental interaction. This study confirms that myostatin
knockdown by exon skipping is a potential therapeutic strategy to counteract muscle
wasting conditions and dual myostatin and dystrophin skipping has potential as a therapy
for DMD.
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