Challenges to oligonucleotides-based therapeutics for Duchenne muscular dystrophy

The development of the antisense oligonucleotides (AO)-based approach started in the late 1970's when the oligonucleotides were used as tools to downregulate the expression of specific genes [1]. The strategy was intuitive: oligonucleotides could be designed to hybridize with a specific mRNA target and mediate its destruction by RNaseH, an enzyme that destroys the RNA in a DNA/RNA complex. Attention rapidly increased with the development of antisense molecules for manipulation of alternative splicing. In this context, oligonucleotides can be used to modulate the ratio of splicing variants or correct splicing defects, which opened far-reaching implications in the treatment of a variety of diseases. The requirements for oligonucleotides that alter splicing are different from those for oligonucleotides used to achieve downregulation. In particular, they must not activate RNaseH, which would destroy the pre-mRNA before it could be spliced. They must also access their target pre-mRNAs within the nuclei of cells to efficiently compete with splicing factors. Several types of modified synthetic oligonucleotides fit these criteria. Among these, oligonucleotides with modifications to the 2' position, such as 2'-O-methyl (2'OMe), 2'-O-methoxyethyl (2'O-MOE) and 2'-O-aminopropyl, are RNaseH inactive and display higher nuclease resistance and affinity for target sequences than their 2'-deoxy counterparts. Similar characteristics are found in oligonucleotides with backbones based on morpholino, peptide nucleic acid (PNA), locked nucleic acid (LNA), phosphoramidate and methyl-phophonate derivatives. These advances in the development of antisense chemistries have led to numerous studies investigating the therapeutic potential of antisense technology (for review see [2]). However, despite early promise, the therapeutic application of AO has proved to be difficult and has been very slow entering the market and standard of care. Only a single AO compound has been approved by the Food