Use of cell-penetrating-peptides in oligonucleotide splice switching therapy

The hydrophobic plasma membrane constitutes an indispensable barrier for cells, allowing influx of essential molecules while preventing access to other macromolecules. Although pivotal for the maintenance of cells, the inability to cross the plasma membrane is one of the major obstacles toward current drug development. Oligonucleotides (ONs) are a group of substances that display great therapeutic potential to interfere with gene expression. Several classes of ONs have emerged either based on double stranded RNAs, such as short interfering RNAs that are utilized to confer gene silencing, or single stranded ONs of various chemistries for antisense targeting of small regulatory micro RNAs or mRNAs. In particular the use of splice switching oligonucleotides (SSOs) to manipulate alternative splicing, by targeting pre-mRNA, has proven to be a highly promising therapeutic strategy to treat various genetic disorders, including Duchenne muscular dystrophy and spinal muscular atrophy. Despite being efficient compounds to alter splicing patterns, their hydrophilic macromolecular nature prohibits efficient cellular internalization.Various chemical drug delivery vehicles have been developed aiming at improving the bioavailability of nucleic acid-based drugs. In the context of SSOs, one group of peptidebased delivery vectors, i.e. cell-penetrating peptides (CPPs), display extremely high potency. CPPs have a remarkable ability to convey various, otherwise impermeable, macromolecules across the plasma membrane of cells in a relatively non-toxic fashion. This review provides insight into the application of CPPs and ONs in gene regulation with particular focus on CPP-assisted delivery of therapeutic SSOs.