Targeted delivery of a splice-switching oligonucleotide by cationic polyplexes of RGD-oligonucleotide conjugate

Nanoparticle-based delivery has become an important strategy to advance therapeutic oligonucleotides into clinical reality. Delivery by nanocarriers can enhance access of oligonucleotides to their pharmacological targets within cells; preferably, targeting ligands are incorporated into nanoparticles for targeting oligonucleotides to disease sites, often by conjugation to delivery carriers. In this study, a splice-switching oligonucleotide (SSO) was conjugated to a bivalent RGD peptide, and then, the RGD-SSO conjugate was formulated into polyplexes with a cationic polymer polyethylenimine. The resultant polyplexes of RGD-oligonucleotide conjugate demonstrated dramatic increase in the pharmacological response of splicing correction compared to free RGD-SSO conjugate or the polyplexes of unconjugated SSO, through integrin-mediated endocytosis and rapid endosomal release. This study has shown that coupling a targeting ligand to cargo oligonucleotide can maintain the integrin targeting ability after the peptide-oligonucleotide conjugate is complexed with cationic polymer. Preliminary study also revealed that integrin targeting redirects intracellular trafficking of the polyplexes to caveolar pathway and thereby generates greater effectiveness of the oligonucleotide. This study provides a new platform technology to construct multifunctional delivery systems of therapeutic oligonucleotides.