Cellular Delivery and Biological Activity of Antisense Oligonucleotides Conjugated to a Targeted Protein Carrier

Targeted delivery can potentially improve the pharmacological effects of antisense and siRNA oligonucleotides. Here we describe a novel bioconjugation approach to the delivery of splice-shifting antisense oligonucleotides (SSOs). The SSOs are linked to albumin via reversible S-S bonds. The albumin is also conjugated with polyethylene glycol (PEG) chains that terminate in an RGD ligand that selectively binds the αvβ3 integrin. As a test system we utilized human melanoma cells that express the αvβ3 integrin and that also contain a luciferase reporter gene that can be induced by delivery of SSOs to the cell nucleus. The RGD-PEG-SSO-albumin conjugates were endocytosed by the cells in an RGD-dependent manner; using confocal fluorescence microscopy evidence was obtained that the SSOs accumulate in the nucleus. The conjugates were able to robustly induce luciferase expression at concentrations in the 25−200nM range. At these levels little short-term or long-term toxicity was observed. Thus the RGD-PEG-Albumin conjugates may provide an effective tool for targeted delivery of oligonucleotides to certain cells and tissues