Enhancing microRNA activity through increased endosomal release mediated by nigericin

Abstract

The therapeutic promise of small RNA therapeutics (siRNAs, miRNAs) is not only limited by the lack of delivery vehicles, but also by the inability of the small RNAs to reach intracellular compartments where they can be biologically active. We previously reported successful delivery of functionally active miRNAs via receptor-mediated endocytosis 1 . This type of targeted therapy still faces one of the major challenges in the delivery field, endosomal sequestration. Here, a new method has been developed to promote endosomal escape of delivered miRNA. The strategy relies on the difference in solute contents between nascent endosomes and the cytoplasm: early endosomes are rich in sodium ions (Na + ) while the intracellular fluid is rich is potassium ions (K + ). Exploiting this difference through favoring the influx of K + into the endosomes without the exchange for a osmotically active ion (Na + ), results in an osmotic differential leading to endosome swelling and bursting. One molecule that is able to exchange K + for an osmotically inactive hydrogen ion is the ionophore nigericin. Via generating an intramolecular miRNA delivery vehicle, containing a ligand, in this case folate, and nigericin we achieve escape of folate-RNA conjugates (e.g. FolamiRs) from their entrapping endosomes into the cytoplasm where they bind the RNA Induced Silencing complex (RISC) and activate the RNAi response.

CHAPTER 14. Strategies for Safe and Targeted Delivery of MicroRNA Therapeutics

“…Various methods to promote endosomal escape and enhance efficiency of delivered miRNAs are being evaluated. 161 Additional targeting ligands that bind to other surface receptors have been developed for RNA delivery. Using the N-acetylgalactosamine (galNAc) ligand, an RNA prodrug was efficiently delivered to the liver, which highly expresses the galNAc receptor asialoglycoprotein (ASgpR).…”