(176ap) Effect of Delivery Vehicle Surface Charge on the Intracellular Trafficking of Sirnas

While the use of short interfering RNAs (siRNAs) for laboratory studies is now common practice, development of siRNAs for therapeutic applications has slowed, due in part to a still limited understanding of the intracellular trafficking of siRNA-containing complexes. As a result, it is difficult to design delivery vehicles for specific cell types, resulting in inefficient delivery, cytotoxicity, or immunogenicity when used in vivo. Our goal is to define the impact of nanoparticle characteristics on the intracellular trafficking of siRNAs. Using silica nanoparticles (sNPs) for their tunable synthesis, we are investigating the effects of surface charge on four processes: membrane translation, endosomal escape, endolysosomal retention, and sequence specific mRNA degradation (silencing).

The presentation will discuss our novel live cell assay that allows independent intracellular trafficking of the delivery vehicle, antisense siRNA strand, and sense siRNA strand. Using confocal microscopy, we track vehicle-siRNA complexes during endocytosis, endosomal trafficking (Early, Late, Recycling, and Lysosome), trafficking to the endoplasmic reticulum or golgi, and exocytosis. Our results to date demonstrate that optimal siRNA delivery occurs when the surface charge of delivery vehicles promotes release of siRNAs in the late endosome. Delivery vehicles with weak surface charge are prone to endosomal recycling, whereas strong surface charge promotes endolysosomal retention.