(549c) Analysis of siRNA Delivery by Polymeric Nanoparticles

Clinical applications of genetic therapies, including delivery of short, interfering RNAs (siRNAs) for RNA interference (RNAi), are limited due to the difficulty of delivering nucleic acids to specific cells of interest while at the same time minimizing toxicity and immunogenicity. The use of cationic polymers to deliver nucleic acid therapeutics has the potential to address these complex issues but is currently limited by low delivery efficiencies. While cell culture studies have shown that some polymers can be used to deliver siRNAs and achieve silencing, it is still not clear what physical or chemical properties are needed to ensure that the polymers form active polymer-siRNA complexes.

Although no general rules exist for designing siRNA delivery vehicles, there are key compositional elements common among effective vehicles such as amine groups or the use of disulfide bonds. In this study, we used multicolor fluorescence confocal microscopy to analyze the cellular uptake of siRNAs delivered by novel propargyl glycolide polymeric nanoparticles (NPs). Delivery was compared to that of linear polyethyleneimine (LPEI) and Lipofectamine 2000 (LF2K), both known as effective delivery vehicles for siRNAs.

Our results showed that LF2K and LPEI both delivered large initial quantities of siRNA rapidly, presumably overwhelming the basal levels of mRNA to initiate silencing. In contrast, our novel polymeric NPs also showed delivery of siRNAs but at initial concentrations too low to achieve silencing. Even so, the low cytotoxicity of our NPs and their simplicity for modification, has made them good candidates for further optimization. We will discuss our recent work on understanding the physical structures formed by our NP-siRNA complexes as well as methods employed to increase siRNA loading into the complexes, such as covalent coupling of multiple siRNA to a single polymer.