These data support the potential of developing sd-rxRNAs as a therapeutic for ocular disease.

Chemical modification of RNA duplexes can provide practical advantages for RNA interference (RNAi) triggering molecules including increased stability, safety and specificity. The impact of nucleotide modifications on Dicer processing, RISC loading and RNAi-mediated mRNA cleavage was investigated with duplexes ≥25 bp in length. It is known that dsRNAs ≥25 bp are processed by Dicer to create classic 19-bp siRNAs with 3′-end overhangs. We demonstrate that the presence of minimal modification configurations on longer RNA duplexes can block Dicer processing and result in the loading of the full-length guide strand into RISC with resultant mRNA cleavage at a defined site. These longer, modified duplexes can be highly potent gene silencers, with EC50s in the picomolar concentration range, demonstrating that Dicer processing is not required for incorporation into RISC or potent target silencing.

RNA interference (RNAi) has been established as an important tool for functional genomics studies and has great promise as a therapeutic intervention for human diseases. In mammalian cells, RNAi is conventionally induced by 19-27-bp RNA duplexes generated by hybridization of two complementary oligonucleotide strands (oligos). Here we describe a novel class of RNAi molecules composed of a single 25-28-nucleotide (nt) oligo. The oligo has a 16-nt mRNA targeting region, followed by an additional 8-10 nt to enable self-dimerization into a partially complementary duplex. Analysis of numerous diverse structures demonstrates that molecules composed of two short helices separated by a loop can efficiently enter and activate the RNAinduced silencing complex (RISC). This finding enables the design of highly effective single-oligo compounds for any mRNA target.

Data will be presented on a method for RNAi compound delivery utilizing RNA encapsulated in beta-glucan shells. When orally administered to mice, these encapsulated nucleic acids traverse the intestinal epithelia by an M cell transcytosis mechanism and are then endocytosed by macrophages. Macrophages containing RNAi compounds migrate to sites of inflammation, and the targeted RNA is specifically inhibited by RNA interference. This formulation presents some unique advantages for therapeutic RNAi including oral mode of administration, low dose (ug/kg/day) and specific delivery to immune cells possibly affording safety advantages for some targets. This novel mode of RNA administration can be used to down-regulate target genes involved in multiple inflammatory disease indications.