(190bf) Endocytosis and Trafficking of siRNA-Containing Complexes

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 endocytosis and 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 aim is to identify which endocytosis and intracellular trafficking pathways lead to active silencing by siRNA-containing complexes. Our work explores the preferential mechanism of endocytosis (whether by clathrin, caveolin, ARF6, GRAF1, flotillin, or macropinocytosis) across multiple cell types (HeLa (cervical), H1299 (lung), HEK293 (kidney), and HepG2 (liver)). Additionally, we will have applied confocal microscopy to independently track the vehicle and siRNA strands across endosomal vesicles (early, late, slow/fast recycling, and lysosomal), endoplasmic reticulum, and Golgi apparatus. To date, we have demonstrated that the preferred endocytic mechanism varies by cell type and that addition of chemical inhibitors can enhance siRNA accumulation through disruption of the function of counterproductive pathways.