MAXIMIZING siRNA DELIVERY EFFICIENCY WITH PHOTO-RESPONSIVE POLYMER NANOCARRIERS

Abstract

In that last few decades, there has been a great interest in developing small interfering RNA (siRNA) therapeutics and research tools. There are many challenges that limit success of siRNA therapeutics, including control over siRNA release, lack of predictability, and the need for multiple doses of siRNA for sustained treatment. In this thesis, those challenges are addressed through the formulation of photo-responsive polyplexes for siRNA delivery. Mixed polyplexes were created by mixing two photo-responsive block copolymers to optimize polyplex properties, including size and zeta potential. It was found that a 50/50 mixture of the two polymers was able to achieve 70% protein knockdown of glyceraldehyde 3-phosphate dehydrogenase (GAPDH), which is suggested to be the maximum amount of GAPDH silencing possible in a single dose of siRNA. Furthermore, a simple kinetic model was employed to predict siRNA, messenger RNA (mRNA), and protein concentrations in the cell over time and develop a dosing schedule. These predictions enabled the application of a second dose of siRNA to suppress gene expression over three days, leading to a further 50% reduction in protein levels compared to a single dose. Polyplexes remained dormant in cells until exposed to light, demonstrating the complete control over siRNA activity as well as the stability of the nanocarriers. Additionally, a layered, photo-responsive system was developed to obtain controlled release over multiple doses of siRNA from a single particle. Thus, this work demonstrates that simple kinetic modeling and straightforward biomaterials design can be used to suppress gene expression maximally for an siRNA delivery system.