| dc.description.abstract | Retinoic acid-inducible gene I (RIG-I) is a cytosolic pattern recognition receptor (PRR) that potently activates antiviral innate immunity in response to binding 5’ triphosphate double-stranded RNA (3pRNA). Accordingly, 3pRNA has emerged as a promising class of therapeutic molecules for vaccine adjuvants, cancer immunotherapies, and antiviral agents; however, like other nucleic acid-based therapeutics, 3pRNA has poor ‘druglike’ properties due to its size, charge, and susceptibility to nuclease degradation. The design of polymeric carriers for the delivery 3pRNA is largely informed by those used for small interfering RNA (siRNA), owing to the structural and chemical similarities between the two classes of RNA, but despite these similarities, formulations of 3pRNA and polymeric carriers can exhibit unacceptable toxicities that are otherwise not observed in similar formulations of siRNA in vivo, highlighting the need for new carriers that are optimized for the delivery of 3pRNA. In order to address this problem, covalent conjugation is evaluated as a strategy for the loading of 3pRNA into nanocarrier, and it is compared to the conventional electrostatic complexation approach. Firstly, the parameters surrounding how the conjugation of 3pRNA related to polymer size, site specificity, and linker chemistry are investigated to highlight their individual contributions to the immune-stimulating activity of 3pRNA conjugates. Next, this information is used to design, develop, and evaluate a panel of polymers to investigate how conjugation of 3pRNA to nanocarrier affects the performance of the overall formulation. These experiments show that covalent conjugation to polymer nanocarrier improves the loading efficiency, nuclease resistance, and serum stability of the 3pRNA according to which part of the carrier it is conjugated to. Finally, covalent conjugation of 3pRNA to polymer nanocarrier improves RIG-I activation efficiency in vivo, allowing for stronger responses with less polymer. Overall, the work in this dissertation highlights a new strategy to improve the efficiency, stability, and in vivo efficacy of 3pRNA formulations. | |
