Date of Award
1-1-2024
Document Type
Dissertation
Degree Name
Ph.D. in Engineering Science
First Advisor
Adam E. Smith
Second Advisor
Thomas A. Werfel
Third Advisor
Nikki Reinemann
School
University of Mississippi
Relational Format
dissertation/thesis
Abstract
This work investigates the development and application of charge-shifting polycations for gene delivery, focusing on their synthesis, characterization, and applications for in vitro mRNA delivery. Utilizing poly(2-vinyl-4,4-dimethylazlactone) (PVDMA), this study explores modifications with various reactive handles, such as alcohol, thiol, and amine groups, to create cationic amide-, ester-, and thioester-functionalized polycations. These modifications aim to exploit the charge-shifting capabilities of the polycations to form stable polyplexes with nucleic acids, promoting the efficient intracellular release of the genetic cargo.Initial studies first examine the charge-shifting properties of PVDMA-based polycations under different pH and temperature conditions. This work highlights the potential of these polymers to enhance nucleic acid delivery by dynamically shifting their net charge in response to physiological triggers, thereby reducing cytotoxicity and improving unpackaging efficiency. Additionally, this research investigates the potential of thioester-modified PVDMA as a glutathione (GSH)-responsive charge-shifting polycation, which can exhibit increased intracellular delivery of nucleic acid cargo. The findings demonstrate that thioester-PVDMA polymers can effectively release nucleic acids in the presence of intracellular GSH, offering a key advantage for targeted gene delivery applications. Furthermore, this study introduces a new thioester-based cationic monomer, dimethylaminoethyl thioester methacrylate, and details its polymerization to create thioester-based polycations. The transfection efficacy of these new polymers is compared to established polycationic vectors like PDMAEMA, providing insights into their potential for gene therapy applications. Comprehensive in vitro studies using HEK293T cells compare the transfection efficiency of these thioester-based polycations with traditional PDMAEMA counterparts. This work concludes by outlining future directions to optimize these novel gene delivery vectors. Recommendations include further enhancing the hydrolytic stability of thioester-PVDMA through chemical modifications, exploring advanced polymer architectures such as star and bottlebrush structures, and conducting in vivo studies to evaluate biocompatibility and therapeutic efficacy. This research seeks to advance the field of non-viral gene delivery systems by offering promising pathways for developing safer and more effective gene therapy treatments.
Recommended Citation
Fortenberry, Alexander, "Cationic Charge-Shifting Polymers as Improved Vectors for Nucleic Acid Delivery" (2024). Electronic Theses and Dissertations. 2934.
https://egrove.olemiss.edu/etd/2934