Date of this Version
Helmink, A. J. (2017). CHITOSAN NANOPARTICLE MODIFICATIONS FOR IMPROVED GENE DELIVERY IN AN ORAL DNA VACCINE APPLICATION (Unpublished Undergraduate Honors Thesis). University of Nebraska - Lincoln.
Vaccines represent one of the most significant medical innovations of the 20th century, resulting in the eradication or near eradication of a handful of deadly diseases. However, many infectious diseases remain resistant to effective vaccination, largely due to a lack full immune activation by traditional protein-based vaccines. A promising alternative vaccination strategy is the emerging development of DNA vaccines, which rely upon the delivery of exogenous genetic material to host cells encoding for a viral or bacterial antigen in order to induce a robust immune response by closely mimicking live infection. The delivery of genetic material requires a carrier in order to avoid degradation by host defenses in vivo. Non-viral gene delivery vectors avoid the immunogenicity and toxicity concerns of viral vectors, but suffer from low efficacy. Thus, techniques to improve the efficiency of gene delivery must be developed. Additionally, while many administration routes are viable, the oral route is preferable due to its ease of administration, high patient compliance, and ability to mimic live infection, which commonly occurs via the oral route. With these considerations in mind, we have previously formulated chitosan-zein nano-in-microparticles (CS-Zn-NIMs) as a dual-material oral gene delivery system consisting of chitosan /pDNA nanoparticle (CS/DNA NP) cores encapsulated within Zn microparticles. While we demonstrated moderate levels of transgene expression in vitro and successful immune responses in vivo, transfection efficiency remains relatively low. Thus, in this study we examine the potential of modifying the CS backbone with hydrophilic and hydrophobic molecules, which have been demonstrated to improve transfection efficiency via increased mucopenetration and cellular uptake, as well as with a molecule to specifically target antigen presenting cells (APCs), immune cells that a play a vital role in initiating robust immune responses. We describe the formulation and in vitro characterization of CS/DNA NPs using CS polymers modified with mannose (Man), poly(ethylene glycol) (PEG), and PEG-Man as well as the attempted formulation of CS-phenylalanine (CS-Phe)/DNA NPs. Modified CS/DNA NPs prepared by ionic gelation with sodium tripolyphosphate (TPP) demonstrated similar sizes to unmodified CS/DNA NPs and high DNA encapsulation efficiency. Furthermore, Man, PEG, and PEG-Man modified CS/DNA NPs exhibited improved transfection levels in vitro as compared to unmodified NPs. This work demonstrates the potential of CS-Man, CS-PEG, and CS-PEG-Man/DNA NPs for use in an oral gene delivery system.
Biological Engineering Commons, Biomaterials Commons, Biomedical and Dental Materials Commons, Molecular, Cellular, and Tissue Engineering Commons, Nanomedicine Commons, Pharmaceutical Preparations Commons