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- All Subjects: Gene Therapy
- Genre: Doctoral Dissertation
- Creators: Rege, Kaushal
- Status: Published
Description
Gold nanoparticles have emerged as promising nanomaterials for biosensing, imaging, photothermal treatment and therapeutic delivery for several diseases, including cancer. We have generated poly(amino ether)-functionalized gold nanorods (PAE-GNRs) using a layer-by-layer deposition approach. Sub-toxic concentrations of PAE-GNRs were employed to deliver plasmid DNA to prostate cancer cells in vitro. PAE-GNRs generated using 1,4C-1,4Bis, a cationic polymer from our laboratory demonstrated significantly higher transgene expression and exhibited lower cytotoxicities when compared to similar assemblies generated using 25 kDa poly(ethylene imine) (PEI25k-GNRs), a current standard for polymer-mediated gene delivery. Additionally, sub-toxic concentrations of 1,4C-1,4Bis-GNR nanoassemblies were employed to deliver expression vectors that express shRNA ('shRNA plasmid') against firefly luciferase gene in order to knock down expression of the protein constitutively expressed in prostate cancer cells. The roles of poly(amino ether) chemistry and zeta-potential in determining transgene expression efficacies of PAE-GNR assemblies were investigated. The theranostic potential of 1,4C-1,4Bis-GNR nanoassemblies was demonstrated using live cell two-photon induced luminescence bioimaging. The PAE class of polymers was also investigated for the one pot synthesis of both gold and silver nanoparticles using a small library poly(amino ethers) derived from linear-like polyamines. Efficient nanoparticle synthesis dependent on concentration of polymers as well as polymer chemical composition is demonstrated. Additionally, the application of poly(amino ether)-gold nanoparticles for transgene delivery is demonstrated in 22Rv1 and MB49 cancer cell lines. Base polymer, 1,4C-1,4Bis and 1,4C-1,4Bis templated and modified gold nanoparticles were compared for transgene delivery efficacies. Differences in morphology and physiochemical properties were investigated as they relate to differences in transgene delivery efficacy. There were found to be minimal differences suggestion that 1,4C-1,4Bis efficacy is not lost following use for nanoparticle modification. These results indicate that poly(amino ether)-gold nanoassemblies are a promising theranostic platform for delivery of therapeutic payloads capable of simultaneous gene silencing and bioimaging.
ContributorsRamos, James (Author) / Rege, Kaushal (Thesis advisor) / Kodibagkar, Vikram (Committee member) / Caplan, Michael (Committee member) / Vernon, Brent (Committee member) / Garcia, Antonio (Committee member) / Arizona State University (Publisher)
Created2014
Description
Gene therapy is a promising technology for the treatment of various nonheritable and genetically acquired diseases. It involves delivery of a therapeutic gene into target cells to induce cellular responses against diseases. Successful gene therapy requires an efficient gene delivery vector to deliver genetic materials into target cells. There are two major classes of gene delivery vectors: viral and non-viral vectors. Recently, non-viral vectors such as cationic polymers have attracted more attention than viral vectors because they are versatile and non-immunogenic. However, cationic polymers suffer from poor gene delivery efficiency due to biological barriers. The objective of this research is to develop strategies to overcome the barriers and enhance polymer-mediated transgene expression. This study aimed to (i) develop new polymer vectors for gene delivery, (ii) investigate the intracellular barriers in polymer-mediated gene delivery, and (iii) explore new approaches to overcome the barriers. A cationic polymer library was developed by employing a parallel synthesis and high-throughput screening method. Lead polymers from the library were identified from the library based on relative levels of transgene expression and toxicity in PC3-PSMA prostate cancer cells. However, transgene expression levels were found to depend on intracellular localization of polymer-gene complexes (polyplexes). Transgene expression was higher when polyplexes were dispersed rather than localized in the cytoplasm. Combination treatments using small molecule chemotherapeutic drugs, e.g. histone deacetylase inhibitors (HDACi) or Aurora kinase inhibitor (AKI) increased dispersion of polyplexes in the cytoplasm and significantly enhanced transgene expression. The combination treatment using polymer-mediated delivery of p53 tumor-suppressor gene and AKI increased p53 expression in PC3-PSMA cells, inhibited the cell proliferation by ~80% and induced apoptosis. Polymer-mediated p53 gene delivery in combination with AKI offers a promising treatment strategy for in vivo and clinical studies of cancer gene therapy.
ContributorsBarua, Sutapa (Author) / Rege, Kaushal (Thesis advisor) / Dai, Lenore (Committee member) / Meldrum, Deirdre R. (Committee member) / Sierks, Michael (Committee member) / Voelkel-Johnson, Christina (Committee member) / Arizona State University (Publisher)
Created2011