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Author Li, Jing
Title Multifunctional bioreducible nanoparticles for gene therapy [electronic resource] / by Jing Li.
Publication Info. 2012.
Location Call No. Status Notes
 Libraries Electronic Books  Electronic Resource - WSU ETD    AVAIL. ONLINE
Note Thesis supervisor: David Oupicky
Thesis Thesis (Ph.D.)--Wayne State University, 2012.
Summary Gene therapy is a promising therapeutic strategy to treat diseases caused by single or multiple gene mutations. Non-viral gene delivery vectors exhibit many advantages over viral vectors, including enhanced safety, versatility in choosing different types of therapeutic nucleic acids, and ease of formulation. However, low transgene efficiency and lack of targeting ability remain major challenges. Bioreducible polyplexes (polyelectrolyte complexes of disulfide-containing polycations and nucleic acids) utilize the redox potential gradient existing between extracellular space and intracellular environment as a physiological stimulus to enhance delivery of therapeutic nucleic acids to the subcellular space. Bioreducible containing polyplexes undergo intracellular reduction mediated GSH, which results in fast disassembly of the polyplexes and degradation of the delivery vector, leading to increased transfection efficiency and decreased cytotoxicity. In this dissertation, we first evaluated the effect of enhanced reductive disassembly on transfection activity of plasmid DNA and antisense oligonucleotide (AON) polyplexes using on a series of synthesized bioreducible poly(amido amine)s (PAA). We found that increasing the disulfide content in PAA increased susceptibility to reduction-triggered DNA and AON release from the polyplexes. However, increasing disulfide content in PAA resulted in increased DNA transfection only and had no effect on AON transfection. We discovered that plasma membrane protein thiols played a key role in the observed enhancement of DNA transfection. We then explored the possibility of introducing targeting moiety (hyaluronic acid, HA) into the design and formulation of the bioreducible polyplexes to stability polyplexes and to achieve selective delivery to CD44 expressing cancers.
Three different approaches to incorporate low- and medium-molecular-weight HA into the structure of bioreducible DNA polyplexes have been explored with the goal of improving steric stability and achieving CD44-selective uptake and transfection. Our results show that higher molecular weight HA is required for steric stabilization of the polyplexes while lower molecular weight HA was beneficial for targeted transfection activity. Further studies and development are needed to combine steric stabilization with efficient, CD44-selective transfection into a single formulation. Our next goal was to develop multifunctional bioreducible polyplexes for combination gene therapy in a single formulation. We have developed a series of bioreducible polycationic copper chelators (RPC) based on 1,4,8,11-tetraazacyclotetradecane (cyclam). We confirmed that the cyclam moieties in the polycations retained their ability to form complexes with Cu(II). The presence of disulfide bonds in the polycations resulted in substantially lower cytotoxicity than control 25 KDa PEI and both the Cu(II)-free and Cu(II) complexed polymers exhibited high transfection efficiency in vitro. This novel type of polycationic Cu(II) chelates represent promising nucleic acid delivery vectors with potential for PET imaging using 64Cu radioisotope. We further designed and developed a cyclam- based dual-function bioreducible polycations that function simultaneously as gene delivery vectors and as CXCR4 antagonists. We found that these dual-function polycations with their own pharmacological activity are able to prevent cancer cell invasion by inhibiting CXCL12 stimulated CXCR4 activation, while at the same time efficiently and safely delivers plasmid DNA into cancer cells.
System Details Mode of access: World Wide Web.
System requirements: Adobe Reader.
Added Author David Oupicky, advisor.
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