Non-viral gene delivery from polymer scaffolds for promoting tissue formation

The combination of gene therapy, biomaterials, and tissue engineering has the potential to direct progenitor cell development into functional tissues. Many tissue engineering approaches employ a polymer scaffold to provide a physical support for cell adhesion and migration, and organize tissue formation. Gene delivery from the scaffold can provide localized expression of tissue inductive factors to promote the development of functional tissue. Importantly, the optimal level and duration of transgene expression for functional tissue formation will certainly depend upon the application. Therefore, identifying the design parameters for DNA releasing polymer scaffolds that determine transgene expression is essential for their application to tissue engineering. I have investigated two mechanisms of polymer-mediated plasmid delivery for their ability to promote gene transfer and tissue formation in order to identify the design parameters. The two mechanisms are (i) sustained release and (ii) substrate-mediated delivery. Sustained release has been proposed to protect DNA from clearance or degradation, and maintain elevated concentrations locally, thereby extending the opportunities for cellular internalization of the DNA. Sustained release of plasmid DNA achieved transgene expression for over 3 months subcutaneously and for at least 6 months intramuscularly. In this approach, the polymer system properties, such as surface area, structural integrity, and degradation rate, as well as the dose of DNA were found to be critical design parameters for regulating and prolonging gene expression in vivo. Alternatively, substrate-mediated delivery involves the immobilization of DNA complexes directly to a biomaterial surface. This system led to high efficiency of transfection with substantially reduced DNA doses (i.e., nanogram quantities). Transgene expression was regulated through balancing several design parameters, including quantity of DNA deposited on the surface, charge ratio of DNA complexes, and surface-modification (i.e., protein-coating, surface hydrolysis). Both sustained and substrate-mediated delivery approaches using polymer scaffolds have the capacity to enhance gene transfer, and these methods have been used to promote tissue formation, as demonstrated by both blood vessel ingrowth and neurite outgrowth. This thesis has identified the design parameters for efficient gene transfer from tissue engineering scaffolds, which will facilitate their application towards coordinating the development of numerous tissues.