Gene therapy offers promise for treatment of many diseases. The success of gene therapy relies on efficient and safe delivery systems. We propose to develop an efficient non-viral DNA delivery system using surfaces loaded with DNA/calcium phosphate (DNA/CaP) composites. Our hypothesis is based on that DNA/CaP rich surfaces can provide high concentration of DNA in the immediate environment of cultured cells and high delivery efficiency can be achieved through optimization of DNA/CaP composite characteristics. We will use PLGA film to support DNA/CaP composite formation for its versatility and compatibility with biological system. Using procedures learned from natural biomineralization process, we will modify PLGA surfaces and form DNA/CaP composites in solutions under predictable and controllable thermodynamic driving forces. The DNA concentration and calcium phosphate solution conditions will be changed systematically to obtain a variety of DNA/CaP composites with controlled properties. The composite characteristics will be carefully examined using state-of-the-art analytical techniques available at Cornell University. Plasmid DNA encoding 2-galactosidase (pVAX/LacZ) and relevant cell lines (Chinese hamster ovarian (CHO) and MG-63 osteoblast-like cells) will be used to study the biocompatibility and transfection efficiency of the composites in vitro. Data analysis will focus on how material characteristics affect transfection efficiency. We believe that DNA/CaP loaded PLGA surfaces with definable and controllable physicochemical properties will make an effective gene delivery system that is not currently available using any other means of production. Because the major component of bone is calcium phosphate, the DNA/CaP loaded PLGA may have direct application in bone tissue engineering. The proposed study will help shed light on the mechanisms of calcium phosphate mediated gene delivery to cells. Moreover, we believe that this study will provide foundation for further development of three dimensional tissue engineering scaffolds with gene delivery capability---a system that will be explored for in vivo studies using animal models and for eventual clinical applications. A future R01 proposal concept will be developed on the basis of this application. This project is to develop a highly efficient calcium phosphate gene delivery system based on strategies learned from the natural biomineralization process. The technology developed in this proposal will have important impact on basic research, biotech industry and gene therapy based medicine. ? ? ?
