Porous silicon nanoneedles for intracellular delivery of small interfering RNA

textThe rational and directed delivery of genetic material to the cell is a formidable tool to investigate the phenotypic effects of gene expression regulation and a promising therapeutic strategy for genetic defects. RNA interference constitutes a versatile approach to gene silencing. Despite the development of numerous strategies the transfection of small interfering RNA (siRNA) is highly dependent on cell type and conditions. Direct physical access to the intracellular compartment is a promising path for high efficiency delivery independently of cell type and conditions. Silicon nanowires grant such access with minimal toxic effects, and allow intracellular delivery of DNA when actuated by atomic force microscope. These findings reveal the potential for porous silicon nanostructures to serve as delivery vectors for nucleic acids due to their porous nature, elevated biocompatibility, and biodegradability. This dissertation illustrates the development a novel platform for efficient siRNA transfection based on an array of porous silicon nanoneedles. The synthesis of biodegradable and biocompatible porous nanowires was accomplished by a novel strategy for electroless etch of silicon that allows anisotropic etch simultaneously with porosification. An ordered array of cone shaped porous silicon nanoneedles with tunable tip size, array density and aspect ratio was obtained coupling this strategy with patterned metal deposition and selective reactive ion etch. This process also granted control over porosity, nanopore size and flexural modulus. The combination of these parameters was appropriately optimized to ensure cell penetration, maximize siRNA loading and minimize cytotoxic effects. Loading of the negatively charged siRNA molecules was optimized by applying an external electric field to the nanoneedles under appropriate voltage conditions to obtain a tenfold increase over open circuit loading, and efficient penetration of the siRNA within the porous volume of the needles. Alternative surface chemistry modification provided a means for effective siRNA loading and sustained release. siRNA transfection was achieved by either imprinting the nanoneedles array chip over a culture of MDA-MB-231 cells or allowing the cells to self-impale over the needles. The procedures allowed the needles to penetrate across the cell membrane without influencing cell proliferation. siRNA was successfully transfected and was effective at suppressing gene expression.Biomedical Engineerin