(622c) Microfluidic Assembly of Multifunctional Polyplexes

The critical bottleneck to translation of human gene therapy remains the lack of safe and efficient methods for delivery of genetic material. Polymeric delivery agents have shown much promise. However, conventional polymeric vectors, which most often comprise plasmid DNA and a single polycation, cannot provide the multiple functionalities (e.g., binding of cellular receptors, intracellular trafficking steps, nuclear translocation, etc.) needed for efficient transfection. To achieve clinically relevant gene delivery activity with synthetic vectors will require more complex assemblies of DNA with multiple materials contributing the needed functions. Furthermore, construction of such multifunctional vectors will require new methods that provide controlled mixing and spatially defined arrangement of the DNA and polymers. We are developing microfluidic systems for construction of multilayered, multifunctional non-viral vectors.  The devices bring solutions of plasmid DNA and polymer(s) into contact under laminar flow providing enhanced control of polymer/DNA stoichiometry and interaction times and, most importantly, allowing introduction of multiple materials in a sequential fashion.  In this presentation, we will present design of a microfluidic device for layer-by-layer assembly of ternary polyanion/polycation/DNA polyplexes. We will compare physicochemical properties of the resulting polyplexes, including polyplex size, size uniformity and architecture, in comparison to polyplexes assembled via conventional bulk mixing. This novel polyplex assembly method is expected to provide the control and reproducibility necessary to enhance transfection efficiencies, facilitate investigation of gene transfer mechanisms, and ultimately enable clinical translation.