(679a) Dynamics of DNA Release From Polyplexes Induced by Inter-Polyelectrolyte Exchange

Polymeric gene delivery systems are attractive alternatives to virus-based systems due to safety concerns. A major problem of polymeric gene carriers is their short circulation half-life in vivo. We carried out a mechanistic study of DNA release induced by inter-polyelectrolyte exchange, which is believed to be one of the main causes of instability for polyplexes in extracellular environment. Bioreducible hyperbranched poly(amido amine) (RHB) has shown improved gene delivery performance in previous studies, and its depolymerization-induced DNA release process has been visualized at the molecular level. In this study RHB as well as poly(ethyleneimine) (PEI) were used as polycations to condense plasmid DNA. Heparin and poly(sodium 4-styrenesulfonate) (PSS) were used as competing polyanions to DNA to induce the inter-polyelectrolyte exchange reaction. The DNA release process was studied at the single polyplex level using in situ real-time AFM imaging. The AFM results were correlated with fluorescence data as well as cell transfection data. A critical polyanion concentration was determined and found to be a function of polycation charge density, chain architecture, and amino to phosphate (N/P) ratio. AFM was used to capture the morphological release pathway with distinct polyplex structures. These characteristic morphologies include core-shell, toroid, nanoparticle decorated toroid, and loose chains held by a compact core structure. This study contributes to basic understanding of DNA release dynamics from single polyplex particles in simulated physiologic conditions and provides synthetic strategies to improve gene delivery efficiency of polymeric vectors.