(232h) pH-Responsive, Polycationic Nanoparticles Designed for Intracellular siRNA Delivery

INTRODUCTION: Harnessing the specificity and potency of siRNA to treat disease by correcting aberrant gene expression is limited by delivery challenges.  The proposed pH-responsive, polycationic nanoparticles are designed to improve delivery by promoting endosomal escape and release of the siRNA into the cytosol (the site of action) following cellular uptake. 

MATERIALS AND METHODS: The polycationic nanoparticles are composed of 2-(diethylamino) ethyl methacrylate (DEAEMA) for pH-responsiveness, poly(ethylene glycol) methyl ether methacrylate (PEGMA) for colloidal stability and biocompatibility, a crosslinking agent for enhanced nanoparticle stability even at low concentrations, and hydrophobic methacrylate monomers to tune the swelling and membrane disruptive characteristics.  The emulsion-based ARGET ATRP scheme for nanoparticle synthesis was adapted from a free radical photoemulsion polymerization developed by Fisher et al.,¹.  This new methacrylate polymerization protocol occurs in water at ambient temperature without UV light.

RESULTS AND DISCUSSION: A panel of polycationic nanoparticles was synthesized and characterized.  Dynamic light scattering indicated pH-responsive swelling near physiological pH which could be tuned as a function of composition.  Hemolysis assays at pH values characteristic of the extracellular matrix (7.4) and early endosomes (6.5) were used as an indicator of endosomolytic activity.  Incorporating a hydrophobic monomer resulted in the desired membrane disruption outcome:  low membrane disruption at pH 7.4 (2-4% of the positive control) and strong membrane disruption at pH 6.5 (10-100% of the positive control). 

Initial in vitro characterization indicates that the nanoparticle cytotoxicity is equivalent to that of particles synthesized using the traditional free radical polymerization scheme developed by Fisher et al.,¹, and Fisher and Peppas².  Additional characterization using a model intestinal epithelial cell culture model will enhance understanding of the polymer carrier cytotoxicity as well as the silencing efficiency of siRNA-loaded carriers. 

ACKNOWLEDGMENTS: This work was supported by the U.S. National Science Foundation (CBET 10-33746) and a National Science Foundation Graduate Research Fellowship to DCF (DGE-1110007).

REFERENCES:

1. Fisher, O.; Kim, T.; Dietz, S.; Peppas, N., Enhanced Core Hydrophobicity, Functionalization and Cell Penetration of Polybasic Nanomatrices. Pharmaceutical Research 2009, 26 (1), 51-60.

2. Fisher, O. Z.; Peppas, N. A., Polybasic Nanomatrices Prepared by UV-Initiated Photopolymerization. Macromolecules 2009, 42 (9), 3391-3398.