(514f) Cationic Precursors to Mixed-Amine Zwitterionic Polymers for Safe and Effective DNA Vaccines

A key limitation to the development of human gene therapy is the lack of safe, efficient, and controllable methods for gene delivery. Current research focuses on non-viral gene delivery agents, but these generally lack the required efficacy, cause toxicity due to both the materials used and nonspecific delivery, and are unstable in vivo when administered systemically. The engineering of specialized vehicles capable of overcoming various gene delivery barriers is critical to achieving successful gene transfection. This work describes efforts to extend a previously-developed integrated platform based on hydrolysable zwitterionic carboxybetaine methacrylate (CBMA) polymers for use as a DNA vaccine. Zwitterionic materials are superlow-fouling in blood, hydrolytically degradable, readily tunable, and biomimetic (nontoxic). The key attribute of CBMA-ester polymers that makes them ideal for gene delivery lies in the fact that they are positively charged in their native form when they condense DNA, but they become zwitterionic CBMA upon hydrolysis in an acidic environment (as in the endosome). In the zwitterionic state, the polymer repels the DNA, and unpacks the nanoparticle to leave a biomimetic pCBMA coproduct. We have previously studied the effect of differently-substituted amines and found an optimal ratio tertiary:quaternary CBMA-ethyl esters for nonspecific gene delivery to COS-7 cells. Here, we adapt the technology for use as a DNA vaccine. DNA vaccines are conceptually similar to gene delivery, except that the genetic material is delivered to macrophage cells for expression. The macrophage cells, upon expression of the gene, display the transcribed protein via MHC Class I-peptide in order to activate T-cells and to mount an immune response that will lead to immunity against the pathogen whose DNA was delivered. Macrophages were successfully transfected with nanoparticles made from luciferase DNA condensed by our mixed-amine cationic precursors to zwitterionic polymers. A major difference is that macrophage cells may also be passively targeted by using microparticles instead of nanoparticles, because only macrophages and a few other types of cells can take up microparticles. Therefore, latex microparticles were coated with our mixed-amine hydrolytic polymers, and DNA was electrostatically buried in the polymer coating. We will report our success selectively transfecting macrophage cells with microparticles, and of activating T-cells co-cultured with the macrophages.