Solely aqueous formulation of hydrophobic cationic polymers for efficient gene delivery.

Abstract Cationic polymers are promising gene delivery vectors due to their ability to bind and protect genetic material. The introduction of hydrophobic moieties into cationic polymers can further improve the vector efficiency, but common formulations of hydrophobic polymers involve harsh conditions such as organic solvents, impairing intactness and loading efficiency of the genetic material. In this study, a mild, aqueous formulation method for the encapsulation of high amounts of genetic material is presented. A well-defined pH-responsive hydrophobic copolymer, i.e. poly((n-butylmethacrylate)-co-(methylmethacrylate)-co-(2-(dimethylamino) ethylmethacrylate)), PBMD) was synthesized by reversible addition fragmentation chain transfer (RAFT) polymerization. Exploiting the pH-dependent solubility behavior of the polymer, stable pDNA loaded nanoparticles were prepared and characterized using analytical ultracentrifugation (AUC), cryo-transmission electron microscopy (cryo-TEM) and dynamic light scattering (DLS). This novel formulation approach showed high transfection efficiencies in HEK293T cells, while requiring 5- to 10-fold less pDNA compared to linear polyethylenimine (LPEI), in particular at short incubation times and in serum-containing media. Furthermore, the formulation was successfully adopted for siRNA and mRNA encapsulation and the commercially approved polymer Eudragit® E(PO/100). Overall, the aqueous formulation approach, accompanied by a tailor-made hydrophobic polymer and detailed physicochemical and application studies, led to improved gene delivery vectors with high potential for further applications.