Gene therapy holds great promise for treating various diseases but relies on safe and efficient delivery vectors for genetic material.[1] Polymers bearing cationic sites can form polyplexes via electrostatic interaction with nucleic acids (NAs). This increases circulation times in the bloodstream by shielding the payload and enhances cellular uptake.[2] However, depending on their structure, polymers with pronounced positive charge may exhibit significant cytotoxicity.[2,3] Therefore, charge density and molecular architecture of polycations must be carefully selected to balance desired and adverse effects.
Our work focuses synthesizing statistical copolymers of cationic Vinylamine (VAm) and charge-neutral hydrophilic N-Vinylpyrrolidone (NVP). VAm was introduced via polymerization of N-Vinylformamide (NVF) and subsequent acidic hydrolysis. The polymers were prepared using photo-iniferter reversible addition-fragmentation chain-transfer (PI-RAFT) polymerization, yielding well defined macromolecules with low polydispersities (Đ = 1.2–1.4). To tailor charge density, we varied the ratio of NVF to NVP in monomer feed. The synthesized polymers show more than 10-fold greater biocompatibility compared to linear Polyethyleneimine and the liposomal carrier Lipofectamine 2000, along with as up to 14-fold increased gene delivery efficacies in luciferase plasmid assays.
To further dictate the shape of polyplexes and modulate their properties as well as NA availability, we prepared sequential bottle brush copolymers via RAFT polymerization, incorporating a block of Polyethylene glycol (PEG), as well as a block containing PVAm-stat-PNVP grafts using a combined grafting-through/grafting-from approach. This architecture aims to reduce cytotoxicity by using PEG grafts to shield the PVAm-NA-polyplex, while still enabling intracellular disassembly and NA delivery.
[1] A. Shahryari, M. Saghaeian Jazi, S. Mohammadi, H. Razavi Nikoo, Z. Nazari, E. S. Hosseini, I. Burtscher, S. J. Mowla, H. Lickert, Frontiers in genetics 2019, 10, 868.
[2] A. Pathak, S. Patnaik, K. C. Gupta, Biotechnology Journal 2009, 4, 1559–1572.
[3] a). B. Yahya, A. M. Alqadhi, Life Sciences 2021, 269; b). Fischer, Y. Li, B. Ahlemeyer, J. Krieglstein, T. Kissel, Biomaterials 2003, 24, 1121–1131.