Neuron-targeted copolymers with sheddable shielding blocks synthesized using a reducible, RAFT-ATRP double-head agent

Adaptation of in vitro optimized polymeric gene delivery systems for in vivo use remains a significant challenge. Most in vivo applications require particles that are sterically stabilized, which significantly compromises transfection efficiency of materials shown to be effective in vitro. We present a multifunctional well-defined block copolymer that forms particles useful for cell targeting, reversible shielding, endosomal release, and DNA condensation. We show that targeted and stabilized particles retain transfection efficiencies comparable to the nonstabilized formulations. A novel, double-head agent that combines a reversible addition-fragmentation chain transfer agent and an atom transfer radical polymerization initiator through a disulfide linkage is used to synthesize a well-defined cationic block copolymer containing a hydrophilic oligoethyleneglycol and a tetraethylenepentamine-grafted polycation. This material effectively condenses plasmid DNA into salt-stable particles that deshield under intracellular reducing conditions. In vitro transfection studies show that the reversibly shielded polyplexes afford up to 10-fold higher transfection efficiencies than the analogous stably shielded polymer in four different mammalian cell lines. To compensate for reduced cell uptake caused by the hydrophilic particle shell, a neuron-targeting peptide is further conjugated to the terminus of the block copolymer. Transfection of neuron-like, differentiated PC-12 cells demonstrates that combining both targeting and deshielding in stabilized particles yields formulations that are suitable for in vivo delivery without compromising in vitro transfection efficiency and are thus promising carriers for in vivo gene delivery applications.