Neuron-specific delivery of nucleic acids mediated by Tet1-modified poly(ethylenimine)

Background: The development of minimally invasive, non-viral gene delivery vehicles for the central nervous system (CNS) is an important technology goal in the advancement of molecular therapies for neurological diseases. One approach is to deliver materials peripherally that are recognized and retrogradely transported by motor neurons toward the CNS. Tet1 is a peptide identified by Boulis and coworkers to possess the binding characteristics of tetanus toxin, which interacts specifically with motor neurons and undergoes fast, retrograde delivery to cell soma. In this work, Tet1-poly(ethylenimine) (Tet1-PEI) was synthesized and evaluated as a neurontargeted delivery vehicle. Methods: Tet1-PEI and NT-PEI (neurotensin-PEI) were synthesized and complexed with plasmid DNA to form polyplexes. Polyplexes were assessed for binding and uptake in differentiated neuron-like PC-12 cells by flow cytometry and confocal microscopy. In order to determine gene delivery efficiency, polyplexes were exposed to PC-12 cells at various stages of differentiation. Targeted binding of polyplexes with primary neurons was studied using dorsal root ganglion cells. Results: Tet1-PEI and NT-PEI polyplexes bound specifically to differentiated PC-12 cells. The specificity of the interaction was confirmed by delivery to non-neuronal cells and by competition studies with free ligands. Tet1-PEI polyplexes preferentially transfected PC-12 cells undergoing NGF-induced differentiation. Finally, neuron-specific binding of Tet1-PEI polyplexes was confirmed in primary neurons. Conclusions: These studies demonstrate the potential of Tet1-PEI as a neuron-targeted material for non-invasive CNS delivery. Tet1-PEI binds specifically and is internalized by neuron-like PC-12 cells and primary dorsal root ganglion. Future work will include evaluation of siRNA delivery with these vector.