Clostridium neurotoxin fragments as potential targeting moieties for liposomal gene delivery to the CNS.
Targeted transfection of the CNS with synthetic, nonviral vectors represents a huge technical challenge. The approach explored here attempts to combine self-assembly ABCD nanoparticles (Kostarelos and Miller, Chem. Soc. Rev. 2005, 34, 970), with the potential of Clostridium neurotoxin fragments to effect receptor-mediated transfection of neuronal cells. Cationic liposome-plasmid DNA complexes were first modified with a PEG stealth layer, before the addition of C-terminal fragments of tetanus toxin (TH(C)), botulinum toxin (BH(C)) or the truncated C-terminal domain of TH(C) as biological "targeting" ligands. First-generation nanoparticles were identified for the transfection of two neuronal cell lines (human SH-5YSY and rat/mouse hybrid N18-RE105); control studies were also performed with HeLa cells. ABCD nanoparticle transfections of the neuronal cell lines were up to 30-fold higher than corresponding control transfections with nanoparticles that lacked the protein ligand. We also demonstrate apparent receptor-mediated uptake by means of competition-binding and real-time confocal experiments. By contrast, nanoparticle transfection of HeLa cells appeared to involve alternative nonspecific enhanced cellular uptake mechanism(s). Receptor-mediated and nonspecific mechanisms appear to be in competition, potentially harming the capacity of receptor-mediated delivery to effect proper targeted delivery of nucleic acids to cells ex vivo and in vivo.
Andreu
Alice
A
Imperial College Genetic Therapies Centre, Department of Chemistry, Flowers Building, Armstrong Road, Imperial College London, London SW7 2AZ, UK.
Fairweather
Neil
N
Miller
Andrew D
AD
D000818:Animals; D001667:Binding, Competitive; D001905:Botulinum Toxins / Q000737:chemistry* / Q000235:genetics; D002460:Cell Line; D002490:Central Nervous System / Q000166:cytology / Q000502:physiology*; D004247:DNA / Q000737:chemistry / Q000235:genetics; D018390:Gene Targeting / Q000379:methods*; D018014:Gene Transfer Techniques; D017930:Genes, Reporter / Q000235:genetics; D006367:HeLa Cells; D006801:Humans; D008081:Liposomes; D051379:Mice; D009474:Neurons / Q000166:cytology / Q000502:physiology; D009498:Neurotoxins / Q000737:chemistry* / Q000235:genetics; D010446:Peptide Fragments / Q000737:chemistry* / Q000235:genetics; D010957:Plasmids / Q000737:chemistry / Q000235:genetics; D051381:Rats; D014162:Transfection / Q000379:methods
D008081:Liposomes; D009498:Neurotoxins; D010446:Peptide Fragments; D004247:DNA; D001905:Botulinum Toxins