title: Tetanus toxin fragment C fusion facilitates protein delivery to CNS neurons from cerebrospinal fluid in mice.
abstract: To improve protein delivery to the CNS following intracerebroventricular administration, we compared the distribution of a human Cu/Zn superoxide dismutase:tetanus toxin fragment C fusion protein (SOD1:TTC) in mouse brain and spinal cord with that of tetanus toxin fragment C (TTC) or human SOD1 (hSOD1) alone, following continuous infusion into the lateral ventricle. Mice infused with TTC or SOD1:TTC showed intense anti-TTC or anti-hSOD1 labeling, respectively, throughout the CNS. In contrast, animals treated with hSOD1 revealed moderate staining in periventricular tissues. In spinal cord sections from animals infused with SOD1:TTC, the fusion protein was found in neuron nuclear antigen-positive (NeuN+) neurons and not glial fibrillary acidic protein-positive (GFAP+) astrocytes. The percentage of NeuN+ ventral horn cells that were co-labeled with hSOD1 antibody was greater in mice treated with SOD1:TTC (cervical cord = 73 +/- 8.5%; lumbar cord = 62 +/- 7.7%) than in mice treated with hSOD1 alone (cervical cord = 15 +/- 3.9%; lumbar cord = 27 +/-4.7%). Enzyme-linked immunosorbent assay for hSOD1 further demonstrated that SOD1:TTC-infused mice had higher levels of immunoreactive hSOD1 in CNS tissue extracts than hSOD1-infused mice. Following 24 h of drug washout, tissue extracts from SOD1:TTC-treated mice still contained substantial amounts of hSOD1, while extracts from hSOD1-treated mice lacked detectable hSOD1. Immunoprecipitation of SOD1:TTC from these extracts using anti-TTC antibody revealed that the recovered fusion protein was structurally intact and enzymatically active. These results indicate that TTC may serve as a useful prototype for development as a non-viral vehicle for improving delivery of therapeutic proteins to the CNS.
authors.affiliation: Cecil B. Day Laboratory for Neuromuscular Research, Department of Neurology, Massachusetts General Hospital, Charlestown, Massachusetts 02129, USA.
authors.forename: Susanna C
authors.initials: SC
authors.lastname: Benn
authors.forename: Ilknur
authors.initials: I
authors.lastname: Ay
authors.forename: Elena
authors.initials: E
authors.lastname: Bastia
authors.forename: Ru-Ju
authors.initials: RJ
authors.lastname: Chian
authors.forename: Samuel A
authors.initials: SA
authors.lastname: Celia
authors.forename: R Blake
authors.initials: RB
authors.lastname: Pepinsky
authors.forename: Paul S
authors.initials: PS
authors.lastname: Fishman
authors.forename: Robert H
authors.initials: RH
authors.lastname: Brown
authors.forename: Jonathan W
authors.initials: JW
authors.lastname: Francis
mesh_terms: D000818:Animals; D015153:Blotting, Western / Q000379:methods; D002452:Cell Count / Q000379:methods; D002490:Central Nervous System / Q000166:cytology* / Q000187:drug effects; D006801:Humans; D007150:Immunohistochemistry / Q000379:methods; D007276:Injections, Intraventricular / Q000379:methods; D008297:Male; D051379:Mice; D008810:Mice, Inbred C57BL; D009474:Neurons / Q000187:drug effects* / Q000378:metabolism*; D010446:Peptide Fragments / Q000378:metabolism / Q000494:pharmacology*; D010751:Phosphopyruvate Hydratase; D011994:Recombinant Proteins / Q000302:isolation & purification / Q000378:metabolism / Q000494:pharmacology; D013482:Superoxide Dismutase / Q000134:cerebrospinal fluid*; D000072105:Superoxide Dismutase-1; D013744:Tetanus Toxin / Q000378:metabolism / Q000494:pharmacology*; D014018:Tissue Distribution / Q000187:drug effects
chemical_list: D010446:Peptide Fragments; D011994:Recombinant Proteins; C000606290:SOD1 protein, human; D013744:Tetanus Toxin; C040635:tetanus toxin fragment C; C000606291:Sod1 protein, mouse; D013482:Superoxide Dismutase; D000072105:Superoxide Dismutase-1; D010751:Phosphopyruvate Hydratase