title: Engineering Botulinum Neurotoxin C1 as a Molecular Vehicle for Intra-Neuronal Drug Delivery.
abstract: Botulinum neurotoxin (BoNT) binds to and internalizes its light chain into presynaptic compartments with exquisite specificity. While the native toxin is extremely lethal, bioengineering of BoNT has the potential to eliminate toxicity without disrupting neuron-specific targeting, thereby creating a molecular vehicle capable of delivering therapeutic cargo into the neuronal cytosol. Building upon previous work, we have developed an atoxic derivative (ad) of BoNT/C1 through rationally designed amino acid substitutions in the metalloprotease domain of wild type (wt) BoNT/C1. To test if BoNT/C1 ad retains neuron-specific targeting without concomitant toxic host responses, we evaluated the localization, activity, and toxicity of BoNT/C1 ad in vitro and in vivo. In neuronal cultures, BoNT/C1 ad light chain is rapidly internalized into presynaptic compartments, but does not cleave SNARE proteins nor impair spontaneous neurotransmitter release. In mice, systemic administration resulted in the specific co-localization of BoNT/C1 ad with diaphragmatic motor nerve terminals. The mouse LD50of BoNT/C1 ad is 5 mg/kg, with transient neurological symptoms emerging at sub-lethal doses. Given the low toxicity and highly specific neuron-targeting properties of BoNT/C1 ad, these data suggest that BoNT/C1 ad can be useful as a molecular vehicle for drug delivery to the neuronal cytoplasm.
authors.affiliation: Department of Biochemistry and Molecular Pharmacology, New York University School of Medicine, New York, NY, 10016, USA.|CytoDel LLC, New York, NY, 10027, USA.|The United States Army Medical Research Institute of Chemical Defense, Aberdeen Proving Ground, MD 21010, USA.
authors.forename: Edwin J
authors.initials: EJ
authors.lastname: Vazquez-Cintron
authors.affiliation: The United States Army Medical Research Institute of Chemical Defense, Aberdeen Proving Ground, MD 21010, USA.
authors.forename: Phillip H
authors.initials: PH
authors.lastname: Beske
authors.affiliation: Department of Biochemistry and Molecular Pharmacology, New York University School of Medicine, New York, NY, 10016, USA.|CytoDel LLC, New York, NY, 10027, USA.
authors.forename: Luis
authors.initials: L
authors.lastname: Tenezaca
authors.affiliation: Excet, Inc., 6225 Brandon Ave., Suite 360, Springfield, VA, 22150, USA.
authors.forename: Bao Q
authors.initials: BQ
authors.lastname: Tran
authors.affiliation: The United States Army Medical Research Institute of Chemical Defense, Aberdeen Proving Ground, MD 21010, USA.
authors.forename: Jonathan M
authors.initials: JM
authors.lastname: Oyler
authors.affiliation: The United States Army Medical Research Institute of Chemical Defense, Aberdeen Proving Ground, MD 21010, USA.
authors.forename: Elliot J
authors.initials: EJ
authors.lastname: Glotfelty
authors.affiliation: Department of Biochemistry and Molecular Pharmacology, New York University School of Medicine, New York, NY, 10016, USA.
authors.forename: Christopher A
authors.initials: CA
authors.lastname: Angeles
authors.affiliation: Department of Biochemistry and Molecular Pharmacology, New York University School of Medicine, New York, NY, 10016, USA.
authors.forename: Aurelia
authors.initials: A
authors.lastname: Syngkon
authors.affiliation: Department of Infectious Diseases and Global Health, Tufts University Clinical and Translational Science Institute, North Grafton, MA, 01536, USA.
authors.forename: Jean
authors.initials: J
authors.lastname: Mukherjee
authors.affiliation: Centers for Disease Control and Prevention, National Center for Environmental Health/Agency for Toxic Substances and Disease Registry, Atlanta, GA 30341, USA.
authors.forename: Suzanne R
authors.initials: SR
authors.lastname: Kalb
authors.affiliation: Department of Biochemistry and Molecular Pharmacology, New York University School of Medicine, New York, NY, 10016, USA.|CytoDel LLC, New York, NY, 10027, USA.|Department of Orthopaedic Surgery, New York University Hospital for Joint Diseases, New York, NY, 10016, USA.
authors.forename: Philip A
authors.initials: PA
authors.lastname: Band
authors.affiliation: The United States Army Medical Research Institute of Chemical Defense, Aberdeen Proving Ground, MD 21010, USA.
authors.forename: Patrick M
authors.initials: PM
authors.lastname: McNutt
authors.affiliation: Department of Infectious Diseases and Global Health, Tufts University Clinical and Translational Science Institute, North Grafton, MA, 01536, USA.
authors.forename: Charles B
authors.initials: CB
authors.lastname: Shoemaker
authors.affiliation: Department of Biochemistry and Molecular Pharmacology, New York University School of Medicine, New York, NY, 10016, USA.
authors.forename: Konstantin
authors.initials: K
authors.lastname: Ichtchenko
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D001905:Botulinum Toxins / Q000235:genetics / Q000378:metabolism* / Q000633:toxicity; D002478:Cells, Cultured; D019281:Dimerization; D004337:Drug Carriers / Q000737:chemistry*; D005260:Female; D007928:Lethal Dose 50; D051379:Mice; D018613:Microscopy, Confocal; D000066450:Mouse Embryonic Stem Cells / Q000166:cytology; D009474:Neurons / Q000166:cytology / Q000378:metabolism; D009435:Synaptic Transmission / Q000187:drug effects; D050825:Synaptosomal-Associated Protein 25 / Q000378:metabolism; D050827:Syntaxin 1 / Q000378:metabolism
chemical_list: D004337:Drug Carriers; D050825:Synaptosomal-Associated Protein 25; D050827:Syntaxin 1; D001905:Botulinum Toxins; C096324:botulinum toxin type C