title: Botulinum neurotoxin serotype A specific cell-based potency assay to replace the mouse bioassay.
abstract: Botulinum neurotoxin serotype A (BoNT/A), a potent therapeutic used to treat various disorders, inhibits vesicular neurotransmitter exocytosis by cleaving SNAP25. Development of cell-based potency assays (CBPAs) to assess the biological function of BoNT/A have been challenging because of its potency. CBPAs can evaluate the key steps of BoNT action: receptor binding, internalization-translocation, and catalytic activity; and therefore could replace the current mouse bioassay. Primary neurons possess appropriate sensitivity to develop potential replacement assays but those potency assays are difficult to perform and validate. This report describes a CBPA utilizing differentiated human neuroblastoma SiMa cells and a sandwich ELISA that measures BoNT/A-dependent intracellular increase of cleaved SNAP25. Assay sensitivity is similar to the mouse bioassay and measures neurotoxin biological activity in bulk drug substance and BOTOX® product (onabotulinumtoxinA). Validation of a version of this CBPA in a Quality Control laboratory has led to FDA, Health Canada, and European Union approval for potency testing of BOTOX®, BOTOX® Cosmetic, and Vistabel®. Moreover, we also developed and optimized a BoNT/A CBPA screening assay that can be used for the discovery of novel BoNT/A inhibitors to treat human disease.
authors.affiliation: Department of Biological Sciences, Allergan Inc., Irvine, California, United States of America. fernandez_ester@allergan.com
authors.forename: Ester
authors.initials: E
authors.lastname: Fernández-Salas
authors.forename: Joanne
authors.initials: J
authors.lastname: Wang
authors.forename: Yanira
authors.initials: Y
authors.lastname: Molina
authors.forename: Jeremy B
authors.initials: JB
authors.lastname: Nelson
authors.forename: Birgitte P S
authors.initials: BP
authors.lastname: Jacky
authors.forename: K Roger
authors.initials: KR
authors.lastname: Aoki
mesh_terms: D000818:Animals; D000911:Antibodies, Monoclonal / Q000737:chemistry; D001681:Biological Assay / Q000379:methods*; D019274:Botulinum Toxins, Type A / Q000737:chemistry / Q000502:physiology*; D045744:Cell Line, Tumor; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods; D006801:Humans; D007700:Kinetics; D051379:Mice; D009474:Neurons / Q000378:metabolism; D009498:Neurotoxins / Q000737:chemistry; D016716:PC12 Cells; D011786:Quality Control; D051381:Rats; D012680:Sensitivity and Specificity; D020349:Surface Plasmon Resonance / Q000379:methods; D050825:Synaptosomal-Associated Protein 25 / Q000235:genetics* / Q000378:metabolism
chemical_list: D000911:Antibodies, Monoclonal; D009498:Neurotoxins; C494209:Snap25 protein, mouse; D050825:Synaptosomal-Associated Protein 25; D019274:Botulinum Toxins, Type A