botulinum neurotoxin serotype a specific cell-based potency assay to replace the mouse bioassay肉毒神经毒素血清型特定细胞效力试验代替鼠标生物测定.pdfVIP

Botulinum Neurotoxin Serotype a Specific Cell-Based Potency Assay to Replace the Mouse Bioassay ´ Ester Fernandez-Salas*, Joanne Wang, Yanira Molina, Jeremy B. Nelson, Birgitte P. S. Jacky, K. Roger Aoki Department of Biological Sciences, Allergan Inc., Irvine, California, United States of America 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 BOTOXH 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 BOTOXH, BOTOXH Cosmetic, and VistabelH. 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. ´ Citation: Fernandez-Salas E, Wang J, Molina Y, Nelson J