A Three Monoclonal Antibody Combination Potently Neutralizes Multiple Botulinum Neurotoxin Serotype E Subtypes

Garcia-Rodriguez C., Razai A., Geren I. N. , Lou J., Conrad F., Wen W., ...More

TOXINS, vol.10, no.3, 2018 (Peer-Reviewed Journal) identifier identifier identifier

  • Publication Type: Article / Article
  • Volume: 10 Issue: 3
  • Publication Date: 2018
  • Doi Number: 10.3390/toxins10030105
  • Journal Name: TOXINS
  • Journal Indexes: Science Citation Index Expanded, Scopus
  • Keywords: botulinum neurotoxin, oligoclonal antibodies, serotype E botulism, recombinant antibodies, antibody engineering, mouse neutralization assay, botulinum antitoxin, DOMAIN-BASED ASSAYS, CLOSTRIDIUM-BOTULINUM, CRYSTAL-STRUCTURE, CHAIN ANTIBODIES, E STRAINS, BINDING, BUTYRICUM, AFFINITY, GENE, IDENTIFICATION


Human botulism is most commonly caused by botulinum neurotoxin (BoNT) serotypes A, B, and E. For this work, we sought to develop a human monoclonal antibody (mAb)-based antitoxin capable of binding and neutralizing multiple subtypes of BoNT/E. Libraries of yeast-displayed single chain Fv (scFv) antibodies were created from the heavy and light chain variable region genes of humans immunized with pentavalent-toxoid- and BoNT/E-binding scFv isolated by Fluorescence-Activated Cell Sorting (FACS). A total of 10 scFv were isolated that bound one or more BoNT/E subtypes with nanomolar-level equilibrium dissociation constants (K-D). By diversifying the V-regions of the lead mAbs and selecting for cross-reactivity, we generated three scFv that bound all four BoNT/E subtypes tested at three non-overlapping epitopes. The scFvs were converted to IgG that had K-D values for the different BoNT/E subtypes ranging from 9.7 nM to 2.28 pM. An equimolar combination of the three mAbs was able to potently neutralize BoNT/E1, BoNT/E3, and BoNT/E4 in a mouse neutralization assay. The mAbs have potential utility as therapeutics and as diagnostics capable of recognizing multiple BoNT/E subtypes. A derivative of the three-antibody combination (NTM-1633) is in pre-clinical development with an investigational new drug (IND) application filing expected in 2018.