A three monoclonal antibody combination potently neutralizes multiple botulinum neurotoxin serotype F subtypes

• Published in: PloS one 2017-03, Vol.12 (3), p.e0174187-e0174187
• Main Authors: Fan, Yongfeng, Garcia-Rodriguez, Consuelo, Lou, Jianlong, Wen, Weihua, Conrad, Fraser, Zhai, Wenwu, Smith, Theresa J, Smith, Leonard A, Marks, James D
• Format: Article
• Language: English
• Subjects: Accreditation; Amino Acid Sequence; Animals; Antibodies, Monoclonal - immunology; Antigenic determinants; Antitoxins; Antitoxins - genetics; Antitoxins - immunology; Biology and Life Sciences; Botulinum toxin type F; Botulinum Toxins - immunology; Botulism; Botulism - diagnosis; Botulism - therapy; Catalytic Domain - immunology; Cloning; Clostridium botulinum; Clostridium botulinum - metabolism; Complications and side effects; Cross Reactions - immunology; Dissociation; Epitope Mapping; Epitopes; Epitopes - immunology; Escherichia coli - genetics; Food contamination; Genetic aspects; Hospitals; Immunization; Immunoglobulin G; Immunoglobulins; Infectious diseases; Laboratory animals; Lead; Licenses; Medicine and Health Sciences; Mice; Monoclonal antibodies; Neurotoxicity syndromes; Neutralization; Physical Sciences; Proteins; Research and Analysis Methods; Risk factors; Saccharomyces cerevisiae - genetics; Serotypes; Single-Chain Antibodies - genetics; Single-Chain Antibodies - immunology; Water-borne diseases; Waterborne diseases; Yeast
• ISSN: 1932-6203; 1932-6203
• DOI: 10.1371/journal.pone.0174187

Human botulism is primarily caused by botulinum neurotoxin (BoNT) serotypes A, B and E, with around 1% caused by serotype F (BoNT/F). BoNT/F comprises at least seven different subtypes with the amino acid sequence difference between subtypes as high as 36%. The sequence differences present a significant challenge for generating monoclonal antibodies (mAbs) that can bind, detect and neutralize all BoNT/F subtypes. We used repertoire cloning of immune mouse antibody variable (V) regions and yeast display to generate a panel of 33 lead single chain Fv (scFv) mAbs that bound one or more BoNT/F subtypes with a median equilibrium dissociation constant (KD) of 4.06 × 10-9 M. By diversifying the V-regions of the lead mAbs and selecting for cross reactivity we generated five mAbs that bound each of the seven subtypes. Three scFv binding non-overlapping epitopes were converted to IgG that had KD for the different BoNT/F subtypes ranging from 2.2×10-8 M to 1.47×10-12 pM. An equimolar combination of the mAbs was able to potently neutralize BoNT/F1, F2, F4 and F7 in the mouse neutralization assay. The mAbs have potential utility as diagnostics capable of recognizing the known BoNT/F subtypes and could be developed as antitoxins to prevent and treat type F botulism.