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Starting date:
1st September 2010
CBtype A
botulinum toxin
54 months
Coordinator: Ministere de la Défense (France)
Partners: 8 Partners (France, Germany, UK, Finland)

Project/SA Objectives

Botulinum neurotoxins (BoNTs) are among the most toxic substances known, whether of biological or chemical origin, and they are part of the “dirty dozen” agents listed as possible bioweapons. Beside voluntarily contamination, naturally-occurring food intoxications, though rare but often severe, are still encountered and intoxication due to the cosmetic use of an unauthorized BoNT has also been reported. Despite extensive research, no small synthetic molecule has been validated for therapeutic use against BoNTs, and Europe relies on an old stockpile of horse polyclonal antibodies as the sole BoNTs-neutralizing medicines. Recombinant antibodies are a highly successful new class of therapeutic molecules, produced by biotechnologies, showing an exponential-like growth. The goal of AntiBotABE is to isolate recombinant antibodies neutralizing BoNT A, B and E as these types are lethal for humans. The heavy and light chains will be targeted for a synergistic effect, thus six recombinant antibodies have to be isolated. For this project, the strategy that allowed to previously isolate neutralizing antibodies against ricin and the lethal toxin of anthrax will be re-utilized.

Description of the work

This project will start with recombinant proteins, part of the light or heavy chains of BoNT A, B and E and utilized as immunogens. The lymphocytes of macaques, immunized with these immunogens, will be used for the construction of immune phage-displayed libraries. These libraries will be screened to isolate high-affinity antibody fragments (scFvs), which will be human-like due to the phylogenetic proximity between macaques and humans. BoNTs present sub-types (A1 and A2, B1 and B2 for instance) and scFvs reacting with these various sub-types will be isolated with a specially-designed panning procedure. To test for neutralization capacities, scFvs directed against heavy chains will be tested in ex vivo assays, and the scFvs directed against the heavy chains will be tested in vitro. At the end of these steps, the scFvs with best neutralizing profile will be selected and super-humanized.

In the third part of the project, neutralizing, super-humanized scFvs will be expressed as full-sized IgGs and tested in a standardized protection model to verify their efficacy against several strains for each targeted serotype.  At various steps of the project, our results will be communicated to the first responders more particularly involved against biothreats.

Expected results

The ideal result is an oligoclonal cocktail of 6 recombinant, super-humanized IgGs, neutralizing the neurotoxins secreted by all strains of Clostridium botulinum A. B and E. These IgGs will then be developed as medicine registered by EMEA, on credits provided by EDA. This medicine is to become available for biodefense  primarily, but also for natural cases of botulinum intoxications in Europe. This dual-use availability, and information given to practitioners in the course of the project, will ensure real improvement in botulism treatment and its perception by EU citizens.

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