A Three-Monoclonal Antibody Combination Potently Neutralizes BoNT/G Toxin in Mice

Area of Science:

• Immunology
• Neuroscience
• Biotechnology

Background:

• Equine-derived antitoxin (BAT®) is the sole treatment for botulism due to botulinum neurotoxin serotype G (BoNT/G).
• BAT® is a foreign protein associated with adverse effects and is not renewable.
• Development of a safe, potent, and renewable antitoxin is crucial.

Purpose of the Study:

• To generate humanized monoclonal antibodies (mAbs) against BoNT/G.
• To develop a safe, potent, and renewable antitoxin for BoNT/G botulism.
• To establish a basis for a fully recombinant heptavalent botulinum antitoxin.

Main Methods:

• Generated yeast-displayed single chain Fv (scFv) libraries from mice immunized with BoNT/G.
• Screened libraries using fluorescence-activated cell sorting (FACS) to isolate BoNT/G-binding scFvs.
• Humanized and affinity-matured selected mAbs, creating IgG antibodies, and tested combinations for efficacy in mice.

Main Results:

• Isolated fourteen scFvs with high affinity (K _D 3.86 nM to 103 nM) for BoNT/G.
• Developed five humanized mAbs (hu6G6.2, hu6G7.2, hu6G9.1, hu6G10, hu6G11.2) with high IgG affinities (K _D 51 pM to 8 pM).
• Three mAb combinations provided complete protection in mice against 10,000 LD ₅₀ s of BoNT/G at a low dose (6.25 μg/mouse).

Conclusions:

• Humanized mAbs are effective against BoNT/G and represent a viable alternative to equine antitoxin.
• These mAbs demonstrate potential for diagnosing and treating BoNT/G botulism.
• The developed mAbs form the foundation for a recombinant heptavalent antitoxin, replacing the current equine product.