Structure Based Discovery of Pan Active Botulinum Neurotoxin Inhibitors

Abstract

Clostridium botulinum neurotoxins (BoNTs) released by the bacterium Clostridium botulinum are the most potent toxins causing the fatal disease called botulism. There are seven distinct serotypes of BoNTs (A to G) released by various strains of botulinum. They all have high sequence homology and similar three-dimensional structure. The toxicity of BoNT follows a four-step process-binding, internalization, translocation, and cleavage of its target protein, one of the three components of the SNARE complex (Soluble N-ethylmaleimde-sensitive factor attachment protein receptor) required for membrane docking and neurotransmitter release. Cleavage of one of the three proteins causes blockage of neurotransmitter release leading to flaccid paralysis. Though anyone of the above four steps could be a target for developing antidotes for botulism, the catalytic domain is the most suitable target for post exposure treatment. Of the seven serotypes BoNT/A, B, E and probably F affect humans, with BoNT/A considered to be the most potent. Development of drugs for botulism is focused on serotype specific inhibitors, but pan-active inhibitor acting on several serotypes is preferable since it is difficult to identify the serotype before the treatment, especially since there is at least a 36 h window before botulism can be diagnosed. Using structure-based drug discovery, we have developed three heptapeptides based on the SNARE proteins which inhibit BoNT/A, B and E equally well. Probable reasons for pan-activity of these peptides are discussed.

Toxicology and pharmacology of botulinum and tetanus neurotoxins: an update

“…In general, up to the present time, inhibitors of the metalloprotease activity of BoNTs were found to be effective only at levels (A) and/or (B) with K i or IC 50 values in the 10 –6 – 10 −8 Molar range. (Caglič et al 2014 ; Kumar et al 2016 ; Bremer et al 2017 ; Vieni et al 2018 ; Patel et al 2018 ; Garland et al 2019 ; Amezcua et al 2021 ; Lin et al 2021 ; Turner et al 2021a , b ). None the less, these studies are very important because they provide the molecular basis to design novel and specific inhibitors of the BoNT active site acting in cultured neurons in the nanomolar range, a value which is a prerequisite figure to proceed to animal studies (level C).…”