Symptomatic treatment of botulism with a clinically approved small molecule.

Edwin Vazquez-Cintron,Kathleen Pagarigan,M Ross Pennington,James Machamer,Celinia Ondeck,Kyle Kelly,Paige Bodner,Brittany Winner,Patrick Mcnutt

doi:10.1172/jci.insight.132891

Abstract

Botulinum neurotoxins (BoNTs) are potent neuroparalytic toxins that cause mortality through respiratory paralysis. The approved medical countermeasure for BoNT poisoning is infusion of antitoxin immunoglobulins. However, antitoxins have poor therapeutic efficacy in symptomatic patients; thus, there is an urgent need for treatments that reduce the need for artificial ventilation. We report that the US Food and Drug Administration-approved potassium channel blocker 3,4-diaminopyridine (3,4-DAP) reverses respiratory depression and neuromuscular weakness in murine models of acute and chronic botulism. In ex vivo studies, 3,4-DAP restored end-plate potentials and twitch contractions of diaphragms isolated from mice at terminal stages of BoNT serotype A (BoNT/A) botulism. In vivo, human-equivalent doses of 3,4-DAP reversed signs of severe respiratory depression and restored mobility in BoNT/A-intoxicated mice at terminal stages of respiratory collapse. Multiple-dosing administration of 3,4-DAP improved respiration and extended survival at up to 5 LD50 BoNT/A. Finally, 3,4-DAP reduced gastrocnemius muscle paralysis and reversed respiratory depression in sublethal models of serotype A-, B-, and E-induced botulism. These findings make a compelling argument for repurposing 3,4-DAP to symptomatically treat symptoms of muscle paralysis caused by botulism, independent of serotype. Furthermore, they suggest that 3,4-DAP is effective for a range of botulism symptoms at clinically relevant time points.

Highlights

Botulinum neurotoxins (BoNTs) are a family of potent protein neurotoxins naturally expressed by members of the Clostridium genus of anaerobic bacteria [1]
We found that human-equivalent doses of 3,4-DAP reversed limb and respiratory paralysis caused by each BoNT serotype
We previously reported that 3,4-DAP increased nerve-elicited end-plate potentials (EPP) and twitch contraction amplitudes in hemidiaphragm preparations paralyzed by BoNT serotypes A, B, or E [24]

Summary

Introduction

Botulinum neurotoxins (BoNTs) are a family of potent protein neurotoxins naturally expressed by members of the Clostridium genus of anaerobic bacteria [1]. BoNTs are categorized into 7 canonical antigenic serotypes (A–G) and further stratified into over 40 subtypes based on primary sequence divergence [3]. HC mediates highly selective and efficient binding to endosomal receptors on the presynaptic membrane of peripheral neurons [5, 6]. Following internalization via synaptic endocytosis, LC translocates across the endosomal membrane to the presynaptic cytosol, where it cleaves neuronal SNARE (soluble N-ethylmaleimide–sensitive factor attachment protein receptor) proteins essential for neurotransmitter exocytosis [2]. Cleavage of SNAP25 by serotypes A, C, or E; SYB1–3 by serotypes B, D, F, and G; or STX1 by serotype C inhibits assembly of the SNARE protein complex that mediates synaptic vesicle fusion with the presynaptic membrane, thereby preventing neurotransmitter release [7, 8]

Methods

Results

Conclusion