Quinoline-based Inhibitors of BoNT/A LC

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DESCRIPTION (provided by applicant): The botulinum neurotoxins (BoNTs) are the most poisonous biological substances known. The lethal intravenous dose of BoNT serotype A (BoNT/A) in humans is 1-5 ng/kg. If accidental exposure to BoNT occurs (e.g., from contaminated foodstuffs), loss of life or life-threatening paralysis can occur. Most importantly, the BoNTs have already been "weaponized" in a highly toxic aerosol form, and they consequently pose a significant threat to both civilian and military populations. Once ingested, BoNTs target the peripheral cholinergic nerve endings and cause death by interrupting autonomic nerve function. The zinc-dependent endopeptidase light chain (LC) portion of BoNTs impairs neuronal exocytosis through proteolysis of essential SNARE (soluble N-ethylmaleimide-sensitive factor attachment protein receptor) components of neurotransmission. The overall goal or this project is to develop small molecule inhibitors of the BoNT/A light chain (LC) metalloprotease activity to treat botulinum poisoning. Our strategy is to optimize a novel chemotype of BoNT/A LC inhibitors to improve potency, selectivity, and drug-like properties. In preliminary studies, we refined the structures to provide a new starting point for medicinal chemistry based on a chemical core with more drug-like properties than the original compound series and with ten-fold better potency. Significant activity was also observed in a chick neuronal cell model of BoNT/A intoxication. These validated early lead compounds are more suitable lead compounds for BoNT/A inhibitor drug discovery than were the original series, and they form the basis for this optimization program to generate an advanced lead compound. Our approach in Phase I is to use proven techniques of medicinal and parallel synthetic chemistry to optimize the potency and selectivity of the lead series of compounds and produce "drug-like" molecules with the potential to advance to pre-clinical development. In an iterative process, we will probe these focused compound libraries for structural features that contribute to tighter binding and more potent inhibition of the BoNT/A metalloprotease by measuring the enzymatic and cellular activities, as well as the specificity of the enzyme inhibitors. We will correlate the in vitro activity of the compounds against BoNT/A LC with activity in a cellular model of BoNT/A action, and use the most promising members of the series to validate the inhibitors in an in vivo mouse model of BoNT/A intoxication. At the end of Phase I we will define an advanced lead compound, which will be selected from the group of in vivo-validated inhibitors that displays suitable in vitro pharmacokinetic properties for further preclinical development. In Phase II, we will further optimize and evaluate the lead for in vivo efficacy, pharmacokinetic properties, toxicity and safety pharmacology in two species, and develop it into a pre-IND clinical candidate, suitable for human clinical trials (Phase III). PUBLIC HEALTH RELEVANCE: The botulinum neurotoxins are some of the most poisonous biologic substances known. Loss of life or life-threatening paralysis can occur following exposure to these neurotoxins from contaminated foodstuffs or acts of bioterrorism. This proposal describes the preparation and development of novel drugs to treat botulinum poisoning.