Combinatorial Strategies and Hypothesis-Based Drug Design in Drug Discovery Targeted to the Protease and Channel Activities of Botulinum Toxin A.

摘要

The ultimate goal of this program is to understand in detail the mechanisms by which botulinum neurotoxins (BoNT) abrogate neurotransmitter release. One facet is focused on the channel-forming domain of the heavy chain (HC). The aim is to identify open channel blockers as a single class of drugs that would be effective against all BoNT isoforms. A major objective is to seek direct demonstration that the HC acts as the molecular conduit for the light chain (LC) thereby allowing the protease activity to reach the cytosol where it acts. Accordingly, purified HC is first reconstituted in lipid bilayers and the direct translocation of isolated LC through the HC channel is measured by single channel recordings and also by direct analytical determination of the LC protease activity. A second facet of the program involves the concept that the peptide products of substrate proteolysis by BoNTs uncouple excitation from secretion pointing to new means of intervention. This notion, discovered for BoNT A, appears to be valid for other BoNT isoforms thereby yielding a diverse repertoire of peptide sequences that may provide insights into the molecular interactions between partner proteins involved in membrane fusion. It is anticipated that this concerted and focused approach will uncover lead compounds that may be developed into selective drugs targeted to prevent or relieve the neurotoxic actions of BoNT.