Mechanistic Characterization of a Small Molecule as a Direct NLRP3 Inhibitor via Binding to the NACHT Domain

The NLRP3 inflammasome has recently emerged as a viable
drug target,
and small-molecule inhibitors of this protein complex are being actively
tested in preclinical disease models. However, the mechanisms of action
for the majority of these compounds remain unclear. Our recent medicinal
chemistry campaign led to the discovery of several potent lead NLRP3
inhibitors from a distinct chemical scaffold. Herein, further characterization
using biophysical, biochemical, chemical biology, and molecular biology
approaches of one of the lead inhibitors revealed direct binding to
the NACHT domain of the NLRP3 protein. The studies also suggested
potentially multiple binding sites within the NACHT domain that can
be targeted by different small-molecule inhibitors. Furthermore, an
activity-based probe with high labeling efficacy was developed and
can serve as a valuable tool to contribute to ongoing efforts in understanding
NLRP3 biology and in developing NLRP3-targeted therapies for various
diseases.