Identification of the first noncompetitive SARM1 inhibitors

Sterile Alpha and Toll Interleukin Receptor Motif–containing protein 1 (SARM1) is a key therapeutic target for diseases that exhibit Wallerian–like degeneration; Wallerian degeneration is characterized by degeneration of the axon distal to the site of injury. These diseases include traumatic brain injury, peripheral neuropathy, and neurodegenerative diseases. SARM1 promotes neurodegeneration by catalyzing the hydrolysis of NAD⁺ to form a mixture of ADPR and cADPR. Notably, SARM1 knockdown prevents degeneration, indicating that SARM1 inhibitors will likely be efficacious in treating these diseases. Consistent with this hypothesis is the observation that NAD⁺ supplementation is axoprotective. To identify compounds that block the NAD⁺ hydrolase activity of SARM1, we developed and performed a high–throughput screen (HTS). This HTS assay exploits an NAD⁺ analog, etheno–NAD⁺ (ENAD) that fluoresces upon cleavage of the nicotinamide moiety. From this screen, we identified berberine chloride and zinc chloride as the first noncompetitive inhibitors of SARM1. Though modest in potency, the noncompetitive mode of inhibition, suggests the presence of an allosteric binding pocket on SARM1 that can be targeted for future therapeutic development. Additionally, zinc inhibition and site–directed mutagenesis reveals that cysteines 629 and 635 are critical for SARM1 catalysis, highlighting these sites for the design of inhibitors targeting SARM1.