Nucleophilic fragments vs SARM1: in situ inhibitor assembly

Recently Practical Fragments wrote about nucleophilic fragments that could react with proteins or cofactors. Previously we’ve also written about in situ chemistry, in which a protein catalyzes the formation of an inhibitor. An interesting marriage of these concepts has just been published (open access) in Mol. Cell by Robert Hughes (Disarm Therapeutics), Thomas Ve (Griffith University) and a group of international collaborators.

 

The researchers were interested in the protein SARM1, which is implicated in the axon degeneration associated with several neurodegenerative disorders. Last year the researchers published a Cell Rep. paper (also open access) in which a biochemical screen of roughly 200,000 molecules led to the discovery of isoquinoline as a 10 µM inhibitor of SARM1. Optimization led to 5-iodoisoqinoline, dubbed DSRM-3716, a 75 nM fragment-sized inhibitor. The paper goes on to demonstrate that the molecule not only prevents axonal degeneration but can even promote recovery of injured axons. The new paper explores the mechanism of action.

 

SARM1 is an NADase: it cleaves the critical cofactor nicotinamide adenine dinucleotide (NAD⁺). While using NMR to study the mechanism of inhibition, the researchers found that DSRM-3716 reacts with NAD⁺ to form the new compound shown. In this sense, DSRM-3716 acts as a prodrug, somewhat analogous to sulfanilamide antibiotics which act as PABA mimics to block folate biosynthesis.

 

What’s behind the inhibition of SARM1? A series of crystallographic and cryo-EM studies of SARM1 reveal that the protein can self-associate into multimers which are either inactive or active depending on the relative orientations of the individual proteins. NAD⁺ normally binds at the interface between two SARM1 proteins. The compound made from NAD⁺ and DSRM-3716 binds here as well, blocking further activity. The crystal structures also revealed a clear halogen bond (see here) with the iodine in DSRM-3716, explaining the increased activity over isoquinoline itself.

 

Unlike the nucleophilic fragments we wrote about last month, isoquinoline probably won’t raise too many eyebrows among medicinal chemists, as the moiety is found in a handful of approved drugs. The researchers also demonstrated that DSRM-3716 itself is selective for SARM1 in a panel of other enzymes that use NAD⁺.

 

This is a lovely case of high-throughput screening in which the hit turns out to be a fragment. Indeed, the highly charged compound that actually inhibits SARM1 would not be cell-permeable, but that's just fine since it is formed inside cells. It is worth noting that nearly 1000 approved drugs could be classified as fragments in terms of molecular weight. In the case of CNS drugs, small is beautiful, and it will be fun to watch how far DSRM-3716 derivatives will be able to advance.