Reactivity of Covalent Fragments and Their Role in Fragment Based Drug Discovery.

Fragment based drug discovery has long been used for the identification of new ligands and interest in targeted covalent inhibitors has continued to grow in recent years, with high profile drugs such as osimertinib and sotorasib gaining FDA approval. It is therefore unsurprising that covalent fragment-based approaches have become popular and have recently led to the identification of novel targets and binding sites, as well as ligands for targets previously thought to be 'undruggable'. Understanding the properties of such covalent fragments is important, and characterizing and/or predicting reactivity can be highly useful. This review aims to discuss the requirements for an electrophilic fragment library and the importance of differing warhead reactivity. Successful case studies from the world of drug discovery are then be examined.

Fragment-based drug discovery-the importance of high-quality molecule libraries.

Fragment-based drug discovery (FBDD) is now established as a complementary approach to high-throughput screening (HTS). Contrary to HTS, where large libraries of drug-like molecules are screened, FBDD screens involve smaller and less complex molecules which, despite a low affinity to protein targets, display more 'atom-efficient' binding interactions than larger molecules. Fragment hits can, therefore, serve as a more efficient start point for subsequent optimisation, particularly for hard-to-drug targets. Since the number of possible molecules increases exponentially with molecular size, small fragment libraries allow for a proportionately greater coverage of their respective 'chemical space' compared with larger HTS libraries comprising larger molecules. However, good library design is essential to ensure optimal chemical and pharmacophore diversity, molecular complexity, and physicochemical characteristics. In this review, we describe our views on fragment library design, and on what constitutes a good fragment from a medicinal and computational chemistry perspective. We highlight emerging chemical and computational technologies in FBDD and discuss strategies for optimising fragment hits. The impact of novel FBDD approaches is already being felt, with the recent approval of the covalent KRASG12C inhibitor sotorasib highlighting the utility of FBDD against targets that were long considered undruggable.

Acrylamide fragment inhibitors that induce unprecedented conformational distortions in enterovirus 71 3C and SARS-CoV-2 main protease.

RNA viruses are critically dependent upon virally encoded proteases to cleave the viral polyproteins into functional proteins. Many of these proteases exhibit a similar fold and contain an essential catalytic cysteine, offering the opportunity to inhibit these enzymes with electrophilic small molecules. Here we describe the successful application of quantitative irreversible tethering (qIT) to identify acrylamide fragments that target the active site cysteine of the 3C protease (3Cpro) of Enterovirus 71, the causative agent of hand, foot and mouth disease in humans, altering the substrate binding region. Further, we re-purpose these hits towards the main protease (Mpro) of SARS-CoV-2 which shares the 3C-like fold and a similar active site. The hit fragments covalently link to the catalytic cysteine of Mpro to inhibit its activity. We demonstrate that targeting the active site cysteine of Mpro can have profound allosteric effects, distorting secondary structures to disrupt the active dimeric unit.