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Biophysical methods such as mass spectrometry, surface plasmon resonance, nuclear magnetic resonance, and both differential scanning isothermal titration calorimetry are now well established as key components of the early drug discovery process. These approaches are used successfully for a range of activities, including assay development, primary screening, hit confirmation and detailed mechanistic characterisation of compound binding. Matching the speed, sensitivity and information content of the various techniques to the generation of critical data and information required at each phase of the drug discovery process has been key. This review describes the framework by which these methods have been applied in the drug discovery process and provides case studies to exemplify the impact.

Mutant RAS family members occur in a wide range of tumour types, and there is a great interest in identifying isoform-specific inhibitors. Here, the authors characterise two designed ankyrin repeat proteins (DARPins) that specifically inhibit the KRAS isoform by binding to the region around the KRAS-specific residue histidine 95 and show that they affect KRAS/effector interactions in different ways.

MTH1 (NUDT1) is an oncologic target involved in the prevention of DNA damage. We investigate the way MTH1 recognises its substrates and present substrate-bound structures of MTH1 for 8-oxo-dGTP and 8-oxo-rATP as examples of novel strong and weak binding substrate motifs. Investigation of a small set of purine-like fragments using 2D NMR resulted in identification of a fragment with weak potency. The protein-ligand X-Ray structure of this fragment provides insight into the role of water molecules in substrate selectivity. Wider fragment screening by NMR resulted in three new protein structures exhibiting alternative binding configurations to the key Asp-Asp recognition element of the protein. These inhibitor binding modes demonstrate that MTH1 employs an intricate yet promiscuous mechanism of substrate anchoring through its Asp-Asp pharmacophore. The structures suggest that water-mediated interactions convey selectivity towards oxidized substrates over their non-oxidised counterparts, in particular by stabilization of a water molecule in a hydrophobic environment through hydrogen bonding. These findings may be useful in the design of inhibitors of MTH1.