Abstract:
Receptor tyrosine kinases (RTKs) wield great influence over cell growth, and their aberrant behaviour is regularly associated with disease. The RTK fibroblast growth factor receptor 1 (FGFR1) has been implicated in multiple types of cancer and is a validated drug target. However, well documented detrimental side effects for the FGFR family come from a lack of drug selectivity, as well as inhibiting a protein so crucial for normal cell signalling. There is a growing interest in targeting protein-protein interfaces (PPIs) with the potential to target selected pathways with high selectivity. In FGFR1, there are three pathways that could be selected for PPI disruption. A major challenge in targeting PPIs with small molecule inhibitors is the nature of these target sites, which are typically broad and relatively featureless. Being small and having high ligand efficiency, fragment compounds show promise in probing for these types of sites. In fragment-based drug discovery (FBDD), fragment “hits” identified in screening experiments can be evolved into functionalised drugs with high specificity towards their target. To investigate their utility in fragment screening FGFR1, differential scanning fluorimetry (DSF) and WaterLOGSY nuclear magnetic resonance (NMR) were used in parallel as preliminary screening methods in this project. These yielded contrasting results, with DSF producing a small number of hits while WaterLOGSY NMR provided a much larger selection. Two fragment “hits” visualised by X-ray crystallography bind to the ATP site of the kinase domain – no surface (PPI) sites were identified. To probe the utility of DSF and WaterLOGSY NMR against a very different active site, the results of a previous fragment screening campaign against aldo-keto reductase 1C3 (AKR1C3) were compared to those of FGFR1. AKR1C3 binds fragments in a deep, buried cavity dominated by an oxyanion site while FGFR1 binding, by contrast, is to a shallow surface exposed groove. DSF in particular showed greater utility against the AKR1C3 active site. This project developed the utility of DSF, WaterLOGSY NMR, and X-ray crystallography in a drug discovery pipeline. Furthermore, the results highlight the necessity for parallel and complementary screening techniques, as well as tailoring screening programmes depending on the target.