The long-term goal of this project is to develop a robust proteomics approach to identify targets of small-molecule compounds and drugs in live cells. Phenotypic screens are commonly used to identify novel bioactive small-molecule probes and develop new therapeutic approaches. The major bottleneck in forward chemical-genomics screens is target identification and validation of novel small-molecule probes. Strikingly, targets are still unknown for many FDA-approved drugs and small-molecule """"""""tool"""""""" compounds. Identifying both the therapeutic targets and """"""""off targets"""""""" allows optimization of a compound's selectivity and should decrease potential side effects in the eventual drug. Our proposal describes a novel strategy to use small-molecule inhibitors as in vivo targeting moieties to biotinylate their molecular targets and their protein complexes for proteomic identification. This novel approach is distinct from traditional biochemical approaches and may allow the identification of much weaker affinity binders and higher order protein-protein interactions. As our approach explores spatial relationships in multi-protein complexes when a small-molecule probe is bound to its protein target, it may provide a novel perspective to characterize the small molecule's mechanisms of action.
The discovery of novel therapeutics relies on understanding the mechanisms of actions of small- molecule probes and drugs. Current approaches for target identification rely on protein fragments or extracted cell lysates under non-native conditions. We propose to develop approaches to study protein targets of small-molecule probes within the cell. Such tools will resolve a major bottleneck in drug discovery programs and can accelerate the discovery of new therapies.