SH2 <i>scan</i> : Mapping SH2 Domain-Ligand Binding Selectivity for Inhibitors and Degraders

Drug discovery targeting SH2 domains (key protein-protein interaction modules) has been hampered by a lack of assay systems evaluating synthetic ligand binding selectivity toward SH2 domains to reduce potential off-target effects. In addition, the molecular determinants for the synthetic ligand engagement to SH2 domains across the target class have yet to be defined. Here, we developed SH2<i>scan</i>, a high-throughput competition binding assay platform to quantify ligand-SH2 domain interactions, covering >80% of the target class. We uncovered unique binding selectivity profiles and quantified a broad range of dissociation constants (<i>K</i>_Ds) for 9 synthetic ligands of SH2 domains from the scientific literature with a range of reported primary targets. These results demonstrate that SH2<i>scan</i> can be used to design more selective compounds targeting the SH2 domains. The platform can be further leveraged for the discovery of new molecular probes for the dissection of cellular protein-protein interaction networks.