The discovery of high affinity and metabolically stable allosteric cyclin-dependent kinase 2 (CDK2) inhibitors from screening through lead optimization

Despite the status of cyclin-dependent kinase 2 (CDK2) as a validated target for both anticancer and contraceptive indications, a CDK2 inhibitor with exquisite selectivity has been historically challenging, largely due to the structural similarity of the ATP-binding site where most kinase inhibitors bind. We previously discovered an allosteric pocket in CDK2 with potential to bind a compound with desirable selectivity. Using high-throughput and virtual screening methods, we discovered and structurally confirmed an anthranilic acid scaffold that binds this pocket with high affinity. We previously reported that these allosteric CDK2 inhibitors demonstrate a negative cooperative relationship with cyclin binding, are selective for CDK2 over the structurally similar kinase CDK1 and show potential as a non-hormonal contraceptive agent. In this work, we describe our screening and lead optimization efforts that led to the discovery of compounds in this series like EF-4-177 with nanomolar affinity for CDK2. EF-4-177 is metabolically stable with a desirably long ½ life and adequate tissue distribution in mice, demonstrating the potential of this series as a therapeutic. This work details the discovery of the highest affinity allosteric CDK inhibitors reported and shows promise for further development of this series to yield an efficacious and selective allosteric CDK2 inhibitor.