Structure-Based Design of Cdk4/6-Specific Inhibitors

摘要

The CDK4/6 kinase has elevated activity in spontaneous breast cancer, making it an excellent candidate for targeted inhibition for the treatment of breast cancer. We proposed to prepare a CDK4/6-specific kinase inhibitor that is based on the naturally occurring CDK4/6 specific inhibitory proteins of the INK4 family. Since the INK4 proteins are notoriously unstable, we have been working towards the preparation and structural characterization of INK4 proteins with improved thermostability that could be used as a scaffold to prepare peptide sub-fragments that may serve as a scaffold to prepare small molecule CDK4/6- specific inhibitors. During the funding period, we successfully prepared p18INK4c proteins with increased thermostability and have characterized the structure of a subset of the more thermostable p18INK4c proteins. This structure-function analysis revealed the chemical basis for the increased stability properties of the more thermostable p18INK4c proteins. Significantly, a thermostable F71N mutant protein also showed enhanced CDK6 interaction and cell cycle inhibitory activity in vivo, as measured using co- immunoprecipitation and transient transfection assays, respectively. Together, our studies show that a structure-based approach to increase the thermodynamic stability of INK4 proteins can be exploited to prepare more biologically active molecules with potential applications for the development of molecules to treat INK4-mediated cancers.