Probing an Allosteric Pocket of CDK2 with Small Molecules

ChemMedChem. 2017 Jan 5;12(1):33-41. doi: 10.1002/cmdc.201600474. Epub 2016 Dec 19.


The availability of well-characterized allosteric modulators is crucial for investigating the allosteric regulation of protein function. In a recently identified inactive conformation of cyclin-dependent kinase 2 (CDK2), an open allosteric pocket was detected and proposed as a site to accommodate allosteric inhibitors. Previous structure-based approaches allowed the identification of a hit compound expected to bind to this pocket. Herein we report the characterization of this compound by X-ray crystallography, which surprisingly provided a chemical structure different from that previously reported. Therefore, the compound was synthesized and completely characterized. X-ray structures of the synthesized and purchased compounds were found to be superimposable. A reaction mechanism was proposed to explain the formation of the structure indicated by crystallography. Moreover, a stereoselective synthesis was developed to evaluate the biological activity of the pure stereoisomers. Modeling studies were performed to unveil the details of the interaction with CDK2. The activity of the obtained compounds was evaluated with various biological assays. Mutagenesis experiments confirmed binding to the allosteric pocket. Finally, the allosteric ligands were shown to inhibit the growth of lung (A549) and ovarian (SKOV3) cancer cell lines. Therefore, this report presents a thorough chemical and biological characterization of the first small-molecule ligands to be used as probes to study the allosteric modulation of CDK2 activity.

Keywords: allosteric modulators; computational chemistry; cyclin-dependent kinase 2; protein kinases; structure elucidation.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Allosteric Regulation / drug effects
  • Allosteric Site / drug effects*
  • Antineoplastic Agents / chemical synthesis
  • Antineoplastic Agents / chemistry
  • Antineoplastic Agents / pharmacology*
  • Cell Line, Tumor
  • Cell Proliferation / drug effects
  • Crystallography, X-Ray
  • Cyclin-Dependent Kinase 2 / antagonists & inhibitors*
  • Cyclin-Dependent Kinase 2 / metabolism
  • Drug Screening Assays, Antitumor
  • Humans
  • Models, Molecular
  • Molecular Structure
  • Quinolines / chemical synthesis
  • Quinolines / chemistry
  • Quinolines / pharmacology*
  • Small Molecule Libraries / chemical synthesis
  • Small Molecule Libraries / chemistry
  • Small Molecule Libraries / pharmacology*


  • Antineoplastic Agents
  • Quinolines
  • Small Molecule Libraries
  • CDK2 protein, human
  • Cyclin-Dependent Kinase 2