4-arylazo-3,5-diamino-1H-pyrazole CDK inhibitors: SAR study, crystal structure in complex with CDK2, selectivity, and cellular effects

J Med Chem. 2006 Nov 2;49(22):6500-9. doi: 10.1021/jm0605740.


In a routine screening of our small-molecule compound collection we recently identified 4-arylazo-3,5-diamino-1H-pyrazoles as a novel group of ATP antagonists with moderate potency against CDK2-cyclin E. A preliminary SAR study based on 35 analogues suggests ways in which the pharmacophore could be further optimized, for example, via substitutions in the 4-aryl ring. Enzyme kinetics studies with the lead compound and X-ray crystallography of an inhibitor-CDK2 complex demonstrated that its mode of inhibition is competitive. Functional kinase assays confirmed the selectivity toward CDKs, with a preference for CDK9-cyclin T1. The most potent inhibitor, 4-[(3,5-diamino-1H-pyrazol-4-yl)diazenyl]phenol 31b (CAN508), reduced the frequency of S-phase cells of the cancer cell line HT-29 in antiproliferation assays. Further observed cellular effects included decreased phosphorylation of the retinoblastoma protein and the C-terminal domain of RNA polymerase II, inhibition of mRNA synthesis, and induction of the tumor suppressor protein p53, all of which are consistent with inhibition of CDK9.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Antimetabolites
  • Azo Compounds / chemical synthesis*
  • Azo Compounds / pharmacology*
  • Bromodeoxyuridine
  • Cell Cycle / drug effects
  • Cell Line, Tumor
  • Cell Proliferation / drug effects
  • Crystallography, X-Ray
  • Cyclin-Dependent Kinase 2 / antagonists & inhibitors
  • Cyclin-Dependent Kinases / antagonists & inhibitors*
  • Enzyme Inhibitors / chemical synthesis*
  • Enzyme Inhibitors / pharmacology*
  • Humans
  • Immunoblotting
  • Models, Molecular
  • Pyrazoles / chemical synthesis*
  • Pyrazoles / pharmacology*
  • RNA / biosynthesis
  • RNA / isolation & purification
  • Reverse Transcriptase Polymerase Chain Reaction
  • Reverse Transcription / drug effects
  • Structure-Activity Relationship
  • Substrate Specificity


  • Antimetabolites
  • Azo Compounds
  • Enzyme Inhibitors
  • Pyrazoles
  • RNA
  • Cyclin-Dependent Kinase 2
  • Cyclin-Dependent Kinases
  • Bromodeoxyuridine

Associated data

  • PDB/2CLX