DFG-out mode of inhibition by an irreversible type-1 inhibitor capable of overcoming gate-keeper mutations in FGF receptors

ACS Chem Biol. 2015 Jan 16;10(1):299-309. doi: 10.1021/cb500674s. Epub 2014 Oct 27.


Drug-resistance acquisition through kinase gate-keeper mutations is a major hurdle in the clinic. Here, we determined the first crystal structures of the human FGFR4 kinase domain (FGFR4K) alone and complexed with ponatinib, a promiscuous type-2 (DFG-out) kinase inhibitor, and an oncogenic FGFR4K harboring the V550L gate-keeper mutation bound to FIIN-2, a new type-1 irreversible inhibitor. Remarkably, like ponatinib, FIIN-2 also binds in the DFG-out mode despite lacking a functional group necessary to occupy the pocket vacated upon the DFG-out flip. Structural analysis reveals that the covalent bond between FIIN-2 and a cysteine, uniquely present in the glycine-rich loop of FGFR kinases, facilitates the DFG-out conformation, which together with the internal flexibility of FIIN-2 enables FIIN-2 to avoid the steric clash with the gate-keeper mutation that causes the ponatinib resistance. The structural data provide a blueprint for the development of next generation anticancer inhibitors through combining the salient inhibitory mechanisms of ponatinib and FIIN-2.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Amino Acid Motifs
  • Animals
  • Cell Line
  • Cell Survival / drug effects
  • Crystallography, X-Ray
  • Drug Resistance, Neoplasm* / drug effects
  • Drug Resistance, Neoplasm* / genetics
  • Humans
  • Mice
  • Models, Molecular
  • Mutation*
  • Oligopeptides / chemistry
  • Protein Kinase Inhibitors / chemistry
  • Protein Kinase Inhibitors / pharmacology*
  • Receptor, Fibroblast Growth Factor, Type 4 / antagonists & inhibitors*
  • Receptor, Fibroblast Growth Factor, Type 4 / chemistry
  • Receptor, Fibroblast Growth Factor, Type 4 / genetics*
  • Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
  • Tandem Mass Spectrometry


  • DFG peptide
  • Oligopeptides
  • Protein Kinase Inhibitors
  • FGFR4 protein, human
  • Receptor, Fibroblast Growth Factor, Type 4