Discovery of a Potent BTK Inhibitor with a Novel Binding Mode by Using Parallel Selections with a DNA-Encoded Chemical Library

Chembiochem. 2017 May 4;18(9):864-871. doi: 10.1002/cbic.201600573. Epub 2017 Feb 8.


We have identified and characterized novel potent inhibitors of Bruton's tyrosine kinase (BTK) from a single DNA-encoded library of over 110 million compounds by using multiple parallel selection conditions, including variation in target concentration and addition of known binders to provide competition information. Distinct binding profiles were observed by comparing enrichments of library building block combinations under these conditions; one enriched only at high concentrations of BTK and was competitive with ATP, and another enriched at both high and low concentrations of BTK and was not competitive with ATP. A compound representing the latter profile showed low nanomolar potency in biochemical and cellular BTK assays. Results from kinetic mechanism of action studies were consistent with the selection profiles. Analysis of the co-crystal structure of the most potent compound demonstrated a novel binding mode that revealed a new pocket in BTK. Our results demonstrate that profile-based selection strategies using DNA-encoded libraries form the basis of a new methodology to rapidly identify small molecule inhibitors with novel binding modes to clinically relevant targets.

Keywords: Bruton's tyrosine kinase; DNA-encoded library; X-ray crystallography; kinase inhibitors; small molecules.

MeSH terms

  • Agammaglobulinaemia Tyrosine Kinase
  • Binding Sites
  • Cell Line
  • Cytochrome P-450 CYP3A / chemistry
  • Cytochrome P-450 CYP3A / metabolism
  • DNA / chemistry*
  • DNA / metabolism
  • Humans
  • Kinetics
  • Molecular Docking Simulation
  • Protein Binding
  • Protein Kinase Inhibitors / chemistry*
  • Protein Kinase Inhibitors / metabolism
  • Protein Structure, Tertiary
  • Protein-Tyrosine Kinases / antagonists & inhibitors*
  • Protein-Tyrosine Kinases / genetics
  • Protein-Tyrosine Kinases / metabolism
  • Small Molecule Libraries / chemistry*
  • Small Molecule Libraries / metabolism


  • Protein Kinase Inhibitors
  • Small Molecule Libraries
  • DNA
  • Cytochrome P-450 CYP3A
  • Protein-Tyrosine Kinases
  • Agammaglobulinaemia Tyrosine Kinase
  • BTK protein, human