A road map to evaluate the proteome-wide selectivity of covalent kinase inhibitors

Nat Chem Biol. 2014 Sep;10(9):760-767. doi: 10.1038/nchembio.1582. Epub 2014 Jul 13.


Kinases are principal components of signal transduction pathways and the focus of intense basic and drug discovery research. Irreversible inhibitors that covalently modify non-catalytic cysteines in kinase active sites have emerged as valuable probes and approved drugs. Many protein classes, however, have functional cysteines, and therefore understanding the proteome-wide selectivity of covalent kinase inhibitors is imperative. Here, we accomplish this objective using activity-based protein profiling coupled with quantitative MS to globally map the targets, both specific and nonspecific, of covalent kinase inhibitors in human cells. Many of the specific off-targets represent nonkinase proteins that, notably, have conserved active site cysteines. We define windows of selectivity for covalent kinase inhibitors and show that, when these windows are exceeded, rampant proteome-wide reactivity and kinase target-independent cell death conjointly occur. Our findings, taken together, provide an experimental road map to illuminate opportunities and surmount challenges for the development of covalent kinase inhibitors.

Publication types

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

MeSH terms

  • Agammaglobulinaemia Tyrosine Kinase
  • Cell Line, Tumor
  • Cell Survival / drug effects
  • Cysteine / chemistry
  • Genes, erbB-1 / genetics
  • Humans
  • Kinetics
  • Protein Kinase Inhibitors / pharmacology*
  • Protein Kinases / metabolism
  • Protein-Tyrosine Kinases / antagonists & inhibitors
  • Proteome / genetics*
  • Pyrazoles / pharmacology
  • Pyrimidines / pharmacology
  • Signal Transduction / drug effects
  • Signal Transduction / genetics


  • Protein Kinase Inhibitors
  • Proteome
  • Pyrazoles
  • Pyrimidines
  • ibrutinib
  • Protein Kinases
  • Protein-Tyrosine Kinases
  • Agammaglobulinaemia Tyrosine Kinase
  • Cysteine