BRAF inhibitors promote intermediate BRAF(V600E) conformations and binary interactions with activated RAS

Sci Adv. 2019 Aug 14;5(8):eaav8463. doi: 10.1126/sciadv.aav8463. eCollection 2019 Aug.


Oncogenic BRAF mutations initiate tumor formation by unleashing the autoinhibited kinase conformation and promoting RAS-decoupled proliferative RAF-MEK-ERK signaling. We have engineered luciferase-based biosensors to systematically track full-length BRAF conformations and interactions affected by tumorigenic kinase mutations and GTP loading of RAS. Binding of structurally diverse αC-helix-OUT BRAF inhibitors (BRAFi) showed differences in specificity and efficacy by shifting patient mutation-containing BRAF reporters from the definitive opened to more closed conformations. Unexpectedly, BRAFi engagement with the catalytic pocket of V600E-mutated BRAF stabilized an intermediate and inactive kinase conformation that enhanced binary RAS:RAF interactions, also independently of RAF dimerization in melanoma cells. We present evidence that the interference with RAS interactions and nanoclustering antagonizes the sequential formation of drug-induced RAS:RAF tetramers. This suggests a previously unappreciated allosteric effect of anticancer drug-driven intramolecular communication between the kinase and RAS-binding domains of mutated BRAF, which may further promote paradoxical kinase activation and drug resistance mechanisms.

Publication types

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

MeSH terms

  • Antineoplastic Agents / pharmacology
  • Biosensing Techniques
  • Catalytic Domain / genetics
  • Cell Line, Tumor
  • Cell Transformation, Neoplastic / genetics
  • HEK293 Cells
  • Humans
  • MAP Kinase Signaling System / drug effects
  • Melanoma / drug therapy*
  • Melanoma / genetics
  • Melanoma / pathology*
  • Molecular Conformation
  • Protein Kinase Inhibitors / pharmacology*
  • Proto-Oncogene Proteins B-raf / antagonists & inhibitors*
  • Proto-Oncogene Proteins B-raf / genetics
  • Proto-Oncogene Proteins p21(ras) / metabolism*
  • Signal Transduction


  • Antineoplastic Agents
  • Protein Kinase Inhibitors
  • BRAF protein, human
  • Proto-Oncogene Proteins B-raf
  • Proto-Oncogene Proteins p21(ras)