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Chemically Linked Vemurafenib Inhibitors Promote an Inactive BRAFV600E Conformation

Journal Article · · ACS Chemical Biology
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  1. Univ. of Pennsylvania, Philadelphia, PA (United States)
  2. Wistar Inst., Philadelphia, PA (United States)
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The BRAF kinase, within the mitogen activated protein kinase (MAPK) signaling pathway, harbors activating mutations in about half of melanomas and to a significant extent in many other cancers. A single valine to glutamic acid substitution at residue 600 (BRAFV600E) accounts for about 90% of these activating mutations. While BRAFV600E-selective small molecule inhibitors, such as debrafenib and vemurafenib, have shown therapeutic benefit, almost all patients develop resistance. Resistance often arises through reactivation of the MAPK pathway, typically through mutation of upstream RAS, downstream MEK, or splicing variants. RAF kinases signal as homo- and heterodimers, and another complication associated with small molecule BRAFV600E inhibition is drug-induced allosteric activation of a wild-type RAF subunit (BRAF or CRAF) of the kinase dimer, a process called “transactivation” or “paradoxical activation.” Here, we used BRAFV600E and vemurafenib as a model system to develop chemically linked kinase inhibitors to lock RAF dimers in an inactive conformation that cannot undergo transactivation. This structure-based design effort resulted in the development of Vem-BisAmide-2, a compound containing two vemurafenib molecules connected by a bis amide linker. We show that Vem-BisAmide-2 has comparable inhibitory potency as vemurafenib to BRAFV600E both in vitro and in cells but promotes an inactive dimeric BRAFV600E conformation unable to undergo transactivation. The crystal structure of a BRAFV600E/Vem-BisAmide-2 complex and associated biochemical studies reveal the molecular basis for how Vem-BisAmide-2 mediates selectivity for an inactive over an active dimeric BRAFV600E conformation. These studies have implications for targeting BRAFV600E/RAF heterodimers and other kinase dimers for therapy.
Research Organization:
Argonne National Laboratory (ANL), Argonne, IL (US)
Sponsoring Organization:
National Institutes of Health (NIH); USDOE Office of Science (SC)
OSTI ID:
1331701
Journal Information:
ACS Chemical Biology, Journal Name: ACS Chemical Biology Journal Issue: 10 Vol. 11; ISSN 1554-8929
Publisher:
American Chemical Society (ACS)Copyright Statement
Country of Publication:
United States
Language:
ENGLISH

References (42)

Mechanism of Activation of the RAF-ERK Signaling Pathway by Oncogenic Mutations of B-RAF journal March 2004
Experimental and computational approaches to estimate solubility and permeability in drug discovery and development settings 1PII of original article: S0169-409X(96)00423-1. The article was originally published in Advanced Drug Delivery Reviews 23 (1997) 3–25. 1 journal March 2001
Acquired Resistance to BRAF Inhibitors Mediated by a RAF Kinase Switch in Melanoma Can Be Overcome by Cotargeting MEK and IGF-1R/PI3K journal December 2010
Mutant BRAF Melanomas—Dependence and Resistance journal January 2011
Allosteric Activation of Functionally Asymmetric RAF Kinase Dimers journal August 2013
Concurrent MEK2 Mutation and BRAF Amplification Confer Resistance to BRAF and MEK Inhibitors in Melanoma journal September 2013
Effects of Raf Dimerization and Its Inhibition on Normal and Disease-Associated Raf Signaling journal February 2013
Identification of a Novel Family of BRAF V600E Inhibitors journal May 2012
Discovery of Dabrafenib: A Selective Inhibitor of Raf Kinases with Antitumor Activity against B-Raf-Driven Tumors journal January 2013
Distinct requirement for an intact dimer interface in wild-type, V600E and kinase-dead B-Raf signalling: V600E and G469A B-Raf are potent homodimerisers journal April 2012
Mutations of the BRAF gene in human cancer journal June 2002
A dimerization-dependent mechanism drives RAF catalytic activation journal September 2009
RAF inhibitors prime wild-type RAF to activate the MAPK pathway and enhance growth journal February 2010
RAF inhibitors transactivate RAF dimers and ERK signalling in cells with wild-type BRAF journal February 2010
Melanomas acquire resistance to B-RAF(V600E) inhibition by RTK or N-RAS upregulation journal November 2010
RAF inhibitor resistance is mediated by dimerization of aberrantly spliced BRAF(V600E) journal November 2011
RAF inhibitors that evade paradoxical MAPK pathway activation journal October 2015
Inhibitors that stabilize a closed RAF kinase domain conformation induce dimerization journal May 2013
Vemurafenib: the first drug approved for BRAF-mutant cancer journal October 2012
The RAF proteins take centre stage journal November 2004
Crystal structure of a BRAF kinase domain monomer explains basis for allosteric regulation journal December 2014
The RTK/RAS/BRAF/PI3K Pathways in Melanoma: Biology, Small Molecule Inhibitors, and Potential Applications journal December 2007
Inhibition of Mutated, Activated BRAF in Metastatic Melanoma journal August 2010
Survival in BRAF V600–Mutant Advanced Melanoma Treated with Vemurafenib journal February 2012
Combined BRAF and MEK Inhibition in Melanoma with BRAF V600 Mutations journal November 2012
Discovery of a selective inhibitor of oncogenic B-Raf kinase with potent antimelanoma activity journal February 2008
Paradoxical activation and RAF inhibitor resistance of BRAF protein kinase fusions characterizing pediatric astrocytomas journal March 2013
Phaser crystallographic software journal July 2007
electronic Ligand Builder and Optimization Workbench ( eLBOW ): a tool for ligand coordinate and restraint generation journal September 2009
MolProbity : all-atom structure validation for macromolecular crystallography journal December 2009
XDS journal January 2010
PHENIX: a comprehensive Python-based system for macromolecular structure solution journal January 2010
Features and development of Coot journal March 2010
Towards automated crystallographic structure refinement with phenix.refine journal March 2012
Cutaneous manifestations of dabrafenib (GSK2118436): a selective inhibitor of mutant BRAF in patients with metastatic melanoma: Cutaneous manifestations of dabrafenib (GSK2118436) journal October 2012
A small-molecule inhibitor of the aberrant transcription factor CBF -SMMHC delays leukemia in mice journal February 2015
Improved antitumor activity of immunotherapy with BRAF and MEK inhibitors in BRAF V600E melanoma journal March 2015
Regulation and Role of Raf-1/B-Raf Heterodimerization journal March 2006
Resistance to BRAF Inhibitors: Unraveling Mechanisms and Future Treatment Options: Figure 1. journal November 2011
Combinations of BRAF, MEK, and PI3K/mTOR Inhibitors Overcome Acquired Resistance to the BRAF Inhibitor GSK2118436 Dabrafenib, Mediated by NRAS or MEK Mutations journal March 2012
Combined BRAF (Dabrafenib) and MEK Inhibition (Trametinib) in Patients With BRAF V600 -Mutant Melanoma Experiencing Progression With Single-Agent BRAF Inhibitor journal November 2014
Dabrafenib; Preclinical Characterization, Increased Efficacy when Combined with Trametinib, while BRAF/MEK Tool Combination Reduced Skin Lesions journal July 2013

Cited By (4)

Mechanism of BRAF Activation through Biochemical Characterization of the Recombinant Full-Length Protein journal August 2018
Biochemical Characterization of Full‐Length Oncogenic BRAF V600E together with Molecular Dynamics Simulations Provide Insight into the Activation and Inhibition Mechanisms of RAF Kinases journal July 2019
A patent review of RAF kinase inhibitors (2010–2018) journal August 2019
Targeting the ERK Signaling Pathway in Melanoma journal March 2019

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