title: Identification of PLX4032-resistance mechanisms and implications for novel RAF inhibitors.
abstract: BRAF inhibitors improve melanoma patient survival, but resistance invariably develops. Here we report the discovery of a novel BRAF mutation that confers resistance to PLX4032 employing whole-exome sequencing of drug-resistant BRAF(V600K) melanoma cells. We further describe a new screening approach, a genome-wide piggyBac mutagenesis screen that revealed clinically relevant aberrations (N-terminal BRAF truncations and CRAF overexpression). The novel BRAF mutation, a Leu505 to His substitution (BRAF(L505H) ), is the first resistance-conferring second-site mutation identified in BRAF mutant cells. The mutation replaces a small nonpolar amino acid at the BRAF-PLX4032 interface with a larger polar residue. Moreover, we show that BRAF(L505H) , found in human prostate cancer, is itself a MAPK-activating, PLX4032-resistant oncogenic mutation. Lastly, we demonstrate that the PLX4032-resistant melanoma cells are sensitive to novel, next-generation BRAF inhibitors, especially the 'paradox-blocker' PLX8394, supporting its use in clinical trials for treatment of melanoma patients with BRAF-mutations.
authors.affiliation: Department of Dermatology, Yale University School of Medicine, New Haven, CT, USA.
authors.forename: Jaehyuk
authors.initials: J
authors.lastname: Choi
authors.forename: Sean F
authors.initials: SF
authors.lastname: Landrette
authors.forename: Tiffany
authors.initials: T
authors.lastname: Wang
authors.forename: Perry
authors.initials: P
authors.lastname: Evans
authors.forename: Antonella
authors.initials: A
authors.lastname: Bacchiocchi
authors.forename: Robert
authors.initials: R
authors.lastname: Bjornson
authors.forename: Elaine
authors.initials: E
authors.lastname: Cheng
authors.forename: Amy L
authors.initials: AL
authors.lastname: Stiegler
authors.forename: Symon
authors.initials: S
authors.lastname: Gathiaka
authors.forename: Orlando
authors.initials: O
authors.lastname: Acevedo
authors.forename: Titus J
authors.initials: TJ
authors.lastname: Boggon
authors.forename: Michael
authors.initials: M
authors.lastname: Krauthammer
authors.forename: Ruth
authors.initials: R
authors.lastname: Halaban
authors.forename: Tian
authors.initials: T
authors.lastname: Xu
mesh_terms: D000595:Amino Acid Sequence; D045744:Cell Line, Tumor; D049109:Cell Proliferation / Q000187:drug effects; D004251:DNA Transposable Elements / Q000235:genetics; D019008:Drug Resistance, Neoplasm / Q000187:drug effects*; D006801:Humans; D007211:Indoles / Q000494:pharmacology*; D020935:MAP Kinase Signaling System / Q000187:drug effects; D008545:Melanoma / Q000201:enzymology / Q000473:pathology; D008958:Models, Molecular; D008969:Molecular Sequence Data; D016254:Mutagenesis, Insertional / Q000235:genetics; D050505:Mutant Proteins / Q000378:metabolism; D009154:Mutation / Q000235:genetics; D047428:Protein Kinase Inhibitors / Q000494:pharmacology*; D048493:Proto-Oncogene Proteins B-raf / Q000037:antagonists & inhibitors* / Q000378:metabolism; D013449:Sulfonamides / Q000494:pharmacology*; D000077484:Vemurafenib
chemical_list: D004251:DNA Transposable Elements; D007211:Indoles; D050505:Mutant Proteins; D047428:Protein Kinase Inhibitors; D013449:Sulfonamides; D000077484:Vemurafenib; D048493:Proto-Oncogene Proteins B-raf
keywords: BRAF; PLX4032; drug resistance; melanoma; paradox blockers