title: RAF inhibitors activate the MAPK pathway by relieving inhibitory autophosphorylation.
abstract: ATP competitive inhibitors of the BRAF(V600E) oncogene paradoxically activate downstream signaling in cells bearing wild-type BRAF (BRAF(WT)). In this study, we investigate the biochemical mechanism of wild-type RAF (RAF(WT)) activation by multiple catalytic inhibitors using kinetic analysis of purified BRAF(V600E) and RAF(WT) enzymes. We show that activation of RAF(WT) is ATP dependent and directly linked to RAF kinase activity. These data support a mechanism involving inhibitory autophosphorylation of RAF's phosphate-binding loop that, when disrupted either through pharmacologic or genetic alterations, results in activation of RAF and the mitogen-activated protein kinase (MAPK) pathway. This mechanism accounts not only for compound-mediated activation of the MAPK pathway in BRAF(WT) cells but also offers a biochemical mechanism for BRAF oncogenesis.
authors.affiliation: Novartis Institutes for Biomedical Research, Emeryville, CA 94608, USA.
authors.forename: Matthew
authors.initials: M
authors.lastname: Holderfield
authors.forename: Hanne
authors.initials: H
authors.lastname: Merritt
authors.forename: John
authors.initials: J
authors.lastname: Chan
authors.forename: Marco
authors.initials: M
authors.lastname: Wallroth
authors.forename: Laura
authors.initials: L
authors.lastname: Tandeske
authors.forename: Huili
authors.initials: H
authors.lastname: Zhai
authors.forename: John
authors.initials: J
authors.lastname: Tellew
authors.forename: Stephen
authors.initials: S
authors.lastname: Hardy
authors.forename: Mohammad
authors.initials: M
authors.lastname: Hekmat-Nejad
authors.forename: Darrin D
authors.initials: DD
authors.lastname: Stuart
authors.forename: Frank
authors.initials: F
authors.lastname: McCormick
authors.forename: Tobi E
authors.initials: TE
authors.lastname: Nagel
mesh_terms: D000255:Adenosine Triphosphate / Q000378:metabolism / Q000502:physiology; D045744:Cell Line, Tumor; D006801:Humans; D020935:MAP Kinase Signaling System / Q000187:drug effects*; D010766:Phosphorylation / Q000187:drug effects; D048493:Proto-Oncogene Proteins B-raf / Q000235:genetics / Q000378:metabolism / Q000502:physiology; D019908:Proto-Oncogene Proteins c-raf / Q000235:genetics / Q000378:metabolism / Q000502:physiology; D048490:raf Kinases / Q000037:antagonists & inhibitors* / Q000235:genetics / Q000378:metabolism
chemical_list: D000255:Adenosine Triphosphate; C482119:BRAF protein, human; D048493:Proto-Oncogene Proteins B-raf; D019908:Proto-Oncogene Proteins c-raf; D048490:raf Kinases