title: Discovery and characterization of small molecules that target the GTPase Ral.
abstract: The Ras-like GTPases RalA and RalB are important drivers of tumour growth and metastasis. Chemicals that block Ral function would be valuable as research tools and for cancer therapeutics. Here we used protein structure analysis and virtual screening to identify drug-like molecules that bind to a site on the GDP-bound form of Ral. The compounds RBC6, RBC8 and RBC10 inhibited the binding of Ral to its effector RALBP1, as well as inhibiting Ral-mediated cell spreading of murine embryonic fibroblasts and anchorage-independent growth of human cancer cell lines. The binding of the RBC8 derivative BQU57 to RalB was confirmed by isothermal titration calorimetry, surface plasmon resonance and (1)H-(15)N transverse relaxation-optimized spectroscopy (TROSY) NMR spectroscopy. RBC8 and BQU57 show selectivity for Ral relative to the GTPases Ras and RhoA and inhibit tumour xenograft growth to a similar extent to the depletion of Ral using RNA interference. Our results show the utility of structure-based discovery for the development of therapeutics for Ral-dependent cancers.
authors.affiliation: Department of Surgery, University of Colorado, Aurora, Colorado 80045, USA.
authors.forename: Chao
authors.initials: C
authors.lastname: Yan
authors.affiliation: Department of Biochemistry, Indiana University School of Medicine, Indianapolis, Indiana 46202, USA.
authors.forename: Degang
authors.initials: D
authors.lastname: Liu
authors.affiliation: Department of Biochemistry, Indiana University School of Medicine, Indianapolis, Indiana 46202, USA.
authors.forename: Liwei
authors.initials: L
authors.lastname: Li
authors.affiliation: Department of Pharmaceutical Sciences, University of Colorado, Aurora, Colorado 80045, USA.
authors.forename: Michael F
authors.initials: MF
authors.lastname: Wempe
authors.affiliation: Department of Surgery, University of Colorado, Aurora, Colorado 80045, USA.
authors.forename: Sunny
authors.initials: S
authors.lastname: Guin
authors.affiliation: Department of Biochemistry, Indiana University School of Medicine, Indianapolis, Indiana 46202, USA.
authors.forename: May
authors.initials: M
authors.lastname: Khanna
authors.affiliation: Cardiovascular Research Center, University of Virginia, Charlottesville, Virginia 22908, USA.
authors.forename: Jeremy
authors.initials: J
authors.lastname: Meier
authors.affiliation: Cardiovascular Research Center, University of Virginia, Charlottesville, Virginia 22908, USA.
authors.forename: Brenton
authors.initials: B
authors.lastname: Hoffman
authors.affiliation: Department of Surgery, University of Colorado, Aurora, Colorado 80045, USA.
authors.forename: Charles
authors.initials: C
authors.lastname: Owens
authors.affiliation: Department of Pharmacology, University of Colorado, Aurora, Colorado 80045, USA.
authors.forename: Christina L
authors.initials: CL
authors.lastname: Wysoczynski
authors.affiliation: Department of Microbiology, Immunology, and Cancer Biology, University of Virginia, Charlottesville, Virginia 22908, USA.
authors.forename: Matthew D
authors.initials: MD
authors.lastname: Nitz
authors.affiliation: Department of Biochemistry, Indiana University School of Medicine, Indianapolis, Indiana 46202, USA.
authors.forename: William E
authors.initials: WE
authors.lastname: Knabe
authors.affiliation: 1] Department of Cardiology, Yale University, New Haven, Connecticut 06511, USA [2] Department of Cell Biology, Yale University, New Haven, Connecticut 06511, USA.
authors.forename: Mansoor
authors.initials: M
authors.lastname: Ahmed
authors.affiliation: Department of Microbiology, Immunology, and Cancer Biology, University of Virginia, Charlottesville, Virginia 22908, USA.
authors.forename: David L
authors.initials: DL
authors.lastname: Brautigan
authors.affiliation: Department of Biochemistry and Molecular Genetics, University of Virginia, Charlottesville, Virginia 22908, USA.
authors.forename: Bryce M
authors.initials: BM
authors.lastname: Paschal
authors.affiliation: 1] Department of Cardiology, Yale University, New Haven, Connecticut 06511, USA [2] Department of Cell Biology, Yale University, New Haven, Connecticut 06511, USA.
authors.forename: Martin A
authors.initials: MA
authors.lastname: Schwartz
authors.affiliation: Department of Pharmacology, University of Colorado, Aurora, Colorado 80045, USA.
authors.forename: David N M
authors.initials: DN
authors.lastname: Jones
authors.affiliation: Department of Pharmaceutical Sciences, University of Colorado, Aurora, Colorado 80045, USA.
authors.forename: David
authors.initials: D
authors.lastname: Ross
authors.affiliation: 1] Department of Biochemistry, Indiana University School of Medicine, Indianapolis, Indiana 46202, USA [2] Department of Chemistry and Chemical Biology, Indiana University - Purdue University, Indianapolis, Indiana 46202, USA.
authors.forename: Samy O
authors.initials: SO
authors.lastname: Meroueh
authors.affiliation: 1] Department of Surgery, University of Colorado, Aurora, Colorado 80045, USA [2] Department of Pharmacology, University of Colorado, Aurora, Colorado 80045, USA [3] University of Colorado Comprehensive Cancer Center, Aurora, Colorado 80045, USA.
authors.forename: Dan
authors.initials: D
authors.lastname: Theodorescu
mesh_terms: D018528:ATP-Binding Cassette Transporters / Q000378:metabolism; D000818:Animals; D045744:Cell Line, Tumor; D049109:Cell Proliferation / Q000187:drug effects; D003198:Computer Simulation; D004354:Drug Screening Assays, Antitumor; D005260:Female; D020690:GTPase-Activating Proteins / Q000378:metabolism; D006801:Humans; D051379:Mice; D008958:Models, Molecular; D058990:Molecular Targeted Therapy; D009369:Neoplasms / Q000188:drug therapy / Q000201:enzymology / Q000378:metabolism / Q000473:pathology; D011485:Protein Binding / Q000187:drug effects; D015398:Signal Transduction / Q000187:drug effects; D054852:Small Molecule Libraries / Q000737:chemistry* / Q000494:pharmacology*; D013379:Substrate Specificity; D023041:Xenograft Model Antitumor Assays; D020725:ral GTP-Binding Proteins / Q000037:antagonists & inhibitors* / Q000737:chemistry / Q000378:metabolism; D018631:ras Proteins / Q000378:metabolism
chemical_list: D018528:ATP-Binding Cassette Transporters; D020690:GTPase-Activating Proteins; C095637:RALBP1 protein, human; D054852:Small Molecule Libraries; D020725:ral GTP-Binding Proteins; D018631:ras Proteins