title: Characterization of Hedgehog Acyltransferase Inhibitors Identifies a Small Molecule Probe for Hedgehog Signaling by Cancer Cells.
abstract: The Sonic Hedgehog (Shh) signaling pathway plays a critical role during embryonic development and cancer progression. N-terminal palmitoylation of Shh by Hedgehog acyltransferase (Hhat) is essential for efficient signaling, raising interest in Hhat as a novel drug target. A recently identified series of dihydrothienopyridines has been proposed to function via this mode of action; however, the lead compound in this series (RUSKI-43) was subsequently shown to possess cytotoxic activity unrelated to canonical Shh signaling. To identify a selective chemical probe for cellular studies, we profiled three RUSKI compounds in orthogonal cell-based assays. We found that RUSKI-43 exhibits off-target cytotoxicity, masking its effect on Hhat-dependent signaling, hence results obtained with this compound in cells should be treated with caution. In contrast, RUSKI-201 showed no off-target cytotoxicity, and quantitative whole-proteome palmitoylation profiling with a bioorthogonal alkyne-palmitate reporter demonstrated specific inhibition of Hhat in cells. RUSKI-201 is the first selective Hhat chemical probe in cells and should be used in future studies of Hhat catalytic function.
authors: Molecular Medicine Section, National Heart & Lung Institute, Imperial College London , London SW7 2AZ, United Kingdom.
authors: Ursula R
authors: UR
authors: Rodgers
authors: Department of Chemistry, Imperial College London , London SW7 2AZ, United Kingdom.
authors: Thomas
authors: T
authors: Lanyon-Hogg
authors: Department of Chemistry, Imperial College London , London SW7 2AZ, United Kingdom.
authors: Naoko
authors: N
authors: Masumoto
authors: Department of Chemistry, Imperial College London , London SW7 2AZ, United Kingdom.
authors: Markus
authors: M
authors: Ritzefeld
authors: Cancer Research UK Cancer Therapeutics Unit, The Institute of Cancer Research , London, SW7 3RP, United Kingdom.
authors: Rosemary
authors: R
authors: Burke
authors: Cancer Research UK Cancer Therapeutics Unit, The Institute of Cancer Research , London, SW7 3RP, United Kingdom.
authors: Julian
authors: J
authors: Blagg
authors: Molecular Medicine Section, National Heart & Lung Institute, Imperial College London , London SW7 2AZ, United Kingdom.
authors: Anthony I
authors: AI
authors: Magee
authors: Department of Chemistry, Imperial College London , London SW7 2AZ, United Kingdom.
authors: Edward W
authors: EW
authors: Tate
mesh_terms: D000217:Acyltransferases / Q000037:antagonists & inhibitors* / Q000378:metabolism; D000818:Animals; D045744:Cell Line, Tumor; D004791:Enzyme Inhibitors / Q000737:chemistry* / Q000494:pharmacology*; D057809:HEK293 Cells; D053823:Hedgehog Proteins / Q000378:metabolism*; D006801:Humans; D054878:Lipoylation / Q000187:drug effects; D051379:Mice; D041681:NIH 3T3 Cells; D009369:Neoplasms / Q000188:drug therapy* / Q000378:metabolism; D015398:Signal Transduction / Q000187:drug effects*
chemical_list: D004791:Enzyme Inhibitors; D053823:Hedgehog Proteins; D000217:Acyltransferases; C105599:HHAT protein, human