title: Identification of autophosphorylation inhibitors of the inositol-requiring enzyme 1 alpha (IRE1α) by high-throughput screening using a DELFIA assay.
abstract: Inositol-requiring enzyme 1 alpha (IRE1α) is a transmembrane sensor protein with both kinase and ribonuclease activity, which plays a crucial role in the unfolded protein response (UPR). Protein misfolding in the endoplasmic reticulum (ER) lumen triggers dimerization and subsequent trans-autophosphorylation of IRE1α. This leads to the activation of its endoribonuclease (RNase) domain and splicing of the mRNA of the transcriptional activator XBP1, ultimately generating an active XBP1 (XBP1s) implicated in multiple myeloma survival. Previously, we have identified human IRE1α as a target for the development of kinase inhibitors that could modulate the UPR in human cells, which has particular relevance for multiple myeloma and other secretory malignancies. Here we describe the development and validation of a 384-well high-throughput screening assay using DELFIA technology that is specific for IRE1α autophosphorylation. Using this format, a focused library of 2312 potential kinase inhibitors was screened, and several novel IRE1α kinase inhibitor scaffolds were identified that could potentially be developed toward new therapies to treat multiple myeloma.
authors.affiliation: Division of Cancer Therapeutics, The Institute of Cancer Research, Sutton, Surrey, UK.
authors.forename: Yvette
authors.initials: Y
authors.lastname: Newbatt
authors.forename: Anthea
authors.initials: A
authors.lastname: Hardcastle
authors.forename: P Craig
authors.initials: PC
authors.lastname: McAndrew
authors.forename: Jade A
authors.initials: JA
authors.lastname: Strover
authors.forename: Amin
authors.initials: A
authors.lastname: Mirza
authors.forename: Gareth J
authors.initials: GJ
authors.lastname: Morgan
authors.forename: Rosemary
authors.initials: R
authors.lastname: Burke
authors.forename: Faith E
authors.initials: FE
authors.lastname: Davies
authors.forename: Ian
authors.initials: I
authors.lastname: Collins
authors.forename: Rob L M
authors.initials: RL
authors.lastname: van Montfort
mesh_terms: D000818:Animals; D002460:Cell Line; D004722:Endoribonucleases / Q000037:antagonists & inhibitors* / Q000737:chemistry / Q000378:metabolism; D057166:High-Throughput Screening Assays / Q000379:methods; D006801:Humans; D007313:Insecta; D008565:Membrane Proteins / Q000037:antagonists & inhibitors / Q000378:metabolism; D009101:Multiple Myeloma / Q000188:drug therapy / Q000201:enzymology / Q000378:metabolism; D010766:Phosphorylation / Q000187:drug effects; D017510:Protein Folding / Q000187:drug effects; D047428:Protein Kinase Inhibitors / Q000737:chemistry* / Q000494:pharmacology*; D017346:Protein Serine-Threonine Kinases / Q000037:antagonists & inhibitors* / Q000737:chemistry / Q000378:metabolism; D056811:Unfolded Protein Response / Q000187:drug effects
chemical_list: D008565:Membrane Proteins; D047428:Protein Kinase Inhibitors; C528229:ERN1 protein, human; D017346:Protein Serine-Threonine Kinases; D004722:Endoribonucleases