Identifying Inhibitors of Inflammation: A Novel High-Throughput MALDI-TOF Screening Assay for Salt-Inducible Kinases (SIKs). (December 2017)

• Record Type: Journal Article
• Authors: Heap, Rachel E.; Hope, Anthony G.; Pearson, Lesley-Anne; Reyskens, Kathleen M. S. E.; McElroy, Stuart P.; Hastie, C. James; Porter, David W.; Arthur, J. Simon C.; Gray, David W.; Trost, Matthias
• Is Part Of: SLAS discovery. Volume 22:Number 10(2017)
• Page Start: 1193
• Page End: 1202
• Publication Date: 2017-12
• Subjects: MALDI TOF -- mass spectrometry -- salt inducible kinases -- kinase -- high-throughput screen -- inflammation -- drug discovery -- macrophage -- interleukin-10; Drugs -- Analysis -- Periodicals; Drugs -- Testing -- Periodicals; Biomolecules -- Analysis -- Periodicals; Biomolecules -- Analysis; Drugs -- Analysis; Drugs -- Testing; Drug Evaluation, Preclinical; Molecular Biology -- methods; Periodicals; Periodicals; 615.1
• DOI: 10.1177/2472555217717473
• Languages: English
• ISSNs: 2472-5552

Abstract

Matrix-assisted laser desorption/ionization time-of-flight (MALDI TOF) mass spectrometry has become a promising alternative for high-throughput drug discovery as new instruments offer high speed, flexibility and sensitivity, and the ability to measure physiological substrates label free. Here we developed and applied high-throughput MALDI TOF mass spectrometry to identify inhibitors of the salt-inducible kinase (SIK) family, which are interesting drug targets in the field of inflammatory disease as they control production of the anti-inflammatory cytokine interleukin-10 (IL-10) in macrophages. Using peptide substrates in in vitro kinase assays, we can show that hit identification of the MALDI TOF kinase assay correlates with indirect ADP-Hunter kinase assays. Moreover, we can show that both techniques generate comparable IC50 data for a number of hit compounds and known inhibitors of SIK kinases. We further take these inhibitors to a fluorescence-based cellular assay using the SIK activity-dependent translocation of CRTC3 into the nucleus, thereby providing a complete assay pipeline for the identification of SIK kinase inhibitors in vitro and in cells. Our data demonstrate that MALDI TOF mass spectrometry is fully applicable to high-throughput kinase screening, providing label-free data comparable to that of current high-throughput fluorescence assays.