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Lookup NU author(s): Rachel Heap,
Professor Matthias TrostORCiD
This work is licensed under a Creative Commons Attribution 4.0 International License (CC BY 4.0).
© 2017, © 2017 Society for Laboratory Automation and Screening. 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.
Author(s): Heap RE, Hope AG, Pearson L-A, Reyskens KMSE, McElroy SP, Hastie CJ, Porter DW, Arthur JSC, Gray DW, Trost M
Publication type: Article
Publication status: Published
Journal: SLAS Discovery
Print publication date: 01/12/2017
Online publication date: 10/07/2017
Acceptance date: 06/06/2017
Date deposited: 18/12/2017
ISSN (print): 2472-5552
ISSN (electronic): 2472-5560
Publisher: SAGE Publications Inc.
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