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SLAS Discov. 2017 Dec;22(10):1193-1202. doi: 10.1177/2472555217717473. Epub 2017 Jul 10.

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

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1 MRC Protein Phosphorylation & Ubiquitylation Unit, University of Dundee, Dundee, Scotland, UK.
2 Institute for Cell and Molecular Biosciences, Newcastle University, Newcastle-upon-Tyne, UK.
3 Drug Discovery Unit, University of Dundee, Dundee, Scotland, UK.
4 Division of Cell Signalling and Immunology, University of Dundee, Dundee, Scotland, UK.


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.


MALDI TOF; drug discovery; high-throughput screen; inflammation; interleukin-10; kinase; macrophage; mass spectrometry; salt inducible kinases

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