Journal
SLAS DISCOVERY
Volume 22, Issue 10, Pages 1193-1202Publisher
SAGE PUBLICATIONS INC
DOI: 10.1177/2472555217717473
Keywords
MALDI TOF; mass spectrometry; salt inducible kinases; kinase; high-throughput screen; inflammation; drug discovery; macrophage; interleukin-10
Funding
- Medical Research Council UK [MC_UU_12016/5, MC_PC_14099, MR/L008785/1]
- BBSRC
- Bruker Daltonics
- Arthritis Research UK [20409]
- Boehringer-Ingelheim
- GlaxoSmithKline
- Merck KGaA
- MRC [MC_UU_12016/5, MC_PC_14099, MC_UP_A500_1020, MC_PC_15036] Funding Source: UKRI
- Medical Research Council [MC_UP_A500_1020, MC_UU_12016/5, MC_PC_15036, MC_PC_14099] Funding Source: researchfish
- Versus Arthritis
- Cancer Research UK [20409] Funding Source: researchfish
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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.
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