MALDI-TOF Mass Spectrometry-Based High-Throughput Screening for Inhibitors of the Cytosolic DNA Sensor cGAS
| Autor: | Jun Li, Frank H. Büttner, Martin Winter, Annekatrin Heimann, Daniel Bischoff, Roman P. Simon, Andreas H. Luippold, Wolfgang Reindl, Robert Ries, Tom Bretschneider, Gisela Schnapp, Ljiljana Zuvela-Jelaska, Carola Kleiner |
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| Rok vydání: | 2020 |
| Předmět: |
0301 basic medicine
Dna sensor High-throughput screening Context (language use) Computational biology Mass spectrometry 01 natural sciences Biochemistry Analytical Chemistry Small Molecule Libraries 03 medical and health sciences Cytosol Drug Discovery Humans Enzyme Inhibitors Physiological reaction Chemistry Drug discovery DNA MALDI-TOF Mass Spectrometry Nucleotidyltransferases Small molecule High-Throughput Screening Assays 0104 chemical sciences 010404 medicinal & biomolecular chemistry 030104 developmental biology Spectrometry Mass Matrix-Assisted Laser Desorption-Ionization Molecular Medicine Biotechnology |
| Zdroj: | SLAS Discovery. 25:372-383 |
| ISSN: | 2472-5552 |
| DOI: | 10.1177/2472555219880185 |
| Popis: | Comprehensive and unbiased detection methods are a prerequisite for high-throughput screening (HTS) campaigns within drug discovery research. Label-free matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF) mass spectrometry (MS) has been introduced as an HTS-compatible readout for biochemical test systems to support the drug discovery process. So far, reported HTS applications were based on surface-modified systems or proof-of-concept studies. We present the utilization of a MALDI-TOF-based screening platform to identify inhibitors of human cyclic GMP-AMP synthase (cGAS), a mediator of innate immune response whose aberration has been causally correlated to a number of inflammatory disorders. In this context, the development and validation of a MALDI-TOF-based activity assay is reported to demonstrate fast, robust, and accurate detection of chemical cGAS inhibition by direct quantification of the physiological reaction product cyclic GMP-ATP (cGAMP). Results from a screen of a diverse library of more than 1 million small molecules in 1536-well format against the catalytic cGAS activity are presented with excellent assay performance and data quality. Identified hits were qualified in dose-response experiments and confirmed by RapidFire-MS measurements. Conclusively, the presented data provide the first proof of applicability of direct automated MALDI-TOF MS as a readout strategy for large-scale drug discovery HTS campaigns. |
| Databáze: | OpenAIRE |
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