The Evolution of MALDI-TOF Mass Spectrometry toward Ultra-High-Throughput Screening: 1536-Well Format and Beyond

Haslam, Carl; Hellicar, John; Dunn, Adrian J.; Fuetterer, Arne; Hardy, Neil; Marshall, Peter S.; Paape, Rainer; Pemberton, Michelle; Resemannand, Anja; Leveridge, Melanie

doi:10.1177/1087057115608605

Cited by 121 publications

(121 citation statements)

References 27 publications

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“…It is a label-free approach as it relies upon the separation and subsequent quantification of typically a substrate and product that has undergone modification that the mass spectrometer is capable of detecting [100]. The current instrument for high throughput mass spectrometry is the Agilent RapidFire that has been used to screen a range of targets with improved quality of the identified hit compounds [101–103]. …”

Section: General Concepts Underlying the Common Deployed Screening Comentioning

confidence: 99%

Epigenetic assays for chemical biology and drug discovery

Gul

2017

Clin Epigenet

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The implication of epigenetic abnormalities in many diseases and the approval of a number of compounds that modulate specific epigenetic targets in a therapeutically relevant manner in cancer specifically confirms that some of these targets are druggable by small molecules. Furthermore, a number of compounds are currently in clinical trials for other diseases including cardiovascular, neurological and metabolic disorders. Despite these advances, the approved treatments for cancer only extend progression-free survival for a relatively short time and being associated with significant side effects. The current clinical trials involving the next generation of epigenetic drugs may address the disadvantages of the currently approved epigenetic drugs.The identification of chemical starting points of many drugs often makes use of screening in vitro assays against libraries of synthetic or natural products. These assays can be biochemical (using purified protein) or cell-based (using for example, genetically modified, cancer cell lines or primary cells) and performed in microtiter plates, thus enabling a large number of samples to be tested. A considerable number of such assays are available to monitor epigenetic target activity, and this review provides an overview of drug discovery and chemical biology and describes assays that monitor activities of histone deacetylase, lysine-specific demethylase, histone methyltransferase, histone acetyltransferase and bromodomain. It is of critical importance that an appropriate assay is developed and comprehensively validated for a given drug target prior to screening in order to improve the probability of the compound progressing in the drug discovery value chain.

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Section: General Concepts Underlying the Common Deployed Screening Comentioning

confidence: 99%

Epigenetic assays for chemical biology and drug discovery

Gul

2017

Clin Epigenet

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“…6t o8s/sample, MALDI-MS at ar ate of ca. 0.3 s/sample [8] and, acoustic droplet ejection MS at rates of 0.5 to 1s/sample. [9] Fort he above techniques,t he sacrifice in separation increases the HTS rate but can lead to loss of specificity and sensitivity in bioassays;m ethods enabling both high-throughput and efficient separation and analysis remain in high demand.…”

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confidence: 99%

High‐Throughput Bioassays using “Dip‐and‐Go” Multiplexed Electrospray Mass Spectrometry

et al. 2019

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Am ultiplexed system based on inductive nanoelectrospray mass spectrometry (nESI-MS) has been developed for high-throughput screening (HTS) bioassays.T his system combines inductive nESI and field amplification microelectrophoresis to achieve a"dip-and-go" sample loading and purification strategy that enables nESI-MS based HTS assays in 96-well microtiter plates.The combination of inductive nESI and micro-electrophoresis makes it possible to perform efficient in situ separations and clean-up of biological samples. The sensitivity of the system is such that quantitative analysis of peptides from 1-10 000 nm can be performed in ab iological matrix. Ap rototype of the automation system has been developed to handle 12 samples (one row of am icrotiter plate) at atime.The sample loading and electrophoretic cleanup of biosamples can be done in parallel within 20 sf ollowed by MS analysis at arate of 1.3 to 3.5 sper sample.The system was used successfully for the quantitative analysis of BACE1catalyzedp eptide hydrolysis,aprototypical HTS assayo f relevance to drug discovery.IC 50 values for this system were in agreement with LC-MS but recorded in times more than an order of magnitude shorter.

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“…To address these issues, we have developed a sensitive and fast assay to quantify in vitro E2/E3 enzyme activity using MALDI TOF MS. It builds on our DUB MALDI TOF assay 32 , which has enabled us to screen successfully for a number of selective DUB 295 inhibitors 56, 57, 58 , and adds to the increasing number of drug discovery assays utilising labelfree high-throughput MALDI TOF MS. Apart from E2/E3 enzymes and DUBs 32 , highthroughput MALDI TOF MS has now successfully been used for drug discovery screening of protein kinases 59 , histone demethylases and acetylcholinesterases 60 as well as histone lysine methyltransferases 61 . 300 Unlike other current assays, all these label-free MALDI TOF MS methods use unmodified substrates, such as mono-ubiquitin.…”

Section: Discussionmentioning

confidence: 99%

The MALDI TOF E2/E3 ligase assay as an universal tool for drug discovery in the ubiquitin pathway

Cesare

Johnson

Barlow

et al. 2017

Preprint

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In many diseases, components of the ubiquitin system -such as E2/E3 ligases and deubiquitylases -are dysregulated. The ubiquitin system has therefore become an emergent 30 target for the treatment of a number of diseases, including cancer, neurodegeneration and autoimmunity. Despite of the efforts in this field, primary screenings of compound libraries to individuate new potential therapeutic molecules targeting the ubiquitin pathway have been strongly limited by the lack of robust and fast high-throughput assays. Here we report the first label-free high-throughput screening (HTS) assay for ubiquitin E2 conjugating enzymes and 35 E3 ligases based on Matrix-Assisted Laser Desorption/Ionization Time-Of-Flight (MALDI TOF) mass spectrometry. The MALDI TOF E2/E3 assay allows us to test E2 conjugating enzymes and E3 ligases for their ubiquitin transfer activity, to identify E2/E3 active pairs, inhibitor potency and specificity and to screen compound libraries in vitro without synthesis of chemical or fluorescent probes. We demonstrate that the MALDI TOF E2/E3 assay is a universal tool 40 for drug discovery screening in the ubiquitin pathway as it is suitable for working with all E3 ligase families and requires a reduced amount of reagents, compared to standard biochemical assays.

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The Evolution of MALDI-TOF Mass Spectrometry toward Ultra-High-Throughput Screening: 1536-Well Format and Beyond

Cited by 121 publications

References 27 publications

Epigenetic assays for chemical biology and drug discovery

Epigenetic assays for chemical biology and drug discovery

High‐Throughput Bioassays using “Dip‐and‐Go” Multiplexed Electrospray Mass Spectrometry

The MALDI TOF E2/E3 ligase assay as an universal tool for drug discovery in the ubiquitin pathway

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