Mass spectrometry-based Cellular Thermal Shift Assay (CETSA®) for target deconvolution in phenotypic drug discovery

Friman, Tomas

doi:10.1016/j.bmc.2019.115174

Cited by 52 publications

(43 citation statements)

References 31 publications

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“…The target deconvolution of small molecules discovered through phenotypic screening remains a considerably significant challenge [45]. Novel techniques, such as mass spectrometrybased cellular thermal shift assay (CETSA), where the drug's binding to proteins in a cell or cell lysate is gauged by the degree of their thermal shifts [46], will be a comprehensive and unbiased method useful in identifying the molecular target or targets of HSG4112.…”

Section: Discussionmentioning

confidence: 99%

Discovery and preclinical efficacy of HSG4112, a synthetic structural analog of glabridin, for the treatment of obesity

Choi

Kang

et al. 2020

Int J Obes

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Background HSG4112 is a clinical-stage drug candidate for the treatment of obesity. Here, we report its discovery and preclinical efficacy. Methods In high-fat diet (HFD)-induced obese male C57BL/6J mice, we tested the weight loss effect of synthetic compounds derived from a structure–activity relationship (SAR) study of glabridin, a natural compound known to reduce body weight and influence energy homeostasis. After selecting HSG4112 as our optimized compound from this discovery method, we characterized its pharmacological effects on parameters related to obesity through in vivo metabolic and biochemical measurements, histology and gene expression analysis, and indirect calorimetry. Results Through the SAR study, we identified four novel components of glabridin pertinent for its anti-obesity activity, and found that HSG4112, an optimized structural analog of glabridin, markedly supersedes glabridin in weight reduction efficacy and chemical stability. Six-week administration of HSG4112 to HFD-induced obese mice led to dose-dependent normalization of obesity-related parameters, including body weight, muscle and adipose tissue weight, adipocyte size, and serum leptin/insulin/glucose levels. The weight reduction induced by HSG4112 was partially mediated by decreased food intake and mainly mediated by increased energy expenditure, with no change in physical activity. Accordingly, the pattern of transcriptional changes was aligned with increased energy expenditure in the liver and muscles. Following significant body weight reduction, robust amelioration of histopathology and blood markers of fatty liver were also observed. Conclusions Our study demonstrates the key chemical components of glabridin pertinent to its weight loss effects and suggests HSG4112 as a promising novel drug candidate for the pharmacological treatment of obesity.

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Section: Discussionmentioning

confidence: 99%

Discovery and preclinical efficacy of HSG4112, a synthetic structural analog of glabridin, for the treatment of obesity

Choi

Kang

et al. 2020

Int J Obes

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“…Knowledge of the direct target of a drug is essential for devising strategies to modulate the compound’s effects. Recently, new proteomics approaches have enabled the investigation of protein–drug interactions in native environments 2 – 4 . One such strategy relies on the use of chemical probes, which allow the enrichment and the identification of protein targets and may include the interaction sites as well 5 .…”

Section: Introductionmentioning

confidence: 99%

A machine learning-based chemoproteomic approach to identify drug targets and binding sites in complex proteomes

et al. 2020

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Chemoproteomics is a key technology to characterize the mode of action of drugs, as it directly identifies the protein targets of bioactive compounds and aids in the development of optimized small-molecule compounds. Current approaches cannot identify the protein targets of a compound and also detect the interaction surfaces between ligands and protein targets without prior labeling or modification. To address this limitation, we here develop LiP-Quant, a drug target deconvolution pipeline based on limited proteolysis coupled with mass spectrometry that works across species, including in human cells. We use machine learning to discern features indicative of drug binding and integrate them into a single score to identify protein targets of small molecules and approximate their binding sites. We demonstrate drug target identification across compound classes, including drugs targeting kinases, phosphatases and membrane proteins. LiP-Quant estimates the half maximal effective concentration of compound binding sites in whole cell lysates, correctly discriminating drug binding to homologous proteins and identifying the so far unknown targets of a fungicide research compound.

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“…This has turned out to be a powerful tool for target deconvolution after phenotypic screening campaigns as well as for the assessment of target specificity of a molecule. 31,32…”

Section: Introductionmentioning

confidence: 99%

A Tale of Two Tails: Efficient Profiling of Protein Degraders by Specific Functional and Target Engagement Readouts

et al. 2021

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Targeted protein degradation represents an area of great interest, potentially offering improvements with respect to dosing, side effects, drug resistance, and reaching “undruggable” proteins compared with traditional small-molecule therapeutics. A major challenge in the design and characterization of degraders acting as molecular glues is that binding of the molecule to the protein of interest (PoI) is not needed for efficient and selective protein degradation; instead, one needs to understand the interaction with the responsible ligase. Similarly, for proteasome targeting chimeras (PROTACs), understanding the binding characteristics of the PoI alone is not sufficient. Therefore, simultaneously assessing the binding to both PoI and the E3 ligase as well as the resulting degradation profile is of great value. The cellular thermal shift assay (CETSA) is an unbiased cell-based method, designed to investigate the interaction of compounds with their cellular protein targets by measuring compound-induced changes in protein thermal stability. In combination with mass spectrometry (MS), CETSA can simultaneously evaluate compound-induced changes in the stability of thousands of proteins. We have used CETSA MS to profile a number of protein degraders, including molecular glues (e.g., immunomodulatory drugs) and PROTACs, to understand mode of action and to deconvolute off-target effects in intact cells. Within the same experiment, we were able to monitor both target engagement by observing changes in protein thermal stability as well as efficacy by simultaneous assessment of protein abundances. This allowed us to correlate target engagement (i.e., binding to the PoI and ligases) and functional readout (i.e., degrader induced protein degradation).

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Mass spectrometry-based Cellular Thermal Shift Assay (CETSA®) for target deconvolution in phenotypic drug discovery

Cited by 52 publications

References 31 publications

Discovery and preclinical efficacy of HSG4112, a synthetic structural analog of glabridin, for the treatment of obesity

Discovery and preclinical efficacy of HSG4112, a synthetic structural analog of glabridin, for the treatment of obesity

A machine learning-based chemoproteomic approach to identify drug targets and binding sites in complex proteomes

A Tale of Two Tails: Efficient Profiling of Protein Degraders by Specific Functional and Target Engagement Readouts

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