One-Pot Time-Induced Proteome Integral Solubility Alteration Assay for Automated and Sensitive Drug–Target Identification DOI
Zhaowei Meng, Amir Ata Saei,

Hezheng Lyu

et al.

Analytical Chemistry, Journal Year: 2024, Volume and Issue: unknown

Published: Nov. 20, 2024

The proteome integral solubility alteration (PISA) assay is widely used for identifying drug targets, but it labor-intensive and time-consuming requires a substantial amount of biological sample. Aiming at enabling automation greatly reducing the sample amount, we developed one-pot time-induced (OPTI)-PISA. Here, demonstrate OPTI-PISA performance on targets multiple drugs in cell lysate scaling down to sub-microgram levels, making PISA method suitable NanoProteomics. can be implemented using only standard equipment proteomics lab.

Language: Английский

Tapioca: a platform for predicting de novo protein–protein interactions in dynamic contexts DOI

Tavis. J. Reed,

Matthew D. Tyl, Alicja Tadych

et al.

Nature Methods, Journal Year: 2024, Volume and Issue: 21(3), P. 488 - 500

Published: Feb. 15, 2024

Language: Английский

Citations

11
High-throughput drug target discovery using a fully automated proteomics sample preparation platform DOI Creative Commons
Qiong Wu, Jiangnan Zheng,

Xintong Sui

et al.

Chemical Science, Journal Year: 2024, Volume and Issue: 15(8), P. 2833 - 2847

Published: Jan. 1, 2024

Drug development is plagued by inefficiency and high costs due to issues such as inadequate drug efficacy unexpected toxicity. Mass spectrometry (MS)-based proteomics, particularly isobaric quantitative offers a solution unveil resistance mechanisms unforeseen side effects related off-targeting pathways. Thermal proteome profiling (TPP) has gained popularity for target identification at the scale. However, it involves experiments with multiple temperature points, resulting in numerous samples considerable variability large-scale TPP analysis. We propose high-throughput discovery workflow that integrates single-temperature TPP, fully automated proteomics sample preparation platform (autoSISPROT), data independent acquisition (DIA) quantification. The autoSISPROT enables simultaneous processing of 96 less than 2.5 hours, achieving protein digestion, desalting, optional TMT labeling (requires an additional 1 hour) 96-channel all-in-tip operations. results demonstrated excellent performance >94% digestion efficiency, >98% >0.9 intra- inter-batch Pearson correlation coefficients. By automatically 87 samples, we identified both known targets potential off-targets 20 kinase inhibitors, affording over 10-fold improvement throughput compared classical TPP. This target/off-target identification.

Language: Английский

Citations

10
Poly-pharmacology of existing drugs: How to crack the code? DOI Creative Commons
Baptiste Mouysset, Marion Le Grand, Luc Camoin

et al.

Cancer Letters, Journal Year: 2024, Volume and Issue: 588, P. 216800 - 216800

Published: March 14, 2024

Drug development in oncology is highly challenging, with less than 5% success rate clinical trials. This alarming figure points out the need to study more details multiple biological effects of drugs specific contexts. Indeed, comprehensive assessment drug poly-pharmacology can provide insights into their therapeutic and adverse effects, optimize utilization maximize Recent technological advances have made possible in-depth investigation poly-pharmacology. review first highlights high-throughput methodologies that been used unveil new mechanisms action existing drugs. Then, we discuss how emerging chemo-proteomics strategies allow effectively dissecting an unsupervised manner.

Language: Английский

Citations

5
Stability-based approaches in chemoproteomics DOI Creative Commons
Amy L. George, María Emilia Dueñas, José Luis Marín‐Rubio

et al.

Expert Reviews in Molecular Medicine, Journal Year: 2024, Volume and Issue: 26

Published: Jan. 1, 2024

Abstract Target deconvolution can help understand how compounds exert therapeutic effects and accelerate drug discovery by helping optimise safety efficacy, revealing mechanisms of action, anticipate off-target identifying opportunities for expansion. Chemoproteomics, a combination chemical biology with mass spectrometry has transformed target deconvolution. This review discusses modification-free chemoproteomic approaches that leverage the change in protein thermodynamics induced small molecule ligand binding. Unlike modification-based methods relying on enriching specific targets, these offer proteome-wide evaluations, driven advancements sensitivity, increasing proteome coverage quantitation methods. Advances based denaturation/precipitation thermal or denaturation, protease degradation are evaluated, emphasising evolving landscape chemoproteomics its potential impact future drug-development strategies.

Language: Английский

Citations

5
Benchmarking of quantitative proteomics workflows for Limited proteolysis mass spectrometry DOI Creative Commons

Tomas Koudelka,

Claudio Bassot, Ilaria Piazza

et al.

Molecular & Cellular Proteomics, Journal Year: 2025, Volume and Issue: unknown, P. 100945 - 100945

Published: March 1, 2025

Limited proteolysis coupled with mass spectrometry (LiP-MS) has emerged as a powerful technique for detecting protein structural changes and drug-protein interactions on proteome-wide scale. However, there is no consensus the best quantitative proteomics workflow analyzing LiP-MS data. In this study, we comprehensively benchmarked two major quantification approaches-data-independent acquisition (DIA) tandem tag (TMT) isobaric labeling-in combination LiP-MS, using drug-target deconvolution assay model system. Our results show that while TMT labeling enabled of more peptides proteins lower coefficients variation (CVs), DIA-MS exhibited greater accuracy in identifying true drug targets stronger dose-response correlation peptides. Additionally, evaluated performance freely available (FragPipe) versus commercial (Spectronaut) software tools analysis, revealing choice between precision sensitivity largely depends specific experimental context. findings underscore importance selecting appropriate strategy based study objectives. This work provides valuable guidelines researchers discovery, highlights how advancements instrumentation, such Astral spectrometer, may further improve sequence coverage, potentially reducing need labeling.

Language: Английский

Citations

0
Streamlined analysis of drug targets by proteome integral solubility alteration indicates organ-specific engagement DOI Creative Commons
Tanveer S. Batth, Marie Locard‐Paulet, Nadezhda T. Doncheva

et al.

Nature Communications, Journal Year: 2024, Volume and Issue: 15(1)

Published: Oct. 16, 2024

Proteins are the primary targets of almost all small molecule drugs. However, even most selectively designed drugs can potentially target several unknown proteins. Identification potential drug facilitate design new and repurposing existing ones. Current state-of-the-art proteomics methodologies enable screening thousands proteins against a limited number molecules. Here we report development label-free quantitative approach that enables proteome-wide organic molecules in scalable, reproducible, rapid manner by streamlining proteome integral solubility alteration (PISA) assay. We used rat organs ex-vivo to determine organ specific medical enzyme inhibitors identify for common such as Ibuprofen. Finally, global profiling revealed overarching trends how affect through either direct or indirect protein interactions.

Language: Английский

Citations

3
Toward the analysis of functional proteoforms using mass spectrometry-based stability proteomics DOI Creative Commons

Ji Hye Kang,

Meena Seshadri,

Kellye A. Cupp‐Sutton

et al.

Frontiers in Analytical Science, Journal Year: 2023, Volume and Issue: 3

Published: June 22, 2023

Functional proteomics aims to elucidate biological functions, mechanisms, and pathways of proteins proteoforms at the molecular level examine complex cellular systems disease states. A series stability methods have been developed protein functionality by measuring resistance a chemical or thermal denaturation proteolysis. These can be applied measure thousands in samples such as cell lysate, intact cells, tissues, other fluids proteome stability. Stability popularly observe shifts upon ligand binding for drug target identification. More recently, these characterize effect structural changes those caused post-translational modifications (PTMs) mutations, which affect structures interactions diversify functions. Here, we discussed current application suite methods, including profiling (TPP), from rates oxidation (SPROX), limited proteolysis (LiP) PTM-induced on We also discuss future perspectives highlighting integration top-down mass spectrometry proteoform understand function variable modifications.

Language: Английский

Citations

7
Chemoproteomics validates selective targeting of Plasmodium M1 alanyl aminopeptidase as an antimalarial strategy DOI Creative Commons
Carlo Giannangelo, Matthew Challis, Ghizal Siddiqui

et al.

eLife, Journal Year: 2024, Volume and Issue: 13

Published: Feb. 2, 2024

New antimalarial drug candidates that act via novel mechanisms are urgently needed to combat malaria resistance. Here, we describe the multi-omic chemical validation of Plasmodium M1 alanyl metalloaminopeptidase as an attractive target using selective inhibitor, MIPS2673. MIPS2673 demonstrated potent inhibition recombinant falciparum ( Pf A-M1) and vivax Pv metalloaminopeptidases, with selectivity over other human aminopeptidases, displayed excellent in vitro activity no significant host cytotoxicity. Orthogonal label-free chemoproteomic methods based on thermal stability limited proteolysis whole parasite lysates revealed solely targets A-M1 parasites, also enabling estimation binding site within ~5 Å determined by X-ray crystallography. Finally, functional investigation untargeted metabolomics inhibits key role haemoglobin digestion. Combined, our unbiased deconvolution confirmed on-target MIPS2673, validated a promising strategy.

Language: Английский

Citations

2
Experimental and data analysis advances in thermal proteome profiling DOI Creative Commons

Amanda M. Figueroa‐Navedo,

Alexander R. Ivanov

Cell Reports Methods, Journal Year: 2024, Volume and Issue: 4(2), P. 100717 - 100717

Published: Feb. 1, 2024

Method development for mass spectrometry (MS)-based thermal shift proteomic assays have advanced to probe small molecules with known and unknown protein-ligand interaction mechanisms specificity, which is predominantly used in characterization of drug-protein interactions. In the discovery target off-target interactions, a thorough investigation method their impact on sensitivity accuracy protein-small molecule protein-protein interactions warranted. this review, we discuss areas improvement at each stage proteome profiling data analysis that includes processing MS-based data, development, effect overall quality profiles. We also overview optimization experimental strategies prioritization an increased number independent biological replicates over evaluated temperatures.

Language: Английский

Citations

2
On the utility of ultrafast MS1-only proteomics in drug target discovery studies based on thermal proteome profiling method DOI
Ivan I. Fedorov, Julia A. Bubis, Elizaveta M. Kazakova

et al.

Analytical and Bioanalytical Chemistry, Journal Year: 2024, Volume and Issue: 416(18), P. 4083 - 4089

Published: May 15, 2024

Language: Английский

Citations

2