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Bowen TJ, Hall AR, Lloyd GR, et al. An Extensive Metabolomics Workflow to Discover Cardiotoxin-Induced Molecular Perturbations in Microtissues. Metabolites. 2021;11(9)doi: 10.3390/metabo11090644.
Bowen, T. J., Hall, A. R., Lloyd, G. R., Weber, R. J. M., Wilson, A., Pointon, A., & Viant, M. R. (2021). An Extensive Metabolomics Workflow to Discover Cardiotoxin-Induced Molecular Perturbations in Microtissues. Metabolites, 11(9), . https://doi.org/10.3390/metabo11090644
Bowen, Tara J, et al. "An Extensive Metabolomics Workflow to Discover Cardiotoxin-Induced Molecular Perturbations in Microtissues." Metabolites vol. 11,9 (2021). doi: https://doi.org/10.3390/metabo11090644
Bowen TJ, Hall AR, Lloyd GR, Weber RJM, Wilson A, Pointon A, Viant MR. An Extensive Metabolomics Workflow to Discover Cardiotoxin-Induced Molecular Perturbations in Microtissues. Metabolites. 2021 Sep 21;11(9). doi: 10.3390/metabo11090644. PMID: 34564460; PMCID: PMC8470535.
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Metabolites. 2021 Sep 21;11(9). doi: 10.3390/metabo11090644.

An Extensive Metabolomics Workflow to Discover Cardiotoxin-Induced Molecular Perturbations in Microtissues.

Metabolites

Tara J Bowen, Andrew R Hall, Gavin R Lloyd, Ralf J M Weber, Amanda Wilson, Amy Pointon, Mark R Viant

Affiliations

  1. School of Biosciences, University of Birmingham, Edgbaston, Birmingham B15 2TT, UK.
  2. Functional and Mechanistic Safety, Clinical Pharmacology and Safety Sciences, R&D, AstraZeneca, Cambridge CB4 0WG, UK.
  3. Phenome Centre Birmingham, University of Birmingham, Edgbaston, Birmingham B15 2TT, UK.
  4. Clinical Pharmacology and Quantitative Pharmacology, Clinical Pharmacology and Safety Sciences, BioPharmaceuticals R&D, AstraZeneca, Cambridge CB4 0WG, UK.

PMID: 34564460 PMCID: PMC8470535 DOI: 10.3390/metabo11090644

Abstract

Discovering modes of action and predictive biomarkers of drug-induced structural cardiotoxicity offers the potential to improve cardiac safety assessment of lead compounds and enhance preclinical to clinical translation during drug development. Cardiac microtissues are a promising, physiologically relevant, in vitro model, each composed of ca. 500 cells. While untargeted metabolomics is capable of generating hypotheses on toxicological modes of action and discovering metabolic biomarkers, applying this technology to low-biomass microtissues in suspension is experimentally challenging. Thus, we first evaluated a filtration-based approach for harvesting microtissues and assessed the sensitivity and reproducibility of nanoelectrospray direct infusion mass spectrometry (nESI-DIMS) measurements of intracellular extracts, revealing samples consisting of 28 pooled microtissues, harvested by filtration, are suitable for profiling the intracellular metabolome and lipidome. Subsequently, an extensive workflow combining nESI-DIMS untargeted metabolomics and lipidomics of intracellular extracts with ultra-high performance liquid chromatography-mass spectrometry (UHPLC-MS/MS) analysis of spent culture medium, to profile the metabolic footprint and quantify drug exposure concentrations, was implemented. Using the synthetic drug and model cardiotoxin sunitinib, time-resolved metabolic and lipid perturbations in cardiac microtissues were investigated, providing valuable data for generating hypotheses on toxicological modes of action and identifying putative biomarkers such as disruption of purine metabolism and perturbation of polyunsaturated fatty acid levels.

Keywords: biomarkers; cardiac microtissues; cardiotoxicity; in vitro metabolomics; mode of action; sample harvesting; sensitivity; untargeted toxicokinetics

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