A Targeted Metabolomics-Based Assay Using Human Induced Pluripotent Stem Cell-Derived Cardiomyocytes Identifies Structural and Functional Cardiotoxicity Potential

Abstract

Implementing screening assays that identify functional and structural cardiotoxicity earlier in the drug development pipeline has the potential to improve safety and decrease the cost and time required to bring new drugs to market. In this study, a metabolic biomarker-based assay was developed that predicts the cardiotoxicity potential of a drug based on changes in the metabolism and viability of human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CM). Assay development and testing was conducted in 2 phases: (1) biomarker identification and (2) targeted assay development. In the first phase, metabolomic data from hiPSC-CM spent media following exposure to 66 drugs were used to identify biomarkers that identified both functional and structural cardiotoxicants. Four metabolites that represent different metabolic pathways (arachidonic acid, lactic acid, 2′-deoxycytidine, and thymidine) were identified as indicators of cardiotoxicity. In phase 2, a targeted, exposure-based biomarker assay was developed that measured these metabolites and hiPSC-CM viability across an 8-point concentration curve. Metabolite-specific predictive thresholds for identifying the cardiotoxicity potential of a drug were established and optimized for balanced accuracy or sensitivity. When predictive thresholds were optimized for balanced accuracy, the assay predicted the cardiotoxicity potential of 81 drugs with 86% balanced accuracy, 83% sensitivity, and 90% specificity. Alternatively, optimizing the thresholds for sensitivity yields a balanced accuracy of 85%, 90% sensitivity, and 79% specificity. This new hiPSC-CM-based assay provides a paradigm that can identify structural and functional cardiotoxic drugs that could be used in conjunction with other endpoints to provide a more comprehensive evaluation of a drug’s cardiotoxicity potential.

Industry Updates from the Field of Stem Cell Research and Regenerative Medicine in February 2020

“…Stemina (WI, USA; www.stemina.com) has published study results showing the high performance of the company's Cardio quickPredict test in identifying cardiotoxic effects for a large number of well-characterized drugs [5,6]. These results suggest the drug screening test could provide drug developers with an early, highly accurate and more comprehensive evaluation of cardiovascular toxicity.…”

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

“…The majority of screening assays developed over recent years have focused on predicting functional cardiotoxicity, i.e., change in the mechanical function of the myocardium, while molecular biomarkers that can predict drug-induced morphological damage of the myocardium and loss of cardiac viability (structural cardiotoxicity) are still lacking [ 8 , 12 , 18 ]. As such, mechanistic insight and predictive biomarkers of structural cardiotoxicity remain a significant unmet need [ 2 , 8 , 12 , 18 ]. Untargeted metabolomics is a promising approach in toxicology for generating hypotheses on toxicity modes of action [ 19 , 20 , 21 , 22 , 23 ] and for discovering metabolic biomarkers [ 18 ].…”

“…As such, mechanistic insight and predictive biomarkers of structural cardiotoxicity remain a significant unmet need [ 2 , 8 , 12 , 18 ]. Untargeted metabolomics is a promising approach in toxicology for generating hypotheses on toxicity modes of action [ 19 , 20 , 21 , 22 , 23 ] and for discovering metabolic biomarkers [ 18 ]. Metabolomics-based in vivo studies have previously detected metabolic perturbations associated with drug-induced cardiotoxicity [ 24 , 25 ].…”

“…Metabolomics-based in vivo studies have previously detected metabolic perturbations associated with drug-induced cardiotoxicity [ 24 , 25 ]. Furthermore, the technology has successively been applied to measure changes to the extracellular metabolome, termed the ‘metabolic footprint’, of hiPSC-CMs induced by cardiotoxins, with the responses of four metabolites (arachidonic acid, lactic acid, 2′-deoxycytidine and thymidine) proposed as biomarkers predictive of the cardiotoxicity potential of drugs [ 18 ]. Measurement of the metabolic footprint of an in vitro model can be achieved by conventional ultra-high performance liquid chromatography-mass spectrometry (UHPLC-MS)-based analyses of spent culture medium, which can be relatively simply and non-invasively sampled [ 18 , 26 ].…”

“…Furthermore, the technology has successively been applied to measure changes to the extracellular metabolome, termed the ‘metabolic footprint’, of hiPSC-CMs induced by cardiotoxins, with the responses of four metabolites (arachidonic acid, lactic acid, 2′-deoxycytidine and thymidine) proposed as biomarkers predictive of the cardiotoxicity potential of drugs [ 18 ]. Measurement of the metabolic footprint of an in vitro model can be achieved by conventional ultra-high performance liquid chromatography-mass spectrometry (UHPLC-MS)-based analyses of spent culture medium, which can be relatively simply and non-invasively sampled [ 18 , 26 ]. However, although it provides an indication of molecular disruption to the in vitro model, the metabolic footprint does not represent all intracellular responses [ 26 , 27 ].…”