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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.

SUBMITTER: Palmer JA

PROVIDER: S-EPMC7098380 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:Cardiotoxicity is a major cause of high attrition rates among newly developed drugs. Moreover, anti-cancer treatment-induced cardiotoxicity is one of the leading reasons of mortality in cancer survivors. Cardiotoxicity screening in vitro may improve predictivity of cardiotoxicity by novel drugs, using human pluripotent stem cell (hPSC)-derived-cardiomyocytes. Anthracyclines, including Doxorubicin, are widely used and highly effective chemotherapeutic agents for the treatment of different forms of malignancies. Unfortunately, anthracyclines cause many cardiac complications early or late after therapy. Anthracyclines exhibit their potent anti-cancer effect primarily via induction of DNA damage during the DNA replication phase in proliferative cells. In contrast, studies in animals and hPSC-cardiomyocytes have revealed that cardiotoxic effects particularly arise from (1) the generation of oxidative stress inducing mitochondrial dysfunction, (2) disruption of calcium homeostasis, and (3) changes in transcriptome and proteome, triggering apoptotic cell death. To increase the therapeutic index of chemotherapeutic Doxorubicin therapy several protective strategies have been developed or are under development, such as (1) reducing toxicity through modification of Doxorubicin (analogs), (2) targeted delivery of anthracyclines specifically to the tumor tissue or (3) cardioprotective agents that can be used in combination with Doxorubicin. Despite continuous progress in the field of cardio-oncology, cardiotoxicity is still one of the major complications of anti-cancer therapy. In this review, we focus on current hPSC-cardiomyocyte models for assessing anthracycline-induced cardiotoxicity and strategies for cardioprotection. In addition, we discuss latest developments toward personalized advanced pre-clinical models that are more closely recapitulating the human heart, which are necessary to support in vitro screening platforms with higher predictivity. These advanced models have the potential to reduce the time from bench-to-bedside of novel antineoplastic drugs with reduced cardiotoxicity.

Project description:Despite preclinical studies demonstrating the functional benefit of transplanting human pluripotent stem cell-derived cardiomyocytes (PSC-CMs) into damaged myocardium, the ability of these immature cells to adopt a more adult-like cardiomyocyte (CM) phenotype remains uncertain. To address this issue, we tested the hypothesis that prolonged in vitro culture of human embryonic stem cell (hESC)- and human induced pluripotent stem cell (hiPSC)-derived CMs would result in the maturation of their structural and contractile properties to a more adult-like phenotype. Compared to their early-stage counterparts (PSC-CMs after 20-40 days of in vitro differentiation and culture), late-stage hESC-CMs and hiPSC-CMs (80-120 days) showed dramatic differences in morphology, including increased cell size and anisotropy, greater myofibril density and alignment, sarcomeres visible by bright-field microscopy, and a 10-fold increase in the fraction of multinucleated CMs. Ultrastructural analysis confirmed improvements in the myofibrillar density, alignment, and morphology. We measured the contractile performance of late-stage hESC-CMs and hiPSC-CMs and noted a doubling in shortening magnitude with slowed contraction kinetics compared to the early-stage cells. We then examined changes in the calcium-handling properties of these matured CMs and found an increase in calcium release and reuptake rates with no change in the maximum amplitude. Finally, we performed electrophysiological assessments in hESC-CMs and found that late-stage myocytes have hyperpolarized maximum diastolic potentials, increased action potential amplitudes, and faster upstroke velocities. To correlate these functional changes with gene expression, we performed qPCR and found a robust induction of the key cardiac structural markers, including ?-myosin heavy chain and connexin-43, in late-stage hESC-CMs and hiPSC-CMs. These findings suggest that PSC-CMs are capable of slowly maturing to more closely resemble the phenotype of adult CMs and may eventually possess the potential to regenerate the lost myocardium with robust de novo force-producing tissue.

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

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