Assessment of beating parameters in human induced pluripotent stem cells enables quantitative in vitro screening for cardiotoxicity

Abstract

Human induced pluripotent stem cell (iPSC)-derived cardiomyocytes show promise for screening during early drug development. Here, we tested a hypothesis that in vitro assessment of multiple cardiomyocyte physiological parameters enables predictive and mechanistically-interpretable evaluation of cardiotoxicity in a high-throughput format. Human iPSC-derived cardiomyocytes were exposed for 30 minutes or 24 hours to 131 drugs, positive (107) and negative (24) for in vivo cardiotoxicity, in up to 6 concentrations (3 nM to 30 μM) in 384-well plates. Fast kinetic imaging was used to monitor changes in cardiomyocyte function using intracellular Ca2+ flux readouts synchronous with beating, and cell viability. A number of physiological parameters of cardiomyocyte beating, such as beat rate, peak shape (amplitude, width, raise, decay, etc.) and regularity were collected using automated data analysis. Concentration-response profiles were evaluated using logistic modeling to derive a benchmark concentration (BMC) point-of-departure value, based on one standard deviation departure from the estimated baseline in vehicle (0.3% dimethylsulfoxide)-treated cells. BMC values were used for cardiotoxicity classification and ranking of compounds. Beat rate and several peak shape parameters were found to be good predictors, while cell viability had poor classification accuracy. In addition, we applied the Toxicological Prioritization Index approach to integrate and display data across many collected parameters, to derive “cardiosafety” ranking of tested compounds. Multi-parameter screening of beating profiles allows for cardiotoxicity risk assessment and identification of specific patterns defining mechanism-specific effects. The data and analysis methods may be used widely for compound screening and early safety evaluation in the drug development process.

Automated Video-Based Analysis of Contractility and Calcium Flux in Human-Induced Pluripotent Stem Cell-Derived Cardiomyocytes Cultured over Different Spatial Scales

“…Based on the electrochemical activation underlying these changes, high-throughput in vitro assays are being developed to monitor the electrical behavior of either single cells or clusters of primary and pluripotent cell-derived cardiomyocytes. [2][3][4] These high-throughput assays complement traditional lower-throughput patch-clamp electrophysiology, and are complemented by novel genetic tools for imaging calcium transients. 5,6 However, these approaches either require that cardiomyocytes be plated onto specialized materials (e.g., multielectrode arrays, microelectrode arrays [MEAs], or synthetic hydrogels embedded with fluorescent particles to track strain 7 ) or involve genetic manipulations.…”

“…Hence, image-processing tools have been applied in attempts to turn such qualitative observations into quantitative data. 3,[8][9][10] Popular image-processing techniques include edge detection and simple pixel intensity monitoring. 8,10 These techniques require the user to specify regions of interest to monitor, which introduces inherent bias.…”

Evaluation of the effects of power-frequency magnetic fields on the electrical activity of cardiomyocytes differentiated from human induced pluripotent stem cells

“…These cause differences in each species' tolerance to chemical agents (Price et al, 2008), and might yield false-positive or -negative results when human toxicity is evaluated using experimental animals and cells. Tests using many kinds of known cardioactive compounds have shown that hiPS-CMs are useful for evaluation of pharmacological responsiveness in humans (Sirenko et al, 2013;Khan et al, 2013). Thus, assay systems using hiPS-CMs are likely to provide accurate and efficient screening of candidate drugs.…”

“…CMs derived from hiPS cells (hiPS-CMs) are artificially generated source of human tissue, and their potential for use in medical and pharmacological applications has been intensively examined (Funakoshi et al, 2016;Kawamura et al, 2012;Khan et al, 2013;Sirenko et al, 2013). In the pharmacological field, hiPS-CMs have been confirmed to show appropriate responsiveness to known cardioactive compounds (Sirenko et al, 2013), and hiPS-CMs are expected to contribute to efficient screening of candidate drugs (Khan et al, 2013). HiPS-CMs are becoming a powerful tool to evaluate human responses.…”

Current approaches and future role of high content imaging in safety sciences and drug discovery

“…Case study e: the aim of the study was to investigate if the in vitro assessment of multiple cardiomyocyte physiological parameters enables predictive and mechanistically interpretable evaluation of cardiotoxicity at high throughput level (Sirenko et al, 2013). Human iPSC-derived cardiomyocytes were exposed for 30 min or 24 h to 131 drugs (incl.…”