Detection of Drug-Induced Torsades de Pointes Arrhythmia Mechanisms Using hiPSC-CM Syncytial Monolayers in a High-Throughput Screening Voltage Sensitive Dye Assay

Abstract

We validated 3 distinct hiPSC-CM cell lines—each of different purity and a voltage sensitive dye (VSD)-based high-throughput proarrhythmia screening assay as a noncore site in the recently completed CiPA Myocyte Phase II Validation Study. Blinded validation was performed using 12 drugs linked to low, intermediate, or high risk for causing Torsades de Pointes (TdP). Commercially sourced hiPSC-CMs were obtained either from Cellular Dynamics International (CDI, Madison, Wisconsin, iCell Cardiomyoyctes2) or Takara Bio (CLS, Cellartis Cardiomyocytes). A third hiPSC-CM cell line (MCH, Michigan) was generated in house. Each cell type had distinct baseline electrophysiological function (spontaneous beat rate, action potential duration, and conduction velocity) and drug responsiveness. Use of VSD and optical mapping enabled the detection of conduction slowing of sodium channel blockers (quinidine, disopyramide, and mexiletine) and drug-induced TdP-like activation patterns (rotors) for some high- and intermediate-risk compounds. Low-risk compounds did not induce rotors in any cell type tested. These results further validate the utility of hiPSC-CMs for predictive proarrhythmia screening and the utility of VSD technology to detect drug-induced APD prolongation, arrhythmias (rotors), and conduction slowing. Importantly, results indicate that different ratios of cardiomyocytes and noncardiomyocytes have important impact on drug response that may be considered during risk assessment of new drugs. Finally, we present the first blinded CiPA hiPSC-CM validation results to simultaneously detect drug-induced conduction slowing, action potential duration prolongation, action potential triangulation, and drug-induced rotors in a proarrhythmia assay.

In vitro model of ischemic heart failure using human induced pluripotent stem cell–derived cardiomyocytes

“…Following hiPSC-CM purification using MACS or metabolic stress selection, cells were replated as electrically and mechanically confluent monolayers for electrophysiological analysis as described before. (20,21,27) First electrophysiology of hiPSC-CM monolayers was investigated using sharp microelectrode recordings or optical mapping with a voltage sensitive dye. Figure 4A shows original action potential recordings and quantification of Maximal Diastolic Potential (MDP) and Action Potential Amplitude (APA).…”

“…These cells were handled, plated and used for optical mapping as outlined before. (20,21) Results of these experiments are presented in supplemental figure 5. First we generated a 96 well plate of these cells, using 50,000 cells per well to form monolayers.…”

“…The MACS approach and subsequent enrichment of the hiPSC-CMs was recently described and validated for use in cardiotoxicity proarrhythmia assays. (20,21) Figure 1A outlines the general cardiac directed differentiation and purification approaches used here. All functional and structural phenotype analysis was performed on syncytial hiPSC-CM monolayers.…”

From atoms to cells: bridging the gap between potency, efficacy, and safety of small molecules directed at a membrane protein

“…Altogether, the data give us an idea that the compound CDN1163 is not leading to significant electrical alterations in iPSC-derived cardiomyocytes. These findings are important because changes in CV and/or APD 90 are associated with pro-arrhythmia effects, such as re-entrant excitation or early/delayed afterdepolarizations 37,39,40 . Therefore, our data indicate that pharmacological stimulation of SERCA2a activity using a relatively selective activator does not induce pro-arrhythmogenic effects in human iPSC-derived cardiac cells.…”

“…Based on the available in situ EC 50 data acquired in this study, we tested CDN1163 and CP-154526 for stimulation of intracellular Ca 2+ dynamics on a 96-format well at compound concentrations of 0.1, 0.5, 1, 5, 10, and 50 µM ( n =6 per concentration), and Ro 41-0960 at compound concentrations of 0.5, 1, 5, 10, 50, and 100 µM ( n =6 per concentration). Additionally, we evaluated the effects of CDN1163 on conduction velocity and APD 90 on a 6-well plate format at compound concentrations of 0.5, 5, and 50 µM ( n =6 per concentration) to screen for potential cardiac toxicity effects induced by this molecule 37,64 . In all cases, data acquisition occurred before (baseline) and after treatment of cells with the compounds.…”