The adoption of human-relevant New Approach Methodologies (NAMs) for non-clinical safety assessment has accelerated following recent regulatory and policy changes, including the FDA Modernisation Act 2.0 and the UK government’s Replacing Animals in Science strategy. In cardiac safety evaluation, assessing individual ion channels such as hERG alone may overlook the integrated electrophysiological mechanisms underlying proarrhythmic risk. Parameters derived from action potential (AP) waveform analysis, including AP duration, triangulation, engagement of repolarisation reserve and early afterdepolarisations (EADs), can provide additional mechanistic insight. This study evaluated whether optical voltage imaging of human iPSC-derived cardiomyocytes (hiPSC-CMs) can detect compound-induced electrophysiological changes beyond hERG inhibition and relate waveform alterations to underlying ion channel pharmacology.
Using fluorescence-based voltage recordings from hiPSC-CM monolayers, we assessed AP waveform parameters following 30 min and 24 h exposure to compounds with selective or multi-ion channel cardiac activity. Selective hERG inhibitors dofetilide and E-4031 produced pronounced increases in triangulation at low concentrations, while multi-ion channel compounds including verapamil, terfenadine, quinidine and ondansetron showed concentration-dependent effects consistent with modulation of inward and outward currents. Changes in APD30 provided additional differentiation of calcium channel effects, while EADs identified action potential instability at higher concentrations. Analysis of E-4031 further demonstrated concentration-dependent engagement of repolarisation reserve, illustrating how interactions between multiple ionic currents can shape the AP waveform. Overall, AP waveform analysis in hiPSC-CMs enabled mechanistic differentiation of multi-ion channel electrophysiological effects beyond hERG inhibition and AP duration alone, supporting its application as a human-relevant approach for cardiac safety assessment.
Development of a robust hNaV1.9 high-throughput screening assay on the Sophion Qube384 platform. This is complemented by a suite of ion channel selectivity assays and sensory neuron recordings to create a versatile screening cascade to support NaV1.9 drug discovery programmes.
Optical voltage imaging of human iPSC-derived cardiomyocytes was used to assess electrophysiological effects of compounds beyond hERG inhibition. Action potential waveform analysis revealed compound-specific and concentration-dependent changes, enabling mechanistic differentiation of multichannel activity and demonstrating a human-relevant approach for translational cardiac safety assessment.