Human induced pluripotent stem cell-derived cardiomyocytes (iPS-CMs) provide a novel assay system to assess cardiac safety in drug development to overcome a problem of species difference in non-clinical testing during drug development. Using the multi-electrode array (MEA) platform, electrophysiological activities of iPS-CMs can be recorded easily to assess QT prolongation and proarrhythmic potential of drug candidates. Here we have established a standardized protocol to evaluate the possibility of iPS-CMs, and shared the protocol with an international consortium. To obtain reproducible and reliable experimental data from these cells, we determined the optimal experimental conditions, such as cell density, MEA coating, culture conditions, high-pass filter frequency, definition of early afterdepolarization or triggered activity, and calibration compounds. Based on the protocol, our validation study using 60 compounds is in progress. Thus, MEA-based experiments using iPS-CMs would be a standard testing method to evaluate QT prolongation and proarrhythmic potentials.

Microphysiological systems (MPS) are gaining global attention as potential game-changers in pharmaceutical development. Since 2013, MPS suppliers from university laboratories in the United States and Europe have competed to develop these devices. After the development phase, the focus shifted to the accumulation of applications using MPS for pharmaceutical companies and end users. In Japan, the AMED-MPS project was launched in 2017, and since then, several MPS devices have been marketed by project participated suppliers. Initially, while Japanese pharmaceutical companies adopted foreign products, they also exhibited interest in domestically produced MPS devices. The utilization of new approach methodologies, including MPS, is expanding in the field of chemical substances risk assessment, and the Organization for Economic Co-operation and Development test guidelines are expected to adopt in vitro evaluation systems as alternatives to animal testing. This publication reviews global and Japanese trends surrounding MPS and outlines activities aimed at the regulatory acceptance of MPS as evaluation systems for medical drugs and chemicals.

Microphysiological system (MPS) are “Cell/tissue culture systems that reproduce in vivo organ functions in vitro by placing organ compartments that mimic the physiological environment of various organs such as the liver, small intestine, and lungs in micro-spaces.” The MPS are attracting attention around the world as tools to improve human predictability in drug discovery research. In the U.S., in 2012, the NIH (National Institutes of Health) allocated a large budget to academia for research development of MPS. In Japan, the National Institute of Advanced Industrial Science and Technology and the NIHS (National Institute of Health Sciences) have been playing a central role in commercialization, performance evaluation, and standardization of MPS devices developed by academia for the liver, small intestine, kidney, and BBB as target organs/tissues in the AMED–MPS project that started in 2017. Pharmaceutical companies are beginning to utilize MPS in drug discovery research. However, MPS have only just been raised as a topic of discussion between regulatory authorities and pharmaceutical companies, and it will be necessary to overcome many barriers before data obtained by MPS can be included in drug approval documents and be widely accepted administratively. In this review, I would like to introduce cardiac safety evaluation as a concrete example to show what paths MPS should take to gain regulatory approval. In addition, I would like also to introduce human 3D heart tissue, which was developed in NIHS, as a cardiac MPS.

TRIC-A is an intracellular cation channel enriched in excitable tissues that is recently identified as a key modulator of sarcoplasmic reticulum (SR) Ca²⁺ homeostasis through direct interaction with type 2 ryanodine receptors (RyR₂). Given the intimate anatomical and functional coupling between the SR and mitochondria, we investigated whether TRIC-A contributes to SR–mitochondrial crosstalk under cardiac stress conditions. Using a transverse aortic constriction (TAC) model, we found that TRIC-A^−/− mice developed more severe cardiac hypertrophy, underwent maladaptive remodeling, and activated apoptotic pathways compared with wild-type littermates. At the cellular level, TRIC-A-deficient cardiomyocytes were more susceptible to H₂O₂-induced mitochondrial injury and displayed abnormal mitochondrial morphology. Live-cell imaging revealed exaggerated mitochondrial Ca²⁺ uptake during caffeine stimulation and increased propensity for store-overload-induced Ca²⁺ release (SOICR). Complementary studies in HEK293 cells expressing RyR₂ demonstrated that exogenous TRIC-A expression attenuates RyR₂-mediated mitochondrial Ca²⁺ overload, preserves respiratory function, and suppresses superoxide generation. Together, these findings identify TRIC-A as a critical regulator of SR–mitochondrial Ca²⁺ signaling. By constraining mitochondrial Ca²⁺ influx and limiting oxidative stress, TRIC-A safeguards cardiomyocytes against SOICR-driven injury and confers protection against pressure overload-induced cardiac dysfunction.

Cardiotoxicity associated with hepatic metabolism and drug–drug interactions is a serious concern. Predicting drug toxicity using animals remains challenging due to species and ethical concerns, necessitating the need to develop alternative approaches. Drug cardiotoxicity associated with hepatic metabolism cannot be detected using a cardiomyocyte-only evaluation system. Therefore, we aimed to establish a system for evaluating cardiotoxicity via hepatic metabolism by co-culturing cryopreserved human hepatocytes (cryoheps) and human iPS cell-derived engineered heart tissues (hiPSC-EHTs) using a stirrer-based microphysiological system. We investigated candidate media to identify a medium that can be used commonly for hepatocytes and cardiomyocytes. We found that the contraction length was significantly greater in the HM Dex (-) medium, the medium used for cryohep culture without dexamethasone, than that in the EHT medium used for hiPSC-EHT culture. Additionally, the beating rate, contraction length, contraction speed, and relaxation speed of hiPSC-EHT cultured in the HM Dex (-) medium were stable throughout the culture period. Among the major CYPs, the expression of CYP3A4 alone was low in cryoheps cultured in the HM Dex (-) medium. However, improved oxygenation using the InnoCell plate increased CYP3A4 expression to levels comparable to those found in the human liver. In addition, CYP3A4 activity was also increased by the improved oxygenation. Furthermore, expression levels of hepatic function-related gene and nuclear receptors in cryoheps cultured in HM Dex (-) medium were comparable to those in the human liver. These results suggest that the HM Dex (-) medium can be applied to co-culture and may allow the evaluation of cardiotoxicity via hepatic metabolism. Moreover, CYP induction by typical inducers was confirmed in cryoheps cultured in the HM Dex (-) medium, suggesting that drug–drug interactions could also be evaluated using this medium. Our findings may facilitate the evaluation of cardiotoxicity via hepatic metabolism, potentially reducing animal testing, lowering costs, and expediting drug development.

Non-clinical pharmacological safety studies are conducted using cells and animals to ensure the safety of pharmaceuticals in humans. Following these studies, drug candidates are administered to humans during clinical trials. Safety must be sufficiently confirmed in non-clinical studies to ensure that test participants suffer no adverse health effects. However, due to species differences, low ability to extrapolate from in vitro to in vivo evaluation methods, and other problems, health hazards may unfortunately still occur. Therefore, sophisticated in vitro evaluation systems using human cells are actively being pursued. The main challenge remains the lack of a reliable methodology for extrapolating in vitro results to in vivo settings. We have attempted to extract parameters that can be predictably translated from in vitro [contractile evaluation in three-dimensional (3D) heart tissue] to in vivo (guinea pig echocardiography) conditions, using cardiac contractile dysfunction induced by anticancer drugs as an example. In this review, we introduce the in vitro methods developed to date to evaluate this cardiac contractile dysfunction, analyze the factors enabling highly accurate prediction of torsades de pointes in humans based on past proarrhythmic risk prediction methods using human induced pluripotent stem cell-derived cardiomyocytes, and apply them to evaluate cardiac contractile dysfunction caused by anticancer drugs using three-dimensional heart tissue. We also introduce the proposed strategy for this evaluation method in this section.