Abstract
Unexpected adverse effects on the cardiovascular system remain a major challenge in the development of novel active pharmaceutical ingredients (API). To overcome the current limitations of animal-based in vitro and in vivo test systems, stem cell derived human cardiomyocyte clusters (hCMC) offer the opportunity for highly predictable pre-clinical testing. The three-dimensional structure of hCMC appears more representative of tissue milieu than traditional monolayer cell culture. However, there is a lack of long-term, real time monitoring systems for tissue-like cardiac material. To address this issue, we have developed a microcavity array (MCA)-based label-free monitoring system that eliminates the need for critical hCMC adhesion and outgrowth steps. In contrast, feasible field potential derived action potential recording is possible immediately after positioning within the microcavity. Moreover, this approach allows extended observation of adverse effects on hCMC. For the first time, we describe herein the monitoring of hCMC over 35 days while preserving the hCMC structure and electrophysiological characteristics. Furthermore, we demonstrated the sensitive detection and quantification of adverse API effects using E4031, doxorubicin, and noradrenaline directly on unaltered 3D cultures. The MCA system provides multi-parameter analysis capabilities incorporating field potential recording, impedance spectroscopy, and optical read-outs on individual clusters giving a comprehensive insight into induced cellular alterations within a complex cardiac culture over days or even weeks.
Highlights
Enormous efforts and investments are made throughout the process of active pharmaceutical ingredients (API) development to identify adverse effects and minimize the risks for patients
While primary cardiomyocyte cells derived from animals have the advantage of allowing the concerted action of many ion channels involved in the action potential as well as structural analysis, interspecies differences in ion channel composition and signal cascades can hinder predictivity when extrapolating from non-human experimental models to the clinical setting [1,4]
In contrast to planar Microelectrode array (MEA), where field potential-derived action potential duration (fAPD) detection is strictly dependent on the adhesion quality of the cells, the microcavity array (MCA) does not require cell adhesion to the electrode
Summary
Enormous efforts and investments are made throughout the process of API development to identify adverse effects and minimize the risks for patients. The pharmaceutical industry has an urgent need for assays that are applicable at an early stage in the process of API development which, at the same time, provide highly predictive and detailed information for the human in vivo situation [1]. This is relevant for the assessment of arrhythmia caused by new chemical entities. The majority of drug-induced adverse events in humans involve a delay in repolarization of the cardiac action potential and an action potential duration (APD) prolongation This is correlated with the prolongation of the clinical relevant QT interval indicating delayed ventricular repolarization and is routinely used as a biomarker of life-threatening ventricular arrhythmia e.g. Torsades de Pointes (TdP). While primary cardiomyocyte cells derived from animals have the advantage of allowing the concerted action of many ion channels involved in the action potential as well as structural analysis, interspecies differences in ion channel composition and signal cascades can hinder predictivity when extrapolating from non-human experimental models to the clinical setting [1,4]
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