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Cardiovascular Engineering

Cardiovascular Engineering - Poster Session C

Poster H12 - Evaluation of Cardiomyocyte Spheroid Models as an Assay Platform for the Prediction of Clinical Cardiotoxicity

Friday, October 25, 2024
10:00 AM - 11:00 AM EST
Location: Exhibit Hall E, F & G

Introduction:  As new drugs are introduced to the market, it is pertinent that there are established processes to assess cardiac safety liabilities to reduce patient risk during development stages. Since these processes are virtually impossible to conduct safely in vivo, there is currently a critical need to produce clinically predictive, high-throughput, in vitro assays to apply in early phases of drug production to identify potential effects on myocardial function and any cardiotoxic consequences. Three of the most relevant testing platforms are 2D screening systems like MEA (monolayer cell culture), 3D screening systems based on bioengineered heart tissue (3D-HTH), and cardiomyocyte spheroids. The aim of this study is to provide a translational safety assay comparison between these predictive platforms, with a specific focus on the cardiomyocyte spheroid approach.

Materials and

Methods:  The first phase of experimentation focused on optimizing the formation of spheroids; testing was conducted to determine which ratio of fibroblasts and cardiomyocytes produced the best beat rate and cohesive spheroid shape. Two cell lines from different vendors (CDI and Celogics cardiomyocytes) were seeded in 384 ULA well plates and allowed to mature. After the necessary periods of time had elapsed (3 and 7 days respectively), the cells were stained with dye and beating patterns were obtained with a simultaneous 384 well reading in the FLIPR. The spheroid assay was further assessed using this calcium transient system; baseline calcium recordings were obtained as RFU, and spheroid contraction was captured by brightfield fluorescence video. A collection of sixteen compounds was then examined; calcium flux-acquired parameters and contraction patterns were recorded within the spheroid assay after exposure to the compounds and compared to pre-existing clinical data after reproducible peak parameters were analyzed, plotted, and inputted into heat maps. These comparisons were used to assess whether this platform accurately showed pharmacological effects at specific compound concentrations and thus could be utilized as a preclinical cardiovascular risk assessment for new treatments. The drugs tested in experimentation include Milrinone, Isoprenaline, Digoxin, Nifedipine, Verapamil, Thapsigargin, Flecainide, Quinidine, Mexiletine, Ranolazine, E-4031, JNJ303, Doxorubicin, Gemcitabine, and Sunitinib. These drugs were separated into 3 groups: inotropic modulators, ion channel modulators, and other cardiotoxic compounds. Individual concentrations were predetermined, and 4 different concentration conditions were tested on each cell type.

Results, Conclusions, and Discussions: The first phase of testing revealed that the optimal spheroid composition was 80% cardiomyocytes and 20% fibroblasts; this condition was used in the formation of spheroids from both the CDI and Celogics cell lines. Each cell line was exposed to 16 different drugs at 4 different concentrations; some wells were reserved for DMSO treatment, and no compound treatment, to act as a control. The parameter trends for peak amplitude and peak beat rate were plotted for each cell line after compound exposure, and results were normalized using data from wells treated with DMSO, and those not exposed to any treatment, before they were translated into heat map form. These heat maps displayed whether the parameters were upregulated or suppressed due to the drug exposure, and by extension whether the drugs were cardiotoxic. These results were compared to the results obtained by the other two modeling techniques and clinical reports. Comparisons revealed that drug responses were variable and dependent on the type of spheroid (CDI vs Celogics), and that the toxic effects might reduce the clinical predictability of this modeling technique. Future steps include retesting each cell line to obtain more reproducible results, testing clinically relevant doses of compounds, and using GCaMP biosensors to do time courses of drug addition and removal as opposed to the dye used during experimentation.

Acknowledgements (Optional):