Spontaneously Beating Cardiac Microtissues: 3D Structural Cardiovascular Toxicity Assay

Detect structural cardiotoxicity of novel therapeutics using Cyprotex’s spontaneously beating tri-cultured cardiac 3D microtissue high content screening (HCS) assay.

Cyprotex delivers consistent, high quality data with the flexibility to adapt protocols based on specific customer requirements.


  • Drug induced cardiovascular toxicity is the leading cause of attrition during drug development. Drugs can exert functional toxicities such as arrhythmia and morphological (structural) damage to the myocardium1. Evaluation of the potential for both types of cardiotoxicity by novel compounds is essential for the discovery of safe drugs.
  • The myocardial tissue comprises 30% cardiomyocytes and 70% non-myocytes, the majority of which are endothelial and fibroblast cells. These non-myocytes are essential to myocardial structure and function2,3 with emerging evidence suggesting important roles within drug induced cardiovascular toxicity4.
  • Mitochondrial disruption, calcium dyshomeostasis and cellular ATP content have been identified as major targets for structural cardiotoxins5.
  • Three dimensional (3D) confocal HCS allows the simultaneous detection of each cell health parameter in combination with a measure of cellular ATP.


3D Microtissue Based Cardiotoxicity Assay Protocol


Data from Cyprotex's 3D Structural Cardiovascular Toxicity Assay

All reference compound toxicities were correctly predicted in the spontaneously beating cardiac tri-culture 3D microtissue model including isoproterenol (MEC 2.1 µM, calcium dyshomeostasis (Table 1 and Figure 2)) and cyclophosphamide (MEC 30.8 µM, mitochondrial mass (Table 1)) which previously went undetected by Pointon et al (2013)5 and Cyprotex’s in-house H9c2 data.

Control compound sunitinib displays cytosolic calcium increase (calcium dyshomeostasis) followed by gross cytotoxicity (microtissue loss) (Figure 2a) while control compound dasatinib displays mitochondrial membrane potential loss without gross cytotoxicity (microtissue loss) (Figure 2b). The combination of an in vitro 3D model that better recapitulates the in vivo cellular physiology of the myocardium with a multiparametric HCS and cytotoxicity assay presents a viable screening strategy for the accurate detection of novel therapeutics that cause drug induced structural cardiovascular toxicity early in drug development.


1) Laverty HG et al., (2011). How can we improve our understanding of cardiovascular safety liabilities to develop safer medicines? Br J Pharmacol 163(4); 675-693
2) Brutsaert DL (2003). Cardiac endothelial-myocardial signaling: Its role in cardiac growth, contractile performance, and rhythmicity. Phys Revs 83; 59-115.
3) Souders CA et al., (2009). Cardiac fibroblast:the renaissance cell. Circ Res 105; 1164-1176
4) Mikaelian I et al., (2010). Primary endothelial damage is the mechanism of cardiotoxicity of tubulin-binding drugs. Tox Sci 117(1); 144-151
5) Pointon A et al., (2013) Phenotypic profiling of structural cardiotoxins in vitro reveals dependency on multiple mechanisms of toxicity. Tox Sci 132(2); 317-326
6) Nam KH et al., (2015) Biomimetic 3D tissue models for advanced high-throughput drug screening. J Lab Autom; In press
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11) Kerkelä R et al., (2006). Cardiotoxicity of the cancer therapeutic agent imatinib mesylate. Nat Med 12(8); 908-916
12)  Chu TF et al., (2007). Cardiotoxicity associated with tyrosine kinase inhibitor sunitinib. Lancet 370; 2011-2019
13) Floyd JD et al., (2005). Cardiotoxicity of cancer therapy. J Clin Oncol 23(30); 7685-7696
14) Zhang J et al., (2008). Isoproterenol-induced cardiotoxicity in Sprague-Dawley rats: correlation of reversible and irreversible myocardial injury with release of cardiac troponin T and roles of iNOS in myocardial injury. Toxicol Pathol 36(2); 277-278


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Sam Bevan

Sam Bevan

Principal Scientist

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Cyprotex enables and enhances the prediction of human exposure, clinical efficacy and toxicological outcome of a drug or chemical. By combining quality data from robust in vitro methods with contemporary in silico technology, we add value, context and relevance to the ADME-Tox data supplied to our partners in the pharmaceutical or chemical industries.