Detect therapeutically relevant pathophysiological hypertrophic cardiotoxicity potential of novel therapeutics using Cyprotex’s 3D combined cardiac hypertrophy and multi-parametric high content screening (HCS) cardiotoxicity assay.
Cyprotex deliver consistent, high quality data with the flexibility to adapt protocols based on specific customer requirements.
Numerous studies have shown that cell responses to drugs in 3D culture are improved from those in 2D, with respect to modeling in vivo tissue functionality, which highlights the advantages of using 3D-based models for preclinical drug screens.
5Nam KH et al., (2015) Biomimetic 3D tissue models for advanced high-throughput drug screening. J Lab Autom 20(3); 201-215
|Spheroid||Induced pluripotent stem cell (iPSC) derived cardiomyocytes|
|Analysis Platform||Brightfield & Confocal Cellomics ArrayScan® XTI (Thermo Scientific).|
|8 point dose response curve with top concentration based on 100x Cmax or solubility limit*
3 replicates per concentration*
|150 µL of a DMSO* solution to achieve 100x Cmax (200x top concentration to maintain 0.5% DMSO) or equivalent amount in solid compound.|
|Time Points||Spheroid hypertrophy: day 3, 7, 10 & 14*
Structural cardiotoxicity HCS & ATP: day 14*
|Quality Controls||Negative control: 0.5% DMSO (vehicle)*
Positive controls: dasatinib (structural cardiotoxin with pathophysiological hypertrophic potential) and mitomycin C (structural cardiotoxicity without hypertrophic potential)
|Data Delivery||Minimum effective concentration (MEC) and AC50 value for each measured parameter; spheroid count and spheroid size (3, 7, 10 & 14 days) and DNA structure (DNA), calcium homeostasis (Ca2+) mitochondrial mass (Mito Mass), mitochondrial membrane potential (MMP) and cellular ATP content (ATP) (Day 14)*|
*Other options available on request.
|Drug||Human exposure Cmax (µM)||In vivo cardiac structural toxicity (P/N)||In vivo cardiac patho-physiological hypertrophy (P/N)||Most senstive structural MEC (µM)||Most sensitive hypertrophy MEC (µM)||Most sensitive combined assay (MEC µM)||Most sensitive feature|
|≤ 1x Cmax|
|≤ 3x Cmax|
|≤ 10x Cmax|
|≥ 10x Cmax|
|Structural toxicity potential||Patho-physiological hypertrophy potential||Cardiac toxicity|
|Correct prediction with a 10x Cmax cut off (%)||94%||81%||100%|
Utilizing the 3D cardiac combined assay approach all reference compound toxicities were correctly predicted within a 10x Cmax cut off. Structural cardiotoxicity was correctly predicted for 94% and pathophysiological hypertrophy potential (PHP) for 81% of the compound set within a 10x Cmax cut off.
The combination of an in vitro 3D model that better recapitulates the in vivo cellular physiology of cardiac tissue with multiparametric temporal HCS and a cytotoxicity assay presents a viable screening strategy for the accurate in vivo relevant detection of novel therapeutics that cause structural cardiotoxicity with pathophysiological hypertrophy potential early in drug development.
1 Laverty H 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 Rev 83(1), 59-11
3 Pointon A et al., (2013) Phenotypic profiling of structural cardiotoxins in vitro reveals dependency on multiple mechanisms of toxicity. Toxicol Sci 132(2), 317-326
4 Cross MJ et al., (2015) Physiological, pharmacological and toxicological considerations of drug-induced structural cardiac injury. Br J Pharmacol 172(4), 957-974
5 Nam KH et al., (2015) Biomimetic 3D tissue models for advanced high-throughput drug screening. J Lab Autom 20(3); 201-215