Detect structural cardiotoxicity of novel therapeutics using Cyprotex’s spontaneously beating tri-cultured cardiac 3D microtissue high content screening (HCS) 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.
6Nam KH, Smith AS, Lone S, Kwon S and Kim DH (2015) Biomimetic 3D Tissue Models for Advanced High-Throughput Drug Screening. J Lab Autom; In press
|Microtissue||Human induced pluripotent stem cell derived cardiomyocytes (iPSC-CM’s), cardiac endothelial cells and cardiac fibroblasts|
|Analysis Platform||Confocal Cellomics ArrayScan® XTI (Thermo Scientific)|
|Test Article Concentration||8 point dose response curve with top concentration based on 100x Cmax or solubility limit
3 replicates per concentration
|Test Article Requirements||50 µL of a DMSO solution at a concentration of 200x top concentration (top concentration = 100x Cmax) or equivalent amount in solid compound|
|Time Points||72 hours (others available on request)|
|Quality Controls||Negative control: 0.5% DMSO (vehicle)
Positive controls: Sunitinib (Ca2+ homeostasis) and dasatinib (mitochondrial membrane potential)
|Data Delivery||Minimum effective concentration (MEC) and AC50 value for each measured parameter (microtissue count, microtissue size, DNA structure, calcium homeostasis (Ca2+), mitochondrial mass (Mito Mass), mitochondrial membrane potential (MMP) and cellular ATP content)|
|Compound||Cmax (µM)||In vivo toxicity||hESC-CM prediction (Pointon et al, 2013)||H9c2 monolayer||H9c2 MTs||Tri-culture cardiac MTs||Most Sensitive Feature|
|Dasatinib7||0.72||Structural cardiotoxin||Positive structural cardiotoxin||0.529||NR||2.08||MMP|
|Cyclophosphamide13||153.20||Negative structural cardiotoxins||NR||NR||30.8||Mito Mass|
|Buspirone HCl||0.03||NR||0.237||NR||Microtissue size|
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.
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Learn more about toxicology in our popular Mechanisms of Drug-Induced Toxicity guide