THE CardioECR/iCELL SYSTEM @ CHARLES RIVER A tool for - PowerPoint PPT Presentation

THE CardioECR/iCELL SYSTEM @ CHARLES RIVER A tool for excitation-contraction coupling in cardiac safety and drug discovery EVERY STEP OF THE WAY EVERY STEP OF THE WAY

MECHANISTIC ANALYSIS OF CONTRACTION INOTROPIC EFFECTS

TOOLS xCELLigence RTCA CardioECR iCELL cardiomyocytes Drugs: Blebbistatin Nifedipine FPL 64176 Omecamtiv mecarbil 3 EVERY STEP OF THE WAY

THE CardioECR/ICell SYSTEM Cell Index = (Impedance @ time t – Impedance @ time 0 ) / 15 Ω Cell Index = (R tn -R t0 )/15 Ω N=45 4 EVERY STEP OF THE WAY

THE IMPEDANCE SIGNAL MATLAB Analysis Grand Average Raw data Local averages 5 EVERY STEP OF THE WAY

MOTION-DEPENDENT EVENTS: BLEBBISTATIN 6 EVERY STEP OF THE WAY

EXCITATION-CONTRACTION COUPLING: L-TYPE CA2+ MODULATORS NIFEDIPINE (BLOCKER) FPL 64176 (AGONIST) 7 EVERY STEP OF THE WAY

MOTION-DEPENDENT EVENTS: OMECAMTIV MECARBIL Contraction: Positive inotropy Omecamtiv mecarbil 1 µM Impedance recordings (CardioECR) 50,000 iCELL 2 cardiomyocytes CL: 1500 ms 8 EVERY STEP OF THE WAY

CONCLUSION Discovery Application • Enhanced software algorithms allow detailed investigation of excitation- contraction coupling and contraction mechanisms. • The system responds to known inotropic agents in a predictive way and could be a valuable tool to investigate contraction in a medium throughput plate-based assay. 9 EVERY STEP OF THE WAY

MECHANISTIC ANALYSIS OF ARRYTHMIC LIABILITY

ARRHYTHMIC LIABILITY You want this … FIELD POTENTIAL IMPEDANCE 11 EVERY STEP OF THE WAY

ARRHYTHMIC LIABILITY But some times you get this at the Cmax ( 140 nM ) Ibutilide 100 nM FIELD POTENTIAL IMPEDANCE 12 EVERY STEP OF THE WAY

ARRHYTHMIC LIABILITY Moxifloxacin: Cmax 11 µM Baseline 10 µM 30 µM 100 µM 300 µM 13 EVERY STEP OF THE WAY

ARRHYTHMIC LIABILITY The CardioECR/iCELL system detects delayed repolarization and arrhythmias 14 EVERY STEP OF THE WAY

CONCLUSION Cardiac Safety Application The CardioECR/iCELL system is a valuable tool to investigate proarrhythmic liability (CiPA) • The system responds to known pharmacological agents in a predictive way. • When tailored with appropriate disease cell models the CardioECR/iCell system is a valuable tool to generate • cardiac disease biomarkers. 15 EVERY STEP OF THE WAY

MECHANISTIC ANALYSIS OF CARDIOTOXICITY

CARDIOTOXIC DRUGS: LONG TERM EFFECTS Vehicle: 8 days 17 EVERY STEP OF THE WAY

CARDIOTOXIC DRUGS: LONG TERM EFFECTS Vehicle and Doxorubicin: 8 days 18 EVERY STEP OF THE WAY

CARDIOTOXIC DRUGS: LONG TERM EFFECTS Effect of Tyrosine Kinase Inhibitors on basal impedance 19 EVERY STEP OF THE WAY

CARDIOTOXIC DRUGS: LONG TERM EFFECTS Crizotinib: Effects at Therapeutic Doses 20 EVERY STEP OF THE WAY

CARDIOTOXIC DRUGS: LONG TERM EFFECTS Erlotinib: Supratherapeutic Effects 21 EVERY STEP OF THE WAY

Ouabain Shortens the FPD (QT-like Signal) • Cardiac glycoside, increases intracellular calcium • Low dose increases contractile force, has been used as heart failure therapy • High dose causes cardiac arrest (used for poison arrows)

Ouabain Does Not Inhibit Ion Channel Currents Ion Channel Inhibition 100.0 Digoxin Ouabain 90.0 80.0 hERG 70.0 Percent Inhibition Nav1.5 60.0 KvLQT1/minK 50.0 Kir2.1 40.0 Cav1.2 30.0 20.0 10.0 0.0 0.03 0.1 0.3 0.03 0.1 0.3 Concentration ( µ M)

Ouabain Toxicity is Revealed in Contraction Measure (Impedance) 10 nM 20 nM 40 nM 5 nM Vehicle 2.5 nM 1 hr 2 hr 3 hr 4 hr

OVERALL CONCLUSIONS • The CardioECR/iCell system is a valuable tool to investigate: • Contractility • Proarrhythmic liability (CiPA) • Cardiotoxicity • Medium throughput analysis of human cells (integrated system) detects liability that would be missed using in vitro ion screening strategies alone 25 EVERY STEP OF THE WAY

CONTACT US Andrew Bruening-Wright, PhD Principal Scientist Safety Assessment | Charles River 251 Ballardvale Street askcharlesriver@crl.com Wilmington, MA 01887 www.criver.com 877.CRIVER.1