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UNIVERSITY OF BUCHAREST
FACULTY OF PHYSICS Guest
2024-11-22 2:22 |
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Conference: Bucharest University Faculty of Physics 2018 Meeting
Section: Biophysics; Medical Physics
Title:
Automated data analysis and optimization algorithms for electrophysiology and pharmacology studies on human induced pluripotent stem cell-derived cardiomyocytes
Authors:
B. AMUZESCU (1), R. AIRINI (1), F. EPUREANU (1), A. F. DEFTU (1), A. T. DEFTU (1), B. RADU (1), D. F. MIHĂILESCU (1), S. MANN (2), J. HEIDE (2), T. KNOTT (3)
*
Affiliation:
1 Dept. Biophysics&Physiology, Faculty of Biology, University of Bucharest,
Romania
2 CytoBioScience GmbH, Cologne, Germany
3 CytoBioScience Inc., San Antonio, TX, USA
E-mail
bogdan@biologie.kappa.ro
Keywords:
cardiac safety pharmacology, cardiac in vitro pro-arrhythmia assay (CiPA), human induced pluripotent stem cell-derived cardiomyocytes, automated patch-clamp, automated data analysis, O'Hara-Rudy2011 model, optimization algorithm
Abstract:
Cardiac safety pharmacology faces a paradigm shift from the “hERG-centric” approach, focused entirely on testing inhibitory effects of drugs on the main cardiac repolarizing K+ current IKr, contributed by hERG channels, to the more comprehensive CiPA (cardiac in vitro pro-arrhythmia assay), a mechanistic 3-step approach addressing multiple cardiac ion channels that may rescue some potential drugs, rejected previously based on hERG testing, without being proarrhythmogenic. We are currently developing an original CiPA assay combining all 3 steps (test of multiple inhibitory effects, in silico modeling using an advanced human cardiomyocyte electrophysiology model like O’Hara-Rudy2011 - ORd2011, and validation of predicted proarrhytmogenic effects via experiments on human induced pluripotent stem cell-derived cardiomyocytes - hiPSC-CM) in a single experiment on a hiPSC-CM approached with the automated patch-clamp platform CytoPatch™2 based on Cytocentering® technology. We used a commercial hiPSC-CM preparation (Cor.4U®, provided by Ncardia), and we developed a complex assay combining several voltage-clamp and current-clamp protocols applied before and during drug exposure. With an automated analysis script we obtained from voltage-clamp recordings applied in physiological conditions (without specific blockers) estimates of surface conductance densities for INa, ICaL, If, Ito, sum of IKr and IKs. Using perforated whole-cell patch-clamp with beta-escin resulted in improved stability in time of current densities and action potential (AP) shape. We modified the ORd2011 model by including temperature dependence of gating for most ion currents (from own experiments) and a 4-state circular allosteric model of If (Männikkö et al., 2005), and then we used this adapted model within a sequential algorithm in order to minimize the sum of squared differences between recorded and model-generated APs, obtaining accurate fits of experimental data.
Acknowledgement:
The study was funded from Competitiveness Operational Programme 2014-2020 project P_37_675 (146/2016), Priority Axis 1, Action 1.1.4, co-financed by the European Funds for Regional Development and Romanian Government.
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