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Impedance based Assay on iPS cardiomyocytes: An efficient way of discriminating between cardioactive and non cardioactive drugs
Abstracts
the question: what is a minimum cardiomyocyte? We conducted comparison studies of human stem cell derived-CM from different sources, cultured over two different time points to allow benchmarking of commercially available stem cell derived cardiomyocytes in terms of functionality and use for drug discovery or safety assessment. This multifactorial evaluation including genetic, structural and electrophysiological measurements of iPSC-CM, ES-CM, adult cardiac stem cell derived CM and cardiac progenitors benchmarked against neonatal and adult human tissue may aid to fill the current lack of standardized guidelines for the utilization of stem cell technology for pre-clinical research.
doi:10.1016/j.vascn.2016.02.149
0153 A highly scalable embryonic stem cell platform for in vitro genetics of drug response Jacqueline Bergseid, Shivanjali Joshi-Barr, Lesley Page, Amy Carlo, Stephanie Brown, Ted Choi Predictive Biology, Inc., Carlsbad, CA, USA We have developed an in vitro genetics platform to identify causal genes, pathways and mechanisms that underlie drug response. The technology combines embryonic stem cells (ESC) with complex trait genetics and high throughput screening in a scalable, broadly applicable technology. Using this approach we can identify causal genes at least 100-fold more efficiently than animal studies or human genomewide association studies (GWAS), which can provide valuable molecular, mechanistically relevant information on a compound or compound series. We produced a panel of 1000 genetically diverse mouse embryonic stem cell lines, which are differentiated to build in vitro genetic screening panels of pharmacologically relevant cell types. The strategy is to rapidly identify causal, mechanistically relevant genes in mouse cells, then test and validate the orthologous human genes and pathways in human cells, combining the power of mouse genetics with the relevance of pharmacological response in human cells. Data will be presented from two large studies underway at Predictive, a cytotoxicity screen of 80 compounds (currently used drugs and environmental toxicants) across 300 ES cell lines, and a cardiotoxicity screen of 30 compounds (anthracyclines, TKIs and CIPA genes) using a novel high throughput visual beating assay developed at Predictive.
doi:10.1016/j.vascn.2016.02.150
0154 Impedance based Assay on iPS cardiomyocytes: An efficient way of discriminating between cardioactive and non cardioactive drugs Aurore Colomar, Lucille Haen, Gaëlle Toussaint, Simon Loïodice, Benjamin Kopp, Franck Atienzar, Jean-Pierre Valentin
development. The use of cardiomyocytes derived from induced Pluripotent Stem cell (iPSC) to detect these liabilities during early drug discovery was investigated. We evaluated the effects of 69 reference drugs on Cell Index (CI), Beating Rate, Amplitude of Contraction (AC) and Beating Rhythm Irregularity on mouse (m)iPSC by impedance based measurements. 13/14 TdP/QTc compounds, 4/5 sodium blockers, 5/5 multi-ion blockers, 4/4 calcium blockers and 10/11 structural cardiotoxicants induced effects on miPSC. Inotropic and chronotropic compounds induced the expected changes in amplitude of contraction or beating rhythm in 8/19 and 4/9 cases respectively. A pattern associating decrease in AC, CI and/or increase of Irregularity was the most common profile of effects. In contrast, 8/8 negative drugs induced no effects or only increase in AC. Condensing the analysis to 3 parameters (i.e. decrease in CI, AC or increase in Irregularity) led to a sensitivity and a specificity of 78.2% and 100% discriminating cardioactive versus non cardioactive drugs. 14 of those drugs were tested also in human (h)iPSC. Effects were seen with higher magnitude at equal or lower concentrations in human compared to mouse iPSC, however hiPSC were less stable in control conditions. In summary, impedance based assay on miPSC allowed to detect cardioactive versus non cardioactive drugs with high sensitivity and specificity.
doi:10.1016/j.vascn.2016.02.151
0155 Cardiotoxicity testing in human iPSC-derived cardiomyocytes for the CiPA pilot study using all-optical electrophysiology Graham Dempsey, Nicholas Atwater, Cuong Nguyen Q-State Biosciences, Cambridge, MA, USA A primary endpoint of the CiPA initiative is to evaluate stem cellderived cardiomyocytes as a cellular substrate for cardiotoxicity testing. We have developed an all-optogenetic platform called Cardiac Optopatch that enables simultaneous stimulation and detection of electrical activity in hiPSC-derived cardiomyocytes. Stimulation is achieved with a channelrhodopsin variant, CheRiff, which provides a stable electrophysiological background. Simultaneous fluorescent detection of action potential (AP) waveforms and calcium transients (CTs) is achieved using a fast, sensitive dualreporter protein called CaViar (calcium and voltage indicator). As part of the CiPA pilot study, we used this platform to test the effects of eight ion channel modulatory compounds on cell lines from two providers, CDI and Axiogenesis. Measurement of electrophysiological parameters included changes in AP waveforms (APD30, APD60, APD90, triangulation, AP rise time, spontaneous beat rate) and CTs (amplitude) at the spontaneous beat rate and at two paced frequencies (1 Hz, 2 Hz). Both cell lines showed AP prolongation upon addition of hERG blockers (moxifloxacin, E-4031). Sodium channel blockers (ranolazine, flecainide, mexiletine, quinidine) led to an increase in rise time and AP prolongation, though larger in effect for CDI cells. The calcium channel blocker nifedipine reduced CT amplitude. EADs were detected in CDI cells for quinidine, flecainide and E-4031. Axiogenesis cells exhibited a faster spontaneous beat rate and slower upstroke compared to CDI cells. Neither cell line showed significant changes upon addition of JNJ313, an IKs blocker. These results demonstrate the utility of Cardiac Optopatch to be used for assessment of drug responses in hiPSC-derived cardiomyocytes.
UCB Biopharma, Braine l’Alleud, Belgium Drug induced cardiac liabilities, such as pro-arrhythmia, chronotropy, inotropy and structural cardiotoxicity, may affect drugs under
381
doi:10.1016/j.vascn.2016.02.152
the question: what is a minimum cardiomyocyte? We conducted comparison studies of human stem cell derived-CM from different sources, cultured over two different time points to allow benchmarking of commercially available stem cell derived cardiomyocytes in terms of functionality and use for drug discovery or safety assessment. This multifactorial evaluation including genetic, structural and electrophysiological measurements of iPSC-CM, ES-CM, adult cardiac stem cell derived CM and cardiac progenitors benchmarked against neonatal and adult human tissue may aid to fill the current lack of standardized guidelines for the utilization of stem cell technology for pre-clinical research.
doi:10.1016/j.vascn.2016.02.149
0153 A highly scalable embryonic stem cell platform for in vitro genetics of drug response Jacqueline Bergseid, Shivanjali Joshi-Barr, Lesley Page, Amy Carlo, Stephanie Brown, Ted Choi Predictive Biology, Inc., Carlsbad, CA, USA We have developed an in vitro genetics platform to identify causal genes, pathways and mechanisms that underlie drug response. The technology combines embryonic stem cells (ESC) with complex trait genetics and high throughput screening in a scalable, broadly applicable technology. Using this approach we can identify causal genes at least 100-fold more efficiently than animal studies or human genomewide association studies (GWAS), which can provide valuable molecular, mechanistically relevant information on a compound or compound series. We produced a panel of 1000 genetically diverse mouse embryonic stem cell lines, which are differentiated to build in vitro genetic screening panels of pharmacologically relevant cell types. The strategy is to rapidly identify causal, mechanistically relevant genes in mouse cells, then test and validate the orthologous human genes and pathways in human cells, combining the power of mouse genetics with the relevance of pharmacological response in human cells. Data will be presented from two large studies underway at Predictive, a cytotoxicity screen of 80 compounds (currently used drugs and environmental toxicants) across 300 ES cell lines, and a cardiotoxicity screen of 30 compounds (anthracyclines, TKIs and CIPA genes) using a novel high throughput visual beating assay developed at Predictive.
doi:10.1016/j.vascn.2016.02.150
0154 Impedance based Assay on iPS cardiomyocytes: An efficient way of discriminating between cardioactive and non cardioactive drugs Aurore Colomar, Lucille Haen, Gaëlle Toussaint, Simon Loïodice, Benjamin Kopp, Franck Atienzar, Jean-Pierre Valentin
development. The use of cardiomyocytes derived from induced Pluripotent Stem cell (iPSC) to detect these liabilities during early drug discovery was investigated. We evaluated the effects of 69 reference drugs on Cell Index (CI), Beating Rate, Amplitude of Contraction (AC) and Beating Rhythm Irregularity on mouse (m)iPSC by impedance based measurements. 13/14 TdP/QTc compounds, 4/5 sodium blockers, 5/5 multi-ion blockers, 4/4 calcium blockers and 10/11 structural cardiotoxicants induced effects on miPSC. Inotropic and chronotropic compounds induced the expected changes in amplitude of contraction or beating rhythm in 8/19 and 4/9 cases respectively. A pattern associating decrease in AC, CI and/or increase of Irregularity was the most common profile of effects. In contrast, 8/8 negative drugs induced no effects or only increase in AC. Condensing the analysis to 3 parameters (i.e. decrease in CI, AC or increase in Irregularity) led to a sensitivity and a specificity of 78.2% and 100% discriminating cardioactive versus non cardioactive drugs. 14 of those drugs were tested also in human (h)iPSC. Effects were seen with higher magnitude at equal or lower concentrations in human compared to mouse iPSC, however hiPSC were less stable in control conditions. In summary, impedance based assay on miPSC allowed to detect cardioactive versus non cardioactive drugs with high sensitivity and specificity.
doi:10.1016/j.vascn.2016.02.151
0155 Cardiotoxicity testing in human iPSC-derived cardiomyocytes for the CiPA pilot study using all-optical electrophysiology Graham Dempsey, Nicholas Atwater, Cuong Nguyen Q-State Biosciences, Cambridge, MA, USA A primary endpoint of the CiPA initiative is to evaluate stem cellderived cardiomyocytes as a cellular substrate for cardiotoxicity testing. We have developed an all-optogenetic platform called Cardiac Optopatch that enables simultaneous stimulation and detection of electrical activity in hiPSC-derived cardiomyocytes. Stimulation is achieved with a channelrhodopsin variant, CheRiff, which provides a stable electrophysiological background. Simultaneous fluorescent detection of action potential (AP) waveforms and calcium transients (CTs) is achieved using a fast, sensitive dualreporter protein called CaViar (calcium and voltage indicator). As part of the CiPA pilot study, we used this platform to test the effects of eight ion channel modulatory compounds on cell lines from two providers, CDI and Axiogenesis. Measurement of electrophysiological parameters included changes in AP waveforms (APD30, APD60, APD90, triangulation, AP rise time, spontaneous beat rate) and CTs (amplitude) at the spontaneous beat rate and at two paced frequencies (1 Hz, 2 Hz). Both cell lines showed AP prolongation upon addition of hERG blockers (moxifloxacin, E-4031). Sodium channel blockers (ranolazine, flecainide, mexiletine, quinidine) led to an increase in rise time and AP prolongation, though larger in effect for CDI cells. The calcium channel blocker nifedipine reduced CT amplitude. EADs were detected in CDI cells for quinidine, flecainide and E-4031. Axiogenesis cells exhibited a faster spontaneous beat rate and slower upstroke compared to CDI cells. Neither cell line showed significant changes upon addition of JNJ313, an IKs blocker. These results demonstrate the utility of Cardiac Optopatch to be used for assessment of drug responses in hiPSC-derived cardiomyocytes.
UCB Biopharma, Braine l’Alleud, Belgium Drug induced cardiac liabilities, such as pro-arrhythmia, chronotropy, inotropy and structural cardiotoxicity, may affect drugs under
381
doi:10.1016/j.vascn.2016.02.152