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1.
Article in English | MEDLINE | ID: mdl-28163191

ABSTRACT

INTRODUCTION: The use of multi-electrode arrays (MEA) in combination with human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs) provides a promising method to predict comprehensive cardiotoxicity, including drug-induced QT prolongation and arrhythmia. We previously demonstrated that MEA in combination with hiPSC-CMs could provide a generalizable platform by using 7 reference drugs at 10 testing facilities. Using this approach, we evaluated responses to reference drugs that modulate a range of cardiac ion currents and have a range of arrhythmogenic effects. METHODS: We used the MEA system (MED64) and commercially available hiPSC-CMs (iCell cardiomyocytes) to evaluate drug effects on the beat rate, field potential duration (FPD), FPD corrected by Fridericia's formula (FPDc), and the incidence of arrhythmia-like waveforms. RESULTS: This assay detected the repolarization effects of Bay K8644, mibefradil, NS1643, levcromakalim, and ouabain; and the chronotropic effects of isoproterenol, ZD7288, and BaCl2. Chronotropy was also affected by K+ and Ca2+ current modulation. This system detected repolarization delays and the arrhythmogenic effects of quinidine, cisapride, thioridazine, astemizole, bepridil, and pimozide more sensitively than the established guinea pig papillary muscle action potential assay. It also predicted clinical QT prolongation by drugs with multiple ion channel effects (fluoxetine, amiodarone, tolterodine, vanoxerine, alfuzosin, and ranolazine). DISCUSSION: MEA in combination with hiPSC-CMs may provide a powerful method to detect various cardiac electrophysiological effects, QT prolongation, and arrhythmia during drug discovery. However, the data require careful interpretation to predict chronotropic effects and arrhythmogenic effects of candidate drugs with multiple ion channel effects.


Subject(s)
Arrhythmias, Cardiac/chemically induced , Cardiotoxins/pharmacology , Heart Rate/drug effects , Induced Pluripotent Stem Cells/drug effects , Ion Channels , Myocytes, Cardiac/drug effects , Arrhythmias, Cardiac/physiopathology , Cardiotonic Agents/pharmacology , Cells, Cultured , Dose-Response Relationship, Drug , Drug Evaluation, Preclinical/methods , Heart Rate/physiology , Humans , Induced Pluripotent Stem Cells/physiology , Ion Channels/agonists , Ion Channels/antagonists & inhibitors , Ion Channels/physiology , Myocytes, Cardiac/physiology
2.
J Pharmacol Toxicol Methods ; 78: 93-102, 2016.
Article in English | MEDLINE | ID: mdl-26657830

ABSTRACT

INTRODUCTION: Drug-induced QT prolongation is a major safety issue during drug development because it may lead to lethal ventricular arrhythmias. In this study, we evaluated the utility of multi-electrode arrays (MEA) with human induced pluripotent stem cell-derived cardiomyocytes (hiPS-CMs) to predict drug-induced QT prolongation and arrhythmia. METHODS: Ten facilities evaluated the effects of 7 reference drugs (E-4031, moxifloxacin, flecainide, terfenadine, chromanol 293B, verapamil, and aspirin) using a MED64 MEA system with commercially available hiPS-CMs. Field potential duration (FPD), beat rate, FPD corrected by Fridericia's formula (FPDc), concentration inducing FPDc prolongation by 10% (FPDc10), and incidence of arrhythmia-like waveform were evaluated. RESULTS: The inter-facility variability of absolute values before drug application was similar to the intra-facility variability for FPD, beat rate, and FPDc. The inter-facility variability of FPDc10 for 5 reference drugs ranged from 1.8- to 5.8-fold. At all 10 facilities, E-4031, moxifloxacin, and flecainide prolonged FPDc and induced arrhythmia-like waveforms at concentrations 1.8- to 6.1-fold higher than their FPDc10. Terfenadine prolonged FPDc and induced beating arrest at 8.0 times the FPDc10. The average FPDc10 values for E-4031, moxifloxacin, and terfenadine were comparable to reported plasma concentrations that caused QT prolongation or Torsade de Pointes in humans. Chromanol 293B, a IKs blocker, also prolonged FPDc but did not induce arrhythmia-like waveforms, even at 7.4 times the FPDc10. In contrast, verapamil shortened FPDc and aspirin did not affect FPDc or FP waveforms. DISCUSSION: MEA with hiPS-CMs can be a generalizable method for accurately predicting both QT prolongation and arrhythmogenic liability in humans.


Subject(s)
Arrhythmias, Cardiac/chemically induced , Cell Culture Techniques/methods , Drug-Related Side Effects and Adverse Reactions , Induced Pluripotent Stem Cells/drug effects , Long QT Syndrome/chemically induced , Myocytes, Cardiac/drug effects , Arrhythmias, Cardiac/diagnosis , Congresses as Topic , Cryopreservation/methods , Drug-Related Side Effects and Adverse Reactions/diagnosis , Humans , Induced Pluripotent Stem Cells/physiology , Long QT Syndrome/diagnosis , Myocytes, Cardiac/physiology , Pharmaceutical Preparations/administration & dosage , Predictive Value of Tests
3.
J Toxicol Sci ; 37(3): 527-37, 2012.
Article in English | MEDLINE | ID: mdl-22687992

ABSTRACT

The 26-week oral toxicity of diheptyl phthalate (DHP), a peroxisome proliferator-activated receptor α (PPARα) agonist, with special emphasis on the potential induction of hepatocellular proliferative lesions was investigated in this study. DHP was administered to male F344 rats via gavage at 0 (control), 1,000 or 2,000 mg/kg/day for 26 weeks. Body weight gain was significantly lower, whereas food and water consumption was significantly higher in DHP-treated rats compared with controls. DHP-treated rats exhibited decreases in blood triglyceride, total cholesterol, phospholipid and glucose levels, which were likely related to biological effects of the PPARα agonist. Absolute and relative organ weights of the livers with pale brown discoloration and dark brown spots significantly increased in DHP-treated rats. Histopathological examinations revealed remarkable diffuse hypertrophy of hepatocytes with ground-glass appearance, intracytoplasmic inclusion bodies and/or vacuolation in the DHP-treated groups. These findings were associated with increases in serum aspartate aminotransferase, alanine aminotransferase, alkaline phosphatase, and γ-glutamyltranspeptidase. The number and area of glutathione S-transferase placental form positive foci, a marker of hepatocellular preneoplastic lesions in rats, significantly increased in DHP-treated groups. Additionally, proliferating cell nuclear antigen positive liver cell counts in DHP-treated groups were significantly higher than those of the controls. Testicular alterations were not detected histopathologically, whereas absolute and relative prostate weights significantly decreased at both doses. These results indicate that DHP induces liver pre-neoplastic foci, and suggest the possibility that DHP is a possible genotoxic carcinogen in the liver of rats.


Subject(s)
Cell Proliferation/drug effects , Liver Neoplasms/pathology , Liver/drug effects , Liver/pathology , Phthalic Acids/toxicity , Precancerous Conditions/pathology , Administration, Oral , Animals , Carcinogenicity Tests , Carcinogens/toxicity , Glutathione Transferase/metabolism , Hepatocytes/drug effects , Hepatocytes/pathology , Liver/metabolism , Liver Neoplasms/chemically induced , Male , Organ Size/drug effects , Precancerous Conditions/chemically induced , Rats , Rats, Inbred F344
4.
J Pharmacol Toxicol Methods ; 59(3): 128-34, 2009.
Article in English | MEDLINE | ID: mdl-19281853

ABSTRACT

INTRODUCTION: : We examined the effects of methylphenidate hydrochloride (MPH) on the cardiovascular system using in vivo and in vitro study methods in accordance with the ICH-S7B guideline. METHODS: MPH was orally administered at doses of 3, 10 and 30 mg/kg to unrestrained conscious dogs implanted with a telemetry transmitter and attached with body surface electrodes, and electrocardiogram (ECG) leads. The QTcF interval was determined while heart rate (HR), and blood pressure (BP) were measured. Action potentials in isolated guinea-pig papillary muscle and the rapid component of the delayed rectifier potassium current (I(Kr)) in HEK-293 cells stably transfected with hERG were also investigated at concentrations of 0.1, 0.3 and 1 microg/mL (0.37, 1.1 and 3.7 micromol/L) of MPH. RESULTS: No ECG changes were observed except for a shortening of the QT interval due to a shortening of the RR interval at the maximum dose tested, 30 mg/kg. The only observed change was an elevation of BP in dogs at the dose of 30 mg/kg, which is approximately 10 times higher than the maximum therapeutic dose for use in children with attention deficit hyperactivity disorder (ADHD). Neither APD prolongation nor I(Kr) inhibition was observed by MPH in the in vitro studies up to the maximum concentration tested, 1 microg/mL (3.7 micromol/L), which is approximately 34 times higher than the clinically attainable unbound plasma MPH concentrations in children with ADHD. DISCUSSION: These results suggest that it is unlikely that MPH affects ventricular repolarization processes at the therapeutically recommended dose levels in patients with ADHD.


Subject(s)
Cardiovascular System/drug effects , Central Nervous System Stimulants/pharmacology , Heart Conduction System/drug effects , Methylphenidate/pharmacology , Action Potentials/drug effects , Administration, Oral , Animals , Blood Pressure/drug effects , Cell Line , Central Nervous System Stimulants/adverse effects , Delayed Rectifier Potassium Channels/antagonists & inhibitors , Dogs , Dose-Response Relationship, Drug , Electrocardiography , Ether-A-Go-Go Potassium Channels/genetics , Ether-A-Go-Go Potassium Channels/physiology , Guinea Pigs , Heart Rate/drug effects , Humans , In Vitro Techniques , Male , Methylphenidate/adverse effects , Papillary Muscles/drug effects , Papillary Muscles/physiology , Telemetry
5.
J Pharmacol Sci ; 99(5): 423-37, 2005.
Article in English | MEDLINE | ID: mdl-16493184

ABSTRACT

To construct a non-clinical database for drug-induced QT interval prolongation, the electrophysiological effects of 11 positive and 10 negative compounds on action potentials (AP) in guinea-pig papillary muscles were investigated in a multi-site study according to a standard protocol. Compounds with a selective inhibitory effect on the rapidly activated delayed rectifier potassium current (IKr) prolonged action potential duration at 90% repolarization (APD90) in a concentration-dependent manner, those showing Ca2+ current (ICa) inhibition shortened APD30, and those showing Na+ current (INa) inhibition decreased action potential amplitude (APA) and Vmax. Some of the mixed ion-channel blockers showed a bell-shaped concentration-response curve for APD90, probably due to their blockade of INa and/or ICa, sometimes leading to a false-negative result in the assay. In contrast, all positive compounds except for terfenadine and all negative compounds with IKr-blocking activity prolonged APD30-90 regardless of their INa- and/or ICa-blocking activities, suggesting that APD30-90 is a useful parameter for evaluating the IKr-blocking activity of test compounds. Furthermore, the assay is highly informative regarding the modulation of cardiac ion channels by test compounds. Therefore, when APD90 and APD30-90 are both measured, the action potential assay can be considered a useful method for assessing the risk of QT interval prolongation in humans in non-clinical safety pharmacology studies.


Subject(s)
Action Potentials/drug effects , Biological Assay , Long QT Syndrome/chemically induced , Papillary Muscles/drug effects , Animals , Databases, Factual , Guinea Pigs , In Vitro Techniques , Male , Papillary Muscles/physiology , Pharmaceutical Preparations
6.
J Pharmacol Sci ; 99(5): 449-57, 2005.
Article in English | MEDLINE | ID: mdl-16493186

ABSTRACT

Certain compounds that prolong QT interval in humans have little or no effect on action-potential (AP) duration used traditionally, but they inhibit rapidly-activated-delayed-rectifier potassium currents (IKr) and/or human ether-a-go-go-related gene (hERG) currents. In this study using isolated guinea-pig papillary muscles, we investigated whether new parameters in AP assays can detect the inhibitory effects of various compounds on IKr and/or hERG currents with high sensitivity. The difference in AP duration between 60% and 30% repolarization, 90% and 60% repolarization, and 90% and 30% repolarization (APD30-60, APD60-90, and APD30-90, respectively) were calculated as the new parameters. All the 15 IKr and/or hERG current inhibitors that have been reported (9 compounds) or not reported (6 compounds) to inhibit calcium currents prolonged APD30-60, APD60-90, and/or APD30-90; and 8 of the 15 inhibitors prolonged APD30-60, APD60-90, and/or APD30-90 more potently than APD90. The APD30-60, APD60-90, and APD30-90 measurements revealed no difference in sensitivity when evaluating the effects of the IKr and/or hERG current inhibitors on the three parameters. On the other hand, compounds with little or no effect on hERG currents had no effect on APD30-60, APD60-90, or APD30-90. Therefore, it is concluded that in AP assays using isolated guinea-pig papillary muscles, APD30-60, APD60-90, and APD30-90 are useful indexes for evaluating the inhibitory effects of compounds including mixed ion-channel blockers on IKr and/or hERG currents.


Subject(s)
Action Potentials/drug effects , Calcium Channel Blockers/pharmacology , Long QT Syndrome/chemically induced , Papillary Muscles/drug effects , Potassium Channel Blockers/pharmacology , Animals , Databases, Factual , Delayed Rectifier Potassium Channels/antagonists & inhibitors , Delayed Rectifier Potassium Channels/physiology , Ether-A-Go-Go Potassium Channels/antagonists & inhibitors , Ether-A-Go-Go Potassium Channels/physiology , Guinea Pigs , In Vitro Techniques , Male , Papillary Muscles/physiology
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