Analysis of torsadogenic and pharmacokinetic profile of E-4031 in dogs bridging the gap of information between in vitro proarrhythmia assay and clinical observation in human subjects

We analyzed torsadogenic and pharmacokinetic profile of E-4031 using chronic atrioventricular block dogs. E-4031 in intravenous doses of 0.03, 0.1 and 0.3 mg/kg over 10 min prolonged QT/QTc, and increased short-term variability of QT in a dose-related manner (n = 4), resulting in onset of torsade de pointes in 1 animal after the middle dose and 4 animals after the high dose, while it attained peak plasma concentrations of 16.5, 60.5 and 182.5 ng/mL at 10 min after their start of administration, respectively (n = 2). These results bridge the gap of information between in vitro proarrhythmia assay and clinical observation in human subjects. Keywords: E-4031, Torsade de pointes, CiP

Application of human induced pluripotent stem cell-derived cardiomyocytes sheets with microelectrode array system to estimate antiarrhythmic properties of multi-ion channel blockers

We examined electrophysiological indices of human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs) sheets in order to quantitatively estimate Na+, K+ and Ca2+ channel blocking actions of bepridil and amiodarone using microelectrode array system in comparison with that of E-4031. We analyzed the field potential duration, effective refractory period, current threshold and conduction property using a programmed electrical stimulation protocol to obtain the post repolarization refractoriness and coefficient a of the relationship between the pacing cycle length and field potential duration. Electropharmacological profile of each drug was successfully characterized; namely, 1) the changes in the current threshold and conduction property provided basic information of Na+ channel blocking kinetics, 2) the relationship between pacing cycle length and field potential duration reflected drug-induced inhibition of human ether-à-go-go-related gene (hERG) K+ channel, 3) the post repolarization refractoriness indicated the relative contribution of these drugs to Na+ and K+ channel blockade, and 4) L-type Ca2+ channel blocking action was more obvious in the field potential waveform of the hiPSC-CMs sheets than that expected in the electrocardiogram in humans. Thus, this information may help to better utilize the hiPSC-CMs sheets for grasping the properties and net effects of drug-induced Na+, Ca2+ and K+ channel blockade. Keywords: Human induced pluripotent stem cell-derived cardiomyocytes, Post repolarization refractoriness, Multichannel blocker, Antiarrhythmic propert

Electropharmacological effects of intracellular Ca2+ handling modulator caldaret on the heart assessed in the halothane-anesthetized dogs

We analyzed how the enhancement of net sarcoplasmic reticulum (SR) Ca2+ uptake may affect cardiac electrophysiological properties in vivo by using caldaret which can decrease SR diastolic Ca2+ leak, enhance SR Ca2+ reuptake and inhibit reverse-mode Na+/Ca2+ exchanger. Caldaret in doses of 0.5, 5 and 50 μg/kg was intravenously administered over 10 min to the halothane-anesthetized beagle dogs (n = 5), attaining pharmacologically active plasma concentration. The low and middle doses of caldaret increased the ventricular contraction, which could be explained by its on-target pharmacological activities. The high dose enhanced the sinus automaticity followed by its suppression in addition to the increase of the total peripheral resistance, which may be unfavorable for treating diastolic heart failure. The low and middle doses enhanced the atrioventricular conduction, which may have some potential for predisposing the atria to the onset of atrial fibrillation via an induction of mitral and/or tricuspid regurgitation. The middle and high doses of caldaret prolonged the ventricular effective refractory period without altering the intraventricular conduction or repolarization period, which may prevent the onset of ventricular arrhythmias. Thus, modulation of intracellular Ca2+ handling by caldaret can induce not only inotropic effect, but also various electrophysiological actions on the in situ heart. Keywords: Caldaret, Ryanodine receptor, Sarco/endoplasmic reticulum Ca2+-ATPase, Reverse-mode Na+/Ca2+ exchange

Development of correction formula for field potential duration of human induced pluripotent stem cell-derived cardiomyocytes sheets

Human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs) have been used in many studies to assess proarrhythmic risks of chemical compounds. In those studies, field potential durations (FPD) of hiPSC-CMs have been corrected by clinically used Fridericia's and/or Bazett's formulae, however, the rationale for the use of these formulae has not been well established. In the present study, we developed a correction formula for experiments using hiPSC-CMs. First, we analyzed the effect of beating rate on FPD in the hiPSC-CMs sheets with electrical stimuli and a HCN channel inhibitor zatebradine. Next, we examined the relationship between the electrophysiological properties and the expression levels of ion channel genes in the cell sheets. Zatebradine slowed the beating rate and allowed to analyze FPD changes at various pacing cycle lengths. Rate-dependent change in the repolarization period was smaller in the cell sheets than that reported on the human hearts, which can be partly explained by lower gene expression level of hKCNJ2 and hKCNE1. Thus, non-linear equation for correcting FPD in the cell sheet; FPDc = FPD/RR0.22 with RR given in second was obtained, which may make it feasible to assess net repolarization delay by various chemical compounds with a chronotropic action

Use of microminipigs for unveiling unknown mechanisms of azithromycin-induced cardiovascular death

Although azithromycin can suppress cardiac INa, IKr, IKs, ICa,L and IK1, its onset mechanisms for cardiovascular death have not been fully investigated. We examined electropharmacological effects of azithromycin in intravenous doses of 0.3, 3 and 30 mg/kg using microminipigs under the halothane anesthesia (n = 4), which provided plasma concentrations of 3.1, 11.2 and 120.4 μg/mL, respectively. The low dose did not alter any of the cardiohemodynamic or electrocardiographic variables. The middle dose significantly shortened QT interval for 10–20 min and QTc for 10–30 min. The high dose significantly decreased mean blood pressure for 5–60 min, prolonged QRS width at 20 min, but shortened QT interval for 15–20 min and QTc for 15–30 min (n = 3). Cardiohemodynamic collapse occurred in 1 animal after the start of the high dose infusion, which might be associated with the cardiovascular death in patients with vasomotor dysfunction. Prolongation of QRS width indicates that azithromycin may suppress ventricular INa in vivo, which may unmask latent type of Brugada electrocardiographic genotype. Meanwhile, abbreviation of the QTc might cause potentially lethal, short QT-related, cardiac arrhythmia syndrome. These findings with microminipigs suggest the possible entry point for analyzing the mechanisms of cardiovascular death clinically seen with this antibiotic. Keywords: Azithromycin, Microminipigs, Short QT syndrome, Brugada syndrome, Hypotensio

Analysis of proarrhythmic potential of an atypical antipsychotic drug paliperidone in the halothane-anesthetized dogs

Fatal cases with the use of atypical antipsychotic drug paliperidone have been reported; however, there was no clinical report describing paliperidone-induced torsade de pointes. In this study we assessed its electropharmacological effects together with its proarrhythmic potential in intravenous doses of 0.03, 0.3 and 3 mg/kg using the halothane-anesthetized dogs (n = 5), which could provide approximately 2, 20 and 200 times higher peak plasma drug concentrations than its therapeutic level, respectively. Paliperidone exerted potent vasodilator effect resulting in hypotension, which may be largely explained by its α1-adrenoceptor blocking action. In vivo electrophysiological results suggest that paliperidone may inhibit human ether-à-go-go-related gene K+ channel in a dose-related manner and modestly suppress Na+ channel in the in situ heart. The high dose of paliperidone may have some potential to induce early afterdepolarization that can trigger lethal ventricular arrhythmias, whereas the low and middle doses lack such proarrhythmic possibility, indicating that at least 20 times higher plasma concentration may be considered to be safe

Changes of electrocardiogram and hemodynamics in response to dipyridamole: In vivo comparative analyses using anesthetized beagle dogs and microminipigs

Microminipigs are expected as a novel animal model for cardiovascular pharmacological experiments. Since inherent vulnerability of coronary circulation of microminipigs has not been characterized, we performed dipyridamole-stress test to both microminipigs and beagle dogs, and compared the results. Dipyridamole in doses of 0.056 and 0.56 mg/kg were intravenously infused over 10 min (n = 4 for each animal). Dipyridamole decreased the systolic/diastolic blood pressures and double product in dogs as well as in microminipigs; but it did not significantly alter the heart rate or the global balance between the myocardial oxygen demand and supply in either animal. While organic coronary arterial stenosis was not detected in either animal, dogs have well-developed epicardial intracoronary networks unlike microminipigs. Like in humans, dipyridamole did not affect the ST segment of microminipigs, whereas it substantially depressed that in dogs. The results indicate the onset of subendocardial ischemia by dipyridamole in dogs may be partly associated with their well-developed native coronary collateral channels. Microminipigs would be more useful to evaluate the drugs which may affect the coronary circulation in the pre-clinical study than dogs. Keywords: Dipyridamole, Collateral arteries, Coronary steal, ST-segment depressio