The rate of infusion of repolarisation delaying agents may influence the dispersion of repolarisation and play a decisive role in the initiation of torsade de pointes.

To improve proarrhythmic predictability of preclinical models, we assessed whether human ventricular-like embryonic stem cell-derived cardiomyocytes (hESC-CMs) can be selected following a standardized protocol. Also, we quantified their arrhythmogenic response and compared this to a contemporary used rabbit Purkinje fiber (PF) model. Multiple transmembrane action potentials (AP) were recorded from 164 hESC-CM clusters (9 different batches), and 12 isolated PFs from New Zealand White rabbits. AP duration (APD), early afterdepolarizations (EADs), triangulation (T), and short-term variability of repolarization (STV) were determined on application of the I(Kr) blocker E-4031 (0.03/0.1/0.3/1 muM). Isoproterenol (0.1 muM) was used to assess adrenergic response. To validate the phenotype, RNA isolated from atrial- and ventricular-like clusters (n=8) was analyzed using low-density Taqman arrays. Based on initial experiments, slow beating rate (<50 bpm) and long APD (>200 ms) were used to select 31 ventricular-like clusters. E-4031 (1 muM) prolonged APD (31/31) and induced EADs only in clusters with APD90>300 ms (11/16). EADs were associated with increased T (1.6+/-0.2 vs 2.0+/-0.3) and STV (2.7+/-1.5 vs 6.9+/-1.9). Rabbit PF reacted in a similar way with regards to EADs (5/12), increased T (1.3+/-0.1 vs 1.9+/-0.4), and STV (1.2+/-0.9 vs 7.1+/-5.6). According to ROC values, hESC-CMs (STV 0.91) could predict EADs at least equivalent to PF (STV 0.69). Isoproterenol shortened APD and completely suppressed EADs. Gene expression analysis revealed that HCN1/2, KCNA5, and GJA5 were higher in atrial/nodal-like cells, whereas KCNJ2 and SCN1B were higher in ventricular-like cells (P<0.05). Selection of hESC-CM clusters with a ventricular-like phenotype can be standardized. The proarrhythmic results are qualitatively and quantitatively comparable between hESC-CMs and rabbit PF. Our results indicate that additional validation of this new safety pharmacology model is warranted.

Atrial fibrillation (AF) is the most common tachyarrhythmia in the adult population and is a major cause of morbidity and mortality. AF can be terminated and sinus rhythm restored by prolonging the action potential duration (APD) and the refractory period. Unfortunately, antiarrhythmic agents that prolong the APD and increase the refractory period via selective inhibition of the rapid delayed rectifier potassium current (IK r ), i.e. class III antiarrhythmic drugs, are associated with an increased risk of the ventricular tachycardia Torsades de Pointes. AZD7009 is an antiarrhythmic agent with predominant actions on atrial electrophysiology that shows high antiarrhythmic efficacy and low proarrhythmic potential in animals and man. The aim of the current studies was to characterize the effect of AZD7009 on cardiac ion currents and APD in order to provide a mechanistic explanation for its predominant atrial effects and low proarrhythmic potential.The human cardiac ion channels hERG (IK r ), Kv1.5 (IK ur ), Kv4.3/KChIP2.2 (I to ), KvLQT1/minK (IK s ), Kir3.1/Kir3.4 (IK ACh ) and Nav1.5 (INa) were expressed in mammalian cells. Whole-cell currents were inhibited by AZD7009 with the following IC 50 values: hERG 0.6 μM, Nav1.5 8 μM, Kv4.3/KChIP2.2 24 μM, Kv1.5 27 μM, Kir3.1/Kir3.4 166 μM and KvLQT1/minK 193 μM. Whole-cell sodium and calcium currents were recorded in isolated rabbit atrial and ventricular myocytes using amphotericin B perforated patch. The late sodium current in rabbit atrial and ventricular myocytes was inhibited by AZD7009 in a concentration dependent way, with approximately 50% inhibition at 10 μM AZD7009. The L-type Ca 2+ current (ICa L ) in rabbit ventricular myocytes was inhibited with an IC 50 of 90 μM. Transmembrane action potentials were recorded in tissue pieces from rabbit atrium, ventricle and Purkinje fibre in control, during exposure to the selective IK r blocker E-4031 and to E-4031 in combination with AZD7009. In Purkinje fibres, but not in ventricular tissue, AZD7009 attenuated the E-4031-induced APD prolongation. In contrast, in atrial cells, AZD7009 further prolonged the APD. In addition, AZD7009 was able to suppress early afterdepolarisations (EADs) induced by E-4031 in Purkinje fibre preparations.In conclusion, AZD7009 delays repolarisation and increases refractoriness in atrial tissue through synergistic inhibition of IK r , I to , IK ur and INa, a mixed ion channel blockade that may underlie its high antiarrhythmic efficacy. Inhibition of the late sodium current, counteracting excessive APD prolongation and EADs in susceptible cells (midmyocardial and Purkinje cells), may explain the low proarrhythmic potential of AZD7009.

AZD1305 is a novel antiarrhythmic agent under clinical evaluation for management of atrial fibrillation. This study assessed its ion channel-blocking potency by the whole cell patch-clamp technique in vitro and its proarrhythmic liability in anesthetized methoxamine-sensitized rabbits in comparison with dofetilide. AZD1305 predominantly blocked the hERG, the L-type calcium and the hNav1.5 currents in a concentration-dependent manner. In vivo AZD1305 increased the QT interval (from 145 +/- 8 to 196 +/- 18 ms, P < 0.01) without inducing ventricular extrasystoles or torsades de pointes (TdP). In contrast, dofetilide prolonged the QT interval from 161 +/- 3 to 256 +/- 15 ms (P < 0.001) and caused TdP in 12/17 rabbits (P < 0.01 vs. AZD1305). During AZD1305 and dofetilide infusion, the QTend-peak interval maximally increased by 14 +/- 4 and 30 +/- 6 ms (P < 0.05 vs. AZD1305) and the beat-by-beat QT interval variability (quantified as the short-term variability, STV) changed from 2 +/- 0.8 to 2 +/- 0.3 ms (NS) and from 2 +/- 0.2 to 12 +/- 1.1 ms (P < 0.001), respectively. Following dofetilide-induced TdP, 6 rabbits each were injected with saline or AZD1305. In contrast to saline, AZD1305 abbreviated the QT interval (from 275 +/- 25 to 216 +/- 9 ms, P < 0.05), reduced the STV to 1 +/- 0.1 ms (P < 0.001) and suppressed TdP in all 6 rabbits (P < 0.01 vs. saline). In conclusion, AZD1305 can be characterised as a combined ion channel blocker that delays repolarization without increasing beat-by-beat variability of repolarization (BVR) or inducing TdP whereas selective IKr blockade by dofetilide prolongs the QT interval and eventually increases BVR resulting in TdP.

By blocking INalate, AZD1305 attenuates excessive APD90 prolongation and repolarization instability following sudden slowing of rhythm implying a low proarrhythmic potential for AZD1305.