Class Ia Antiarrhythmic Drugs Research Articles

Abstract 16851: Tricyclic Antidepressant Induced Wide Complex Tachycardia With Brugada Pattern

Case Description: A 52-year-old male with bipolar disorder and depression was brought to the ED after he was found confused following an episode of syncope. He reported taking 5 pills of his new dose of Nortriptyline 75mg instead of 5 pills of his previous dose of Nortriptyline 25mg. ECG from the field showed a wide complex tachycardia (WCT) with a rightward axis and a Brugada pattern in V1 and V2 at a rate of 120-130s. Upon arrival in the ED, he continued to have WCT, and was treated with Amiodarone, Magnesium Sulfate, and Sodium Bicarbonate. Following treatment, his heart rate improved and the QRS duration shortened with a gradual loss of Brugada pattern (Figure 1). Six hours from initial presentation, his QRS completely normalized. Transthoracic echocardiogram (TTE) showed normal LV function. His presentation was consistent with tricyclic antidepressant (TCA) overdose. Discussion: The cardiotoxic effects of TCAs are mediated through their behavior as class IA antiarrhythmic drugs. This results in lengthening of both depolarization and repolarization of the cardiac action potential resulting in a widened QRS complex and QT prolongation. Sodium channel blockade can precipitate a Brugada phenotype which resolves after Sodium Bicarbonate administration. Similar to class IA agents, TCAs also exhibit use dependence and in our patient, the Brugada pattern began to resolve soon after presentation, possibly due to slowing of the tachycardia. Recognition of TCA toxicity is important to allow for timely administration of bicarbonate which facilitates prompt reversal.

Impact of Short-Acting Disopyramide on Left Ventricular Mechanics Evaluated by Strain Analysis in Patients with Hypertrophic Obstructive Cardiomyopathy.

Disopyramide is a class Ia antiarrhythmic drug that has been used for the second-line treatment of symptomatic hypertrophic obstructive cardiomyopathy (HOCM). The aim of the study was to assess the impact of short-acting disopyramide in patients with hypertrophic obstructive cardiomyopathy (HOCM) using two-dimensional speckle-tracking echocardiography. This prospective study included patients with HOCM on chronic treatment with short-acting disopyramide. Two sequential comprehensive echocardiographic examinations were performed: after temporary disopyramide suspension and 2.5 h after disopyramide intake. 19 patients were included in the study. The effect of disopyramide on the left ventricle was not uniform. After the intake of disopyramide, the mean global strain peak was -17 ± 2% before disopyramide intake and -14 ± 2% after (p < 0.0001). There was a significant reduction in strain in the basal septal (p = 0.015), basal inferior (p = 0.019), basal posterior (p = 0.05), apical anterior (p = 0.0001), and apical lateral segments (p = 0.021). In all other segments, there was no significant change. Disopyramide also caused a significant accentuation of the base-apex strain gradients (p = 0.036). No change was noted in circumferential and left atrial strain. While the left ventricular ejection fraction and outflow gradients did not change, the significant reduction in global and segmental longitudinal strain demonstrated the acute negative inotropic effect of disopyramide on the myocardium in patients with HOCM. A strain analysis may be a useful tool to assess the negative inotropic effect of cardiovascular medication on the left ventricle in patients with HOCM.

Structural Basis for Pore Blockade of the Human Cardiac Sodium Channel Nav1.5 by the Antiarrhythmic Drug Quinidine**

AbstractNav1.5, the primary voltage‐gated Na+ (Nav) channel in heart, is a major target for class I antiarrhythmic agents. Here we present the cryo‐EM structure of full‐length human Nav1.5 bound to quinidine, a class Ia antiarrhythmic drug, at 3.3 Å resolution. Quinidine is positioned right beneath the selectivity filter in the pore domain and coordinated by residues from repeats I, III, and IV. Pore blockade by quinidine is achieved through both direct obstruction of the ion permeation path and induced rotation of an invariant Tyr residue that tightens the intracellular gate. Structural comparison with a truncated rat Nav1.5 in the presence of flecainide, a class Ic agent, reveals distinct binding poses for the two antiarrhythmics within the pore domain. Our work reported here, along with previous studies, reveals the molecular basis for the mechanism of action of class I antiarrhythmic drugs.

Structural Basis for Pore Blockade of the Human Cardiac Sodium Channel Nav 1.5 by the Antiarrhythmic Drug Quinidine*.

Nav 1.5, the primary voltage-gated Na+ (Nav ) channel in heart, is a major target for class I antiarrhythmic agents. Here we present the cryo-EM structure of full-length human Nav 1.5 bound to quinidine, a class Ia antiarrhythmic drug, at 3.3 Å resolution. Quinidine is positioned right beneath the selectivity filter in the pore domain and coordinated by residues from repeats I, III, and IV. Pore blockade by quinidine is achieved through both direct obstruction of the ion permeation path and induced rotation of an invariant Tyr residue that tightens the intracellular gate. Structural comparison with a truncated rat Nav 1.5 in the presence of flecainide, a class Ic agent, reveals distinct binding poses for the two antiarrhythmics within the pore domain. Our work reported here, along with previous studies, reveals the molecular basis for the mechanism of action of class I antiarrhythmic drugs.

Crystal Structures of Antiarrhythmic Drug Disopyramide and Its Salt with Phthalic Acid

Disopyramide (DPA) is as a class IA antiarrhythmic drug and its crystallization from cyclohexane at ambient condition yields lower melting form crystals which belong to the monoclinic centrosymmetric space group P21/n, having two molecules in an asymmetric unit. Crystal structure analysis of pure DPA revealed closely associated DPA molecules aggregates via amide–amide dimer synthon through the N–H∙∙∙O hydrogen bond whereas the second amide hydrogen N–H engaged in an intramolecular N–H∙∙∙N hydrogen bond with N-nitrogen of 2-pyridine moieties. Crystallization of DPA and phthalic acid (PA) in 1: 1 stoichiometric molar ratio from acetone at ambient condition yielded block shape crystals of 1:1 DPA_PA salt. Its X-ray single crystal structure revealed the formation of salt by transfer of acidic proton from one of the carboxylic acidic groups of PA to the tertiary amino group of chain moiety (N3-nitrogen atom) of DPA molecules. DPA_PA salt crystals belong to the monoclinic centrosymmetric space group P21/n, comprising one protonated DPA and one PA¯ anion (hydrogen phthalate counterion) in an asymmetric unit and linked by N–H∙∙∙O and C–H∙∙∙O hydrogen bonds. Pure DPA and DPA_PA salt were further characterized by differential calorimetric analysis, thermal gravimetric analysis, powder x-ray diffraction and infrared spectroscopy.

Long-term efficacy and safety of hydroquinidine in patients with Brugada syndrome

Abstract Background Brugada syndrome (BrS) is an inherited channelopathy with an increased risk of supraventricular, ventricular arrhythmias (VAs) and sudden cardiac death (SCD). Implantable cardioverter-defibrillator (ICD) is a cornerstone of SCD prevention, but it does not reduce the incidence of life-threatening VAs and it can carry substantial complications. Hydroquinidine (HQ) is a class IA antiarrhythmic drug used for electrical storms, to reduce ICD's appropriate discharges and as an alternative to ICD in children with BrS or in patients with a contraindication to ICD. Nevertheless, HQ's side effects may undermine treatment compliance. Purpose The aim of this study was to evaluate the efficacy and safety of HQ in reducing VAs (ventricular fibrillation, sustained and non-sustained ventricular tachycardia) inducibility at electrophysiology study (EPS) and atrial fibrillation/flutter (AF/AFL) or VAs recurrence in patients with BrS. Methods From the prospective Piedmont Brugada Registry, patients treated with HQ were selected and divided into three groups according to the indication for HQ initiation: index EPS positive for VAs induction (group 1), secondary prevention of AF/AFl (group 2), secondary prevention of VAs (group 3). In group 3 recurrence of VAs was monitored by implantable devices or by periodic 24-hour ECG Holter monitoring. In 5 patients HQ was started for reasons different from the above mentioned, so they were considered only for safety outcomes. Safety was assessed considering the occurrence of HQ side effects and their impact on treatment discontinuation. Results A total of 98 patients (79 males, 80,6%) were included. Median follow-up was 61 months (IQR 31–89 months). None of the baseline clinical characteristics was associated with arrhythmic recurrences. Among 46 patients in group 1 HQ was effective in reducing EPS inducibility in 91.9% of patients (p<0.0001); in group 2 (31 patients) HQ reduced palpitations [before HQ 83,8%, with HQ 27,6%, RRR 67.1%, NNT 1.8; p<0.0001] and no AF/AFL recurrence was recorded during follow-up (p<0.0001); in group 3 (17 patients; 70.6% with ICD/LR implanted) VAs recurrences were significantly reduced in patients with HQ (5.9% recurrence rate, p<0.0001). Overall, no cardiac arrest occurred during follow-up. At ECG, HQ determined a significant increase in QTc duration (V5-lead mean QTc duration pre-HQ 406 ms vs with HQ 428 ms; p=0.001). Overall, 28.6% of patients presented HQ-related side effects, mainly due to gastrointestinal intolerance (18.3%). Treatment discontinuation rate was 25% but only about half of these patients discontinued HQ for adverse events (29.2% for GI intolerance, 16.7% for drug-induced QTc prolongation, 8.3% for elevated liver enzymes; 45.8% self-discontinuation). Conclusions In patients with BrS, HQ was effective in reducing VAs inducibility at EPS, AF/AFL and VAs recurrences; moreover, it was effective in reducing symptoms. Overall, HQ proved to be safe and well-tolerated. Funding Acknowledgement Type of funding source: None

Pathomechanisms and Clinical Implications of Myasthenic Syndromes Exacerbated and Induced by Medical Treatments.

Myasthenic syndromes are typically characterized by muscle weakness and increased fatigability due to an impaired transmission at the neuromuscular junction (NMJ). Most cases are caused by acquired autoimmune conditions such as myasthenia gravis (MG), typically with antibodies against the acetylcholine receptor (AChR). Different drugs are among the major factors that may complicate pre-existing autoimmune myasthenic conditions by further impairing transmission at the NMJ. Some clinical observations are substantiated by experimental data, indicating that presynaptic, postsynaptic or more complex pathomechanisms at the NMJ may be involved, depending on the individual compound. Most robust data exist for the risks associated with some antibiotics (e.g., aminoglycosides, ketolides, fluoroquinolones) and cardiovascular medications (e.g., class Ia antiarrhythmics, beta blockers). Apart from primarily autoimmune-mediated disorders of the NMJ, de novo myasthenic manifestations may also be triggered by medical treatments that induce an autoimmune reaction. Most notably, there is growing evidence that the immune checkpoint inhibitors (ICI), a modern class of drugs to treat various malignancies, represent a relevant risk factor to develop severe and progressive medication-induced myasthenia via an immune-mediated mechanism. From a clinical perspective, it is of utmost importance for the treating physicians to be aware of such adverse treatment effects and their consequences. In this article, we aim to summarize existing evidence regarding the key molecular and immunological mechanisms as well as the clinical implications of medication-aggravated and medication-induced myasthenic syndromes.

Action potential clamp characterization of the S631A hERG mutation associated with short QT syndrome.

The hERG potassium channel is critical to normal repolarization of cardiac action potentials (APs) and loss‐ and gain‐of‐function hERG mutations are associated, respectively, with long and short QT syndromes, pathological conditions that can lead to arrhythmias and sudden death. hERG current (I h ERG) exhibits uniquely fast inactivation involving conformational changes to the channel pore. The S631A hERG pore mutation was originally engineered to interrogate hERG channel inactivation, but has very recently been found in a family with short QT syndrome (SQTS). Accordingly, this study characterized the effects of the S631A mutation on I h ERG profile during ventricular, atrial, and Purkinje fiber (PF) AP waveforms, using patch clamp recording from hERG expressing HEK 293 cells at 37°C. Under conventional voltage clamp, the current–voltage (I–V) relation for I h ERG exhibited a marked right‐ward shift in the region of negative slope at positive membrane potentials. Under ventricular AP clamp, the S631A mutation resulted in augmented I h ERG, which also peaked much earlier during the AP plateau than did wild‐type (WT) I h ERG. Instantaneous I–V relations showed a marked positive shift in peak repolarizing current during the ventricular AP in the S631A setting, while the instantaneous conductance‐voltage relation showed an earlier and more sustained rise in S631A compared to WT I h ERG conductance during ventricular repolarization. Experiments with atrial and PF APs in each case also showed augmented and positively shifted I h ERG in the S631A setting, indicating that the S631A mutation is likely to accelerate repolarization in all three cardiac regions. Ventricular AP clamp experiments showed retained effectiveness of the class Ia antiarrhythmic drug quinidine (1 μmol/L) against S631A I h ERG. Quinidine is thus likely to be effective in reducing excessively fast repolarization in SQTS resulting from the S631A hERG mutation.

Ajmaline blocks INa and IKr without eliciting differences between Brugada syndrome patient and control human pluripotent stem cell-derived cardiac clusters

The class Ia anti-arrhythmic drug ajmaline is used clinically to unmask latent type I ECG in Brugada syndrome (BrS) patients, although its mode of action is poorly characterised.Our aims were to identify ajmaline's mode of action in human induced pluripotent stem cell (hiPSC)-derived cardiomyocytes (CMs), and establish a simple BrS hiPSC platform to test whether differences in ajmaline response could be determined between BrS patients and controls.Control hiPSCs were differentiated into spontaneously contracting cardiac clusters. It was found using multi electrode array (MEA) that ajmaline treatment significantly lengthened cluster activation-recovery interval. Patch clamping of single CMs isolated from clusters revealed that ajmaline can block both INa and IKr.Following generation of hiPSC lines from BrS patients (absent of pathogenic SCN5A sodium channel mutations), analysis of hiPSC-CMs from patients and controls revealed that differentiation and action potential parameters were similar. Comparison of cardiac clusters by MEA showed that ajmaline lengthened activation-recovery interval consistently across all lines.We conclude that ajmaline can block both depolarisation and repolarisation of hiPSC-CMs at the cellular level, but that a more refined integrated tissue model may be necessary to elicit differences in its effect between BrS patients and controls.

Dual Mechanism for Inhibition of Inwardly Rectifying Kir2.x Channels by Quinidine Involving Direct Pore Block and PIP2-interference.

Class IA antiarrhythmic drug quinidine was one of the first clinically used compounds to terminate atrial fibrillation and acts as multichannel inhibitor with well-documented inhibitory effects on several cardiac potassium channels. In the mammalian heart, heteromeric assembly of Kir2.1-2.3 channels underlies IK1 current. Although a low-affinity block of quinidine on Kir2.1 has already been described, a comparative analysis of effects on other Kir2.x channels has not been performed to date. Therefore, we analyzed the effects of quinidine on wild-type and mutant Kir2.x channels in the Xenopus oocyte expression system. Quinidine exerted differential inhibitory effects on Kir2.x channels with the highest affinity toward Kir2.3 subunits. Onset of block was slow and solely reversible in Kir2.2 subunits. Quinidine inhibited Kir2.x currents in a voltage-independent manner. By means of comparative Ala-scanning mutagenesis, we further found that residues E224, F254, D259, and E299 are essential for quinidine block in Kir2.1 subunits. Analogously, quinidine mediated Kir2.3 inhibition by binding corresponding residues E216, D247, D251, and E291. In contrast, Kir2.2 current block merely involved corresponding residue D260. Using channel mutants with altered (phosphatidylinositol 4,5-bisphosphate PIP2) affinities, we were able to demonstrate that high PIP2 affinities (i.e., Kir2.3 I214L) correlate with low quinidine sensitivity. Inversely, mutant channels interacting only weakly with PIP2 (i.e., Kir2.1 K182Q, and L221I) are prone to a higher inhibitory effect. Thus, we conclude that inhibition of Kir2.x channels by quinidine is mediated by joint modes of action involving direct cytoplasmic pore block and an impaired channel stabilization via interference with PIP2.

Efficacy of Intravenous and Oral Sotalol in Pharmacologic Conversion of Atrial Fibrillation: A Systematic Review and Meta-Analysis

Objectives: The role of sotalol is well established for the maintenance of sinus rhythm after successful conversion of atrial fibrillation (AF). However, its role in pharmacologic conversion of AF is poorly defined. The purpose of this study is to compare the efficacy of sotalol to that of other antiarrhythmic agents for AF conversion. Methods: Standard methods of meta-analysis were employed. Full-text publications of clinical trials in English that compared the efficacy of sotalol to that of other antiarrhythmics or placebo/no treatment were eligible for inclusion. Results: A systematic review revealed 10 eligible publications. Sotalol was superior to placebo and/or no antiarrhythmic therapy in AF conversion, with a relative success of 24 (95% CI 4.7-119, p < 0.001). Sotalol was not significantly different from class IA antiarrhythmic drugs. Similarly, sotalol was not different from class IC antiarrhythmic drugs or amiodarone in terms of conversion efficacy. In one study, sotalol was less effective than high-dose ibutilide (2 mg), with a relative success of 0.248 (95% CI 0.128-0.481, p < 0.001). Ibutilide caused more proarrhythmia. Conclusions: Sotalol is as effective as class IA and class IC antiarrhythmic agents, and it is also as effective as amiodarone for pharmacologic conversion of AF. Only ibutilide at a high dose showed a greater conversion rate of AF.

Influence of Gender on the Tolerability, Safety, and Efficacy of Quinidine Used for Treatment of Supraventricular and Ventricular Arrhythmias

Quinidine, a class IA antiarrhythmic drug (AAD), has been used for the treatment of arrhythmias since the early 1900s. Use has decreased recently because of the availability of newer AADs and concerns about side effects and safety. Quinidine can cause QT prolongation, and women have longer QT intervals and are more susceptible to torsades de pointes (TdP) than are men. We sought to evaluate the influence of gender on quinidine tolerability, safety, and efficacy. We performed retrospective analyses of patients at our institution prescribed quinidine as an AAD between 2000 and 2012. Time to quinidine discontinuation and arrhythmia recurrence were evaluated using Cox proportional hazards models. In 179 patients, 23.5% were women and median age was 65.8 years. Quinidine indication was supraventricular arrhythmias in 68.7% and ventricular arrhythmias in 27.9% of patients. At 3 years after quinidine initiation, Kaplan-Meier probability of quinidine discontinuation was 65.7% for men and 82.4% for women (p = 0.015). Women were more likely than men to discontinue quinidine for QT prolongation (14.3 vs 4.4%, p = 0.036) and TdP (4.8 vs 0%, p = 0.054). After multivariate adjustment, female gender remained independently associated with quinidine discontinuation (adjusted hazard ratio 1.97, p = 0.014). Gender had no influence on arrhythmia recurrence: 1 year after quinidine initiation, Kaplan-Meier probability of freedom from recurrent arrhythmia was 62.4% in men and 57.9% in women (p = 0.33). Quinidine is highly effective in both genders. However, women are more likely than men to experience QT prolongation and TdP on quinidine and are more likely to discontinue quinidine independent of these side effects.

Impact of chronic use of cibenzoline on left ventricular pressure gradient and left ventricular remodeling in patients with hypertrophic obstructive cardiomyopathy

BackgroundCibenzoline, a class Ia antiarrhythmic drug, is useful for reducing the left ventricular pressure gradient (LVPG) in patients with hypertrophic obstructive cardiomyopathy (HOCM). However, chronic effects of cibenzoline on LVPG and left ventricular (LV) remodeling are unknown. MethodsForty-one patients with HOCM participated in this study. Echocardiographic, electrocardiographic, and brain natriuretic peptide (BNP) data collected before and after cibenzoline treatment were compared. From the relation between LVPG and plasma concentration of cibenzoline, an efficacious plasma concentration of cibenzoline was estimated. ResultsThe mean follow-up period was 74.2±47.1 months. The LVPG decreased from 104.8±62.6mmHg to 27.6±30.5mmHg (p<0.0001). The LV end-diastolic dimension increased from 42.8±5.8mm to 46.2±5.4mm (p<0.0001), but neither LV end-systolic dimension nor LV fractional shortening changed significantly. The left atrial dimension decreased from 40.0±4.7mm to 36.2±5.1mm (p<0.0001). The E-wave velocity/A-wave velocity ratio increased, early diastolic annular velocity (Ea) increased, and E/Ea ratio decreased. The interventricular septal wall thickness, LV posterior wall thickness, the Sokolow-Lyon index, and the depth of negative T wave decreased. The heart rate-corrected QT interval was shortened. Plasma BNP level decreased from 418.8±423.7pg/ml to 213.7±154.1pg/ml (p<0.02). The safe and efficacious plasma concentration of cibenzoline was between 300ng/mL and 1500ng/mL. ConclusionsLong-term treatment with cibenzoline attenuated LVPG, improved LV diastolic dysfunction, and induced LV hypertrophy regression in patients with HOCM without causing serious complications.

Inverstigating Stereoselectivity of Pharmacological Inhibition of Herg Channels

The potential for stereoselectivity of hERG K+ channel inhibition by drugs that exist as racemic mixtures is recognised; stereoselectivity has been established for some compounds (reviewed in (1)). Both the class Ia antiarrhythmic drug disopyramide (DISO) and the serotonin-selective reuptake inhibitor citalopram exist as racemic mixtures. Both have been associated with cases of drug-induced QT prolongation, with its attendant risk of torsades de pointes (TdP). Prior work has established that both compounds inhibit hERG current (IhERG) significantly at low micromolar concentrations (eg 2,3). In this study, we have investigated the potential for stereoselective hERG current (IhERG) inhibition by these drugs.Whole-cell patch-clamp measurements of IhERG were made at 37°C from hERG-expressing HEK 293 cells. All observations reported here are from at least 5 replicates. For citalopram, action potential voltage-clamp experiments were performed on racemic citalopram and the S(+) enantiomer (escitalopram). Application of 3μM citalopram inhibited maximal IhERG during the repolarisation phase of the AP by 69.6±3.6%, whilst escitalopram blocked the current by 63.4±7.1% (p>0.05, student t test). On the other hand, with a conventional screening protocol (S+) DISO was found to block IhERG ∼3 -fold more potently than did (R-) DISO (IC50 values of 3.9±0.6 μM and 12.9±1.4 μM respectively with normal (4mM) [K+]e). With high (94 mM) [K+]e, the IC50 values for (S+) and (R-) DISO were 16.9 ±3.0 μM and 61.1±7.4 μM respectively.Our data provide evidence that disopyramide exhibits stereoselective inhibition of IhERG. Our initial findings with citalopram are suggestive of a lack of notable stereoselectivity for that compound, though this needs further investigation.Funded by the British Heart Foundation(1) Grilo LS, et al, Front Pharmacol, 2010;1:137.(2) Ashok AP,et al, BBSRC, 2001, 280:1243-1250.(3) Witchel HJ, et al, FEBS Lett 2002;512:59-66.

Atrial Fibrillation Through the Years

Information about a real patient is presented in stages (boldface type) to an expert clinician (Dr Valentin Fuster), who responds to the information, sharing his reasoning with the reader (regular type). A discussion by the authors follows. A 61-year-old man presents with 2 weeks of exertional dyspnea. Pertinent medical history includes hypertension, nephrolithiasis, and internal hemorrhoids. He takes no medications and has no known drug allergies. His father died after a myocardial infarction at 57 years of age. He formerly smoked 1 pack of cigarettes daily for 15 years but ceased tobacco use 10 years before presentation. He ingests 2 glasses of alcohol weekly and denies illicit drug use. His caffeine intake is limited. He is an architect and is married, with healthy children. On physical examination, his temperature is 98.0°F, blood pressure is 130/85 mm Hg bilaterally, pulse is irregular at 130 beats per minute, and respiratory rate is 18 breaths per minute with an oxygen saturation of 97% while breathing room air. He is a slender white man in no distress. His jugular venous pressure is elevated at 14 cm H 2 O. There is no thyromegaly, and the carotid upstrokes are brisk, without bruits. Cardiovascular examination reveals a rapid and irregular heart rhythm with variation in the intensity of the first heart sound. The point of maximal impulse is not displaced. The remainder of the chest and abdominal examination is within normal limits. The extremities are warm and show mild pitting edema. Laboratory testing is significant for normal renal function and electrolytes, but a hemogram reveals a mild thrombocytopenia of 90 000 platelets/μL. ECG demonstrates atrial fibrillation (AF) with an average ventricular rate of 123 bpm ( Figure 1 ). Figure 1. The 12-lead ECG showing atrial fibrillation with a rapid ventricular rate. Dr Valentin Fuster : This is a …

Contribution of Local Anesthetic Binding Site Residues F1760 and Y1767 to Block of the Cardiac Na+ Channel, hNaV1.5, by Ranolazine

Amino acid residues F1760 and Y1767 in S6/Domain IV of voltage gated Na+ channels are part of the local anesthetic binding site in the inner mouth of the pore. Mutagenesis experiments have shown that use-dependent block (UDB) of the cardiac hNaV1.5 Na+ channel by the Class Ia anti-arrhythmic drug lidocaine is significantly reduced by alanine substitutions at these two sites. The antianginal drug ranolazine, which has structural similarity to lidocaine, was shown previously to block the cardiac late sodium current (INa) and to block peak and late INa in a use-dependent manner (Rajamani et al., 2009). As with lidocaine, UDB of peak INa by ranolazine was reduced in the F1760A mutant channel (Fredj et al., 2006; Wang et al., 2008). We used the F1760A and Y1767A mutants to investigate further the block by ranolazine of the closed, inactivated, and open states of the channel. To measure affinity to the open state, we used an inactivation-deficient hNaV1.5 channel (L409C/A410W, Edrich et al., 2005) in the presence and absence of the F1760A or Y1767A mutations. We found that both mutations (F1760A and Y1767A) significantly decreased UDB of peak INa, block of the open state, and block of late INa by ranolazine compared to WT channels (14-24 fold shift of the value of IC50 for ranolazine). The effect of the mutations on block by ranolazine of the resting and inactivated states was modest (1.3-1.9 fold shift of the value of IC50 for ranolazine). The results indicate that both the F1760A and Y1767A mutations reduce the effect of ranolazine to cause UDB and open state block of hNaV1.5.

A novel method of selecting human embryonic stem cell-derived cardiomyocyte clusters for assessment of potential to influence QT interval

Physiologically relevant assessment of delayed repolarization is necessary in drug development. In our preliminary experiments on the evaluation using a multielectrode recording system, we had found that the responsiveness of field potential duration (FPD), as QT-like intervals, to hERG channel blockers differed greatly from non-responders to excessive responders in human embryonic stem cell-derived cardiomyocyte clusters. Thus, we report a novel method of selecting clusters suitable for evaluating compounds for the assessment. Clusters were treated with cisapride, a hERG channel blocker, at 100nM, and selected with criteria of 5–20% of corrected FPD (FPDc) prolongation. Then, selected clusters were treated with reference compounds. FPDc was prolonged by blockade of the hERG channel (E-4031 and dl-sotalol) and KvLQT1 channel (chromanol 293B and HMR1556), and by activation of the sodium channel (veratridine) and calcium channel (Bay K8644). FPDc was shortened by calcium channel blockage (verapamil, nifedipine and diltiazem) and by KATP channel activation (pinacidil). Class Ia antiarrhythmic drugs, quinidine and disopyramide, prolonged FPDc. Selected clusters are appropriate for assessing the effects of compounds on ion channels affecting QT intervals. This is the first report of the establishment of an assessment system of potential to influence QT interval, using pharmacologically selected clusters.

Molecular determinants of hERG potassium channel inhibition by disopyramide

The Class Ia antiarrhythmic drug disopyramide (DISO) causes QT interval prolongation that is potentially dangerous in acquired Long QT Syndrome but beneficial in short QT syndrome, through inhibition of the hERG-encoded channels responsible for rapid delayed rectifier K+ current (IKr). In this study, alanine mutants of hERG S6 and pore helix residues and MthK-based homology modelling and ligand docking were used to investigate molecular determinants of DISO binding to hERG. Whole-cell hERG current (IhERG) recordings were made at 37°C from HEK-293 cells expressing WT or mutant hERG channels. WT outward IhERG tails were inhibited with an IC50 of 7.3μM, whilst inward IhERG tails in a high [K+]e of 94mM were blocked with an IC50 of 25.7μM. The IC50 for the Y652A mutation was ~55-fold that of WT IhERG; this mutation also abolished a leftward shift in voltage-dependent IhERG activation present for WT hERG. The IC50 for F656A IhERG was ~51 fold its corresponding WT control. In contrast to previously studied methanesulphonanilide hERG inhibitors, neither the G648A S6 nor the T623A and S624A pore helical mutations modified DISO IC50. Computational docking with the hERG model showed that DISO did not exhibit a single unique binding pose; instead several low energy binding poses at the lower end of the pore cavity favoured interactions with Y652 and F656. In the WT hERG model DISO did not interact directly with residues at the base of the pore helix, consistent with the minimal effect of mutation of these residues on drug block.

Molecular Determinants of HERG Potassium Channel Inhibition by Disopyramide

The class Ia antiarrhythmic drug disopyramide (DISO) delays cardiac ventricular repolarization through inhibition of the hERG-encoded, pore-forming subunit of the rapid delayed rectifier K+ current (IKr). In contrast with high-affinity hERG blockers DISO's inhibitory action on hERG is less dependent upon channel inactivation, raising the possibility that DISO binding to hERG may in some way be distinct [1,2]. Here the sensitivity to DISO of mutants of two aromatic residues (Y652A and F656A) in the S6 domain was investigated. Whole-cell patch clamp recordings were made at 37oC from HEK-293 cells stably expressing WT hERG or hERG mutants Y652A or F656A. WT hERG current (IhERG) tails at −40 mV following depolarization to +20 mV were inhibited with an IC50 of 7.2 ± 0.06 μM. The Y652A mutation greatly attenuated DISO inhibition of IhERG tail (IC50=402 ± 0.04 μM) and also abolished voltage-dependence of inhibition that was present for WT IhERG. F656A IhERG tails at −120 mV (recorded with a high [K+]e of 94 mM to facilitate current measurement) yielded a very high IC50of 1.85 ±0.07 mM. Preliminary docking analysis using a homology model built onto the KvAP crystal structure template suggests that in low-energy binding configurations the aromatic groups of DISO may reside at the lower end of the pore cavity. Collectively, these findings suggest a critical role for canonical binding residues Y652 and F656 in DISO inhibition of hERG and implicate Y652 in the voltage-dependence of hERG inhibition by DISO. This work was supported by the British Heart Foundation. 1. Paul AA et al (2001) BBRC, 280; 1243-50 2. McPate MJ et al (2008) Br J Pharmacol, 155, 957-66

Drug-Induced Arrhythmia

Prolongation of the QTc interval has been associated with proarrhythmia resulting from a potentially fatal form of polymorphic ventricular tachycardia called torsades de pointes (TdP). Genetic forms of the long-QT syndrome (LQTS) associated with high arrhythmic risk have been causally related to mutations in ion channels responsible for the cardiac action potential; genetic factors associated with milder degrees of QTc prolongation and arrhythmic risk have also been described. Acquired forms of QTc prolongation and proarrhythmia, particularly related to drug therapy, are frequently related to drug effects on the same ion channels involved in genetic forms of LQTS. As is true for genetic forms of LQTS, there is a wide spectrum of potential drug effects on the QTc interval ranging from trivial to potentially lethal. Drug-induced QTc prolongation is a complicated phenomenon related not just to the properties or dose of a particular drug but also to drug-drug interactions and a variety of patient factors, including age, gender, the presence and severity of underlying heart disease, and genetic predisposition. Additionally, drug effects on other ion channels, including blockade of sodium channels (causing prolongation of the QRS interval rather than the QTc interval), are an important cause of drug-induced proarrhythmia. The purpose of this review is to summarize the available data related to acquired forms of LQTS, with particular focus on drug-related LQTS and proarrhythmia. A brief summary is presented here of genetic LQTS because the relevant genes and proteins form the basis for an understanding of the causes of acquired LQTS, and in any given patient, the QTc is determined by genetic, substrate, and environmental factors. Emphasis is placed on practical strategies for clinicians, with the goal of minimizing the risk of dangerous proarrhythmia related to the use of both cardiac and noncardiac drugs with the potential for QTc prolongation. Genetic …