Block of the hERG channel by bupivacaine: Electrophysiological and modeling insights towards stereochemical optimization

The hERG voltage-gated potassium channel mediates the cardiac I(Kr) current, which is crucial for the duration of the cardiac action potential. Undesired block of the channel by certain drugs may prolong the QT interval and increase the risk of malignant ventricular arrhythmias. Although the molecular determinants of hERG block have been intensively studied, not much is known about its stereoselectivity. Levo-(S)-bupivacaine was the first drug reported to have a higher affinity to block hERG than its enantiomer. This study strives to understand the principles underlying the stereoselectivity of bupivacaine block with the help of mutagenesis analyses and molecular modeling simulations. Electrophysiological measurements of mutated hERG channels allowed for the identification of residues involved in bupivacaine binding and stereoselectivity. Docking and molecular mechanics simulations for both enantiomers of bupivacaine and terfenadine (a non-stereoselective blocker) were performed inside an open-state model of the hERG channel. The predicted binding modes enabled a clear depiction of ligand-protein interactions. Estimated binding affinities for both enantiomers were consistent with electrophysiological measurements. A similar computational procedure was applied to bupivacaine enantiomers towards two mutated hERG channels (Tyr652Ala and Phe656Ala). This study confirmed, at the molecular level, that bupivacaine stereoselectively binds the hERG channel. These results help to lay the foundation for structural guidelines to optimize the cardiotoxic profile of drug candidates in silico.

Patient with syncope and LQTS carrying a mutation in the PAS domain of the hERG1 channel

We report the case of a woman with syncope and persistently prolonged QTc interval. Screening of congenital long QT syndrome (LQTS) genes revealed that she was a heterozygous carrier of a novel KCNH2 mutation, c.G238C. Electrophysiological and biochemical characterizations unveiled the pathogenicity of this new mutation, displaying a 2-fold reduction in protein expression and current density due to a maturation/trafficking-deficient mechanism. The patient's phenotype can be fully explained by this observation. This study illustrates the importance of performing genetic analyses and mutation characterization when there is a suspicion of congenital LQTS. Identifying mutations in the PAS domain or other domains of the hERG1 channel and understanding their effect may provide more focused and mutation-specific risk assessment in this population.

Nedd4-2-dependent ubiquitylation and regulation of the cardiac potassium channel hERG1

The voltage-gated cardiac potassium channel hERG1 (human ether-à-gogo-related gene 1) plays a key role in the repolarization phase of the cardiac action potential (AP). Mutations in its gene, KCNH2, can lead to defects in the biosynthesis and maturation of the channel, resulting in congenital long QT syndrome (LQTS). To identify the molecular mechanisms regulating the density of hERG1 channels at the plasma membrane, we investigated channel ubiquitylation by ubiquitin ligase Nedd4-2, a post-translational regulatory mechanism previously linked to other ion channels. We found that whole-cell hERG1 currents recorded in HEK293 cells were decreased upon neural precursor cell expressed developmentally down-regulated 4-2 (Nedd4-2) co-expression. The amount of hERG1 channels in total HEK293 lysates and at the cell surface, as assessed by Western blot and biotinylation assays, respectively, were concomitantly decreased. Nedd4-2 and hERG1 interact via a PY motif located in the C-terminus of hERG1. Finally, we determined that Nedd4-2 mediates ubiquitylation of hERG1 and that deletion of this motif affects Nedd4-2-dependent regulation. These results suggest that ubiquitylation of the hERG1 protein by Nedd4-2, and its subsequent down-regulation, could represent an important mechanism for modulation of the duration of the human cardiac action potential.

Stereoselective Inhibition of the hERG1 Potassium Channel

A growing number of drugs have been shown to prolong cardiac repolarization, predisposing individuals to life-threatening ventricular arrhythmias known as Torsades de Pointes. Most of these drugs are known to interfere with the human ether à-gogo related gene 1 (hERG1) channel, whose current is one of the main determinants of action potential duration. Prolonged repolarization is reflected by lengthening of the QT interval of the electrocardiogram, as seen in the suitably named drug-induced long QT syndrome. Chirality (presence of an asymmetric atom) is a common feature of marketed drugs, which can therefore exist in at least two enantiomers with distinct three-dimensional structures and possibly distinct biological fates. Both the pharmacokinetic and pharmacodynamic properties can differ between enantiomers, as well as also between individuals who take the drug due to metabolic polymorphisms. Despite the large number of reports about drugs reducing the hERG1 current, potential stereoselective contributions have only been scarcely investigated. In this review, we present a non-exhaustive list of clinically important molecules which display chiral toxicity that may be related to hERG1-blocking properties. We particularly focus on methadone cardiotoxicity, which illustrates the importance of the stereoselective effect of drug chirality as well as individual variations resulting from pharmacogenetics. Furthermore, it seems likely that, during drug development, consideration of chirality in lead optimization and systematic assessment of the hERG1 current block with all enantiomers could contribute to the reduction of the risk of drug-induced LQTS.