The antiarrhythmic compound efsevin directly modulates voltage-dependent anion channel 2 by binding to its inner wall and enhancing mitochondrial Ca2+ uptake.

Fabiola Wilting,Robin Kopp,Ohyun Kwon,Philip A Gurnev,Thomas Gudermann,Annette Nicke,Nathan J Dupper,Johann Schredelseker,Anna Schedel

doi:10.1111/bph.15022

Abstract

Background and PurposeThe synthetic compound efsevin was recently identified to suppress arrhythmogenesis in models of cardiac arrhythmia, making it a promising candidate for antiarrhythmic therapy. Its activity was shown to be dependent on the voltage‐dependent anion channel 2 (VDAC2) in the outer mitochondrial membrane. Here, we investigated the molecular mechanism of the efsevin–VDAC2 interaction.Experimental ApproachTo evaluate the functional interaction of efsevin and VDAC2, we measured currents through recombinant VDAC2 in planar lipid bilayers. Using molecular ligand‐protein docking and mutational analysis, we identified the efsevin binding site on VDAC2. Finally, physiological consequences of the efsevin‐induced modulation of VDAC2 were analysed in HL‐1 cardiomyocytes.Key ResultsIn lipid bilayers, efsevin reduced VDAC2 conductance and shifted the channel's open probability towards less anion‐selective closed states. Efsevin binds to a binding pocket formed by the inner channel wall and the pore‐lining N‐terminal α‐helix. Exchange of amino acids N207, K236 and N238 within this pocket for alanines abolished the channel's efsevin‐responsiveness. Upon heterologous expression in HL‐1 cardiomyocytes, both channels, wild‐type VDAC2 and the efsevin‐insensitive VDAC2AAA restored mitochondrial Ca2+ uptake, but only wild‐type VDAC2 was sensitive to efsevin.Conclusion and ImplicationsIn summary, our data indicate a direct interaction of efsevin with VDAC2 inside the channel pore that leads to modified gating and results in enhanced SR‐mitochondria Ca2+ transfer. This study sheds new light on the function of VDAC2 and provides a basis for structure‐aided chemical optimization of efsevin.

Highlights

Cardiovascular diseases represent the primary cause of death and hospitalization worldwide (Benjamin et al, 2018)
We demonstrate that the observed changes in zebrafish voltage-dependent anion channel 2 (VDAC2) (zVDAC2) electrophysiology translate into enhanced mitochondrial Ca2+ uptake and thereby explain the antiarrhythmic effect of efsevin
Efsevin suppresses arrhythmogenesis in both, models for Ca2+ overload and models for inherited arrhythmias like catecholaminergic polymorphic

Summary

| INTRODUCTION

Cardiovascular diseases represent the primary cause of death and hospitalization worldwide (Benjamin et al, 2018). Action and binding structure aided-drug unclear whether efsevin directly interacts with the VDAC2 peptide and thereby modulates the electrophysiological properties of the channel or if the observed effects require a yet unidentified protein partner. To address this issue, we expressed and purified recombinant zebrafish VDAC2 (zVDAC2) protein and inserted it into planar lipid bilayers. Statistical analysis was only performed for sample sizes >5 from independent experiments (i.e. individual insertions in lipid bilayers, individual runs of computational docking, and individual cultures of HL-1 cardiomyocytes), and the obtained data were used for statistical analysis without normalization and outlier removal. Key protein targets and ligands in this article are hyperlinked to corresponding entries in http://www.guidetopharmacology.org, the common portal for data from the IUPHAR/BPS Guide to PHARMACOLOGY (Harding et al, 2018), PHARMACOLOGY

| RESULTS

Findings

| DISCUSSION