IQM-110, a new selective inhibitor of KChIP and its pharmacological effects on Kv4., Kv4./KChIP and Kv4./KChIP currents

Abstract

Kv4 channels generate outward currents activated at subthreshold membrane potentials and are responsible for the repolarization of both cardiac and neuronal action potentials through ITO (transient outward current) and IA (A-type K+ current), respectively. Kv4 dysfunctions have been identified in cardiac diseases (i.e. Brugada syndrome or atrial fibrillation) as well as in neuronal diseases (i.e. schizophrenia, epilepsy or Alzheimer’s disease). However, Kv4 channels need to assemble with regulatory or β subunits to fully reproduce ITO and IA currents. Among them, we will focus on KChIPs (potassium channel interacting proteins), being KChIP3 predominant in the brain and KChIP2 in both brain and heart. The assembly of these β subunits alters not only the biophysical properties of the channel, but also its pharmacology. For this reason, we have analysed the electrophysiological effects of IQM-22110 (a new compound designed to bind KChIP3) on the currents generated by Kv4.3, Kv4.3/KChIP2 and Kv4.3/KChIP3 channels. CHO cells were transiently transfected (Kv4.3, Kv4.3/KChIP2 or Kv4.3/KChIP3), and the potassium currents were recorded using the whole-cell patch-clamp technique. Our results indicate that IQM-22110 exerts differential effects on Kv4.3, Kv4.3/KChIP2 and Kv4.3/KChIP3 currents. First of all, the concentration-dependence of inhibition was modified, being the IC50 = 5.5 μM for Kv4.3 channels, 10.4 μM for Kv4.3/KChIP2 and exhibited a biphasic curve when KChIP3 is present (IC50(1) = 0.03 μM and IC50(2) = 26.3 μM), suggesting two binding sites for IQM-22110 in this complex. With molecular dynamics and sitedirected mutagenesis, we have identified two aminoacids (Y130 and K166) in KChIP3 critical to the binding of IQM-22110 to its high affinity site. Also, we selected equipotent concentrations of IQM22110 to further study the biophysical effects of this compound on each situation, demonstrating that IQM-22110 binds to the closed-active state of the channel. In this study we concluded that: i) at low concentrations, IQM-22110 is a selective inhibitor of Kv4.3/KChIP3, ii) Y130 and K166 in KChIP3 are critical to the binding of IQM-22110 to the high affinity site in Kv4.3/KChIP3, and iii) IQM-22110 binds to the closed-active state of Kv4.3, Kv4.3/KChIP2 and Kv4.3/KChIP3.