The endocannabinoid ARA-S acts as an endogenous rescuing factor of Kv7.1/KCNE1 carrying Long QT syndrome mutations and paves the way for drug development strategies

Abstract

Abstract Funding Acknowledgements Type of funding sources: Public grant(s) – EU funding. Main funding source(s): European Union’s Horizon 2020 research and innovation program Long QT Syndrome (LQTS) is a cardiac arrhythmia characterized by delayed ventricular repolarization. LQTS occurs when there are loss-of-function mutations in genes encoding for voltage-gated channels and their beta subunits that contribute to the cardiac action potential. Among them, the most mutated channel is Kv7.1/KCNE1, which encodes for the slowly activating component of the delayed rectifier K+ current (IKs). Families with inherited mutations in the Kv7.1/KCNE1 exhibit inconsistent correlations between genotypes and phenotypes making the clinical management of LQTS challenging. A possible strategy to help guide clinical approaches involves exploring endogenous compounds capable of modulating the Kv7.1/KCNE1 channel function, acting as potential disease modifiers. Using the two-electrode voltage clamp technique, we previously found that the endogenous endocannabinoid-like compound ARA-S, which has an arachidonic acid tail and a serine head group, facilitates activation of the wild-type Kv7.1/KCNE1 channel expressed in Xenopus oocytes. This is seen as a left shift in the voltage dependence of channel opening and an increase in conductance. Moreover, we showed that ARA-S restores a physiological QT interval in isolated guinea pig hearts suffering from drug-induced LQTS. Hence, ARA-S is an interesting endogenous Kv7.1/KCNE1 channel modulator that may influence disease severity. However, it is uncertain whether ARA-S also activates the Kv7.1/KCNE1 when it carries LQTS-associated mutations. Therefore, the current study involves studying the effect of ARA-S on mutated Kv7.1/KCNE1 channels using two-electrode voltage clamp electrophysiology. We find that ARA-S facilitates the activation of all tested LQTS mutants. However, the magnitude of the effect varies among them, suggesting that the compound´s effect is dependent on the genetic background. To understand why ARA-S has different effects, we use computational simulation approaches to study how mutations in the putative ARA-S binding sites change ARA-S efficacy. This understanding aids in developing new substances that can activate different mutations, by, for example, modifying the ARA-S tail to enhance its efficacy. We also try to understand the concentration of ARA-S required to restore a wild-type-like function of each mutant, considering the mutant's intrinsic behavior and the apparent affinity of ARA-S for the specific mutant. Altogether, this work finds ARA-S to be an efficient activator of LQTS-associated mutant Kv7.1/KCNE1 channels and is a first step towards understanding how the genetic background determines the effect of this endogenous modulator.