HERG Heteromeric 1A/1B and Homomeric 1A Channels Exhibit Differential Pharmacological Sensitivities

Abstract

The majority of hERG screens aiming to minimize the risk of drug-induced long QT syndrome have been conducted using heterologous systems expressing the hERG 1a subunit, yet both hERG 1a and 1b subunits contribute to the channels producing the repolarizing current IKr. Previous studies show that differences in gating in heteromeric 1a/1b vs. homomeric 1a channels markedly increase repolarizing current during the ventricular action potential and protect against QT prolongation in computational models. We conducted a pharmacological analysis of 50 compounds targeting hERG channels and selected for their chemical diversity to evaluate differences in sensitivity that may influence safety margins or contribute to a stratified risk analysis. Experiments were carried out using the IonWorksTM plate-based electrophysiology device. Non-cumulative, 8-point concentration effect curves were generated, with each point representing data from 20 to 30 cells. Potency was determined as IC50 values (∈1/4M) obtained from data normalized to vehicle and 100% blocking levels and fitted to the Hill equation. To minimize possible sources of variability, compound potency was assessed using test plates arranged in alternating columns of 1a and 1a/1b cells. Although most compounds had similar potencies at both variants, some surprising differences were observed. For example, fluoxetine (Prozac) was 6-fold more potent at blocking hERG 1a/1b compared to 1a channels. The results were robust when compounds were tested against the hERG 1a and 1a/1b cell lines in parallel, but statistical analysis encompassing longitudinal variation indicates such differences may not be sufficient to warrant routine use of hERG 1a/1b in preclinical high throughput screens. However, our findings have uncovered several important candidates for further risk evaluation as we learn more about native subunit composition in different populations or changes in subunit composition during development.