Rb+ Flux through hERG Channels Affects the Potency of Channel Blocking Drugs: Correlation with Data Obtained Using a High-Throughput Rb+ Efflux Assay

Abstract

The nonradioactive Rb⁺ efflux assay has become a reliable and efficient high-throughput hERG screening method, but it is limited by its low sensitivity for potent hERG blockers. Using the patch clamp technique, the authors found that the low sensitivity is due in part to the use of Rb⁺ as the permeating cation in the assay. The affinities of the drugs measured by patch clamp technique in the presence of Rb⁺ were 3- to 10-fold lower than when measured by the same method in the presence of K⁺ ions. The apparent affinity of the drugs decreased even further when monitored bytheRb⁺ efflux assay. It was also observed that Rb⁺ had minimal effects on the activation properties of channels while there was a significant change in the half-inactivation potential. This voltage shift reduces hERG channel inactivation at efflux assay potentials, and will reduce the affinity of hERG-blocking drugs that bind to inactivated states of the channel. In combination with the effects of elevated extracellular ion concentrations, it is likely that Rb⁺ modulation of hERG channel inactivation is largely responsible for the reduced drug potencies observed in the Rb⁺ efflux assay.