Protocol-Dependent Differences in IC₅₀ Values Measured in Human Ether-Á-Go-Go-Related Gene Assays Occur in a...

Current guidelines around preclinical screening for drug-induced arrhythmias require the measurement of the potency of block of voltage-gated potassium channel subtype 11.1 (K_v11.1) as a surrogate for risk. A shortcoming of this approach is that the measured IC₅₀ of K_v11.1 block varies widely depending on the voltage protocol used in electrophysiological assays. In this study, we aimed to investigate the factors that contribute to these differences and to identify whether it is possible to make predictions about protocol-dependent block that might facilitate the comparison of potencies measured using different assays. Our data demonstrate that state preferential binding, together with drug-binding kinetics and trapping, is an important determinant of the protocol dependence of K_v11.1 block. We show for the first time that differences in IC₅₀ measured between protocols occurs in a predictable way, such that machine-learning algorithms trained using a selection of simple voltage protocols can indeed predict protocol-dependent potency. Furthermore, we also show that the preference of a drug for binding to the open versus the inactivated state of K_v11.1 can also be inferred from differences in IC₅₀ values measured between protocols. Our work therefore identifies how state preferential drug binding is a major determinant of the protocol dependence of IC₅₀ values measured in preclinical K_v11.1 assays. It also provides a novel method for quantifying the state dependence of K_v11.1 drug binding that will facilitate the development of more complete models of drug binding to K_v11.1 and improve our understanding of proarrhythmic risk associated with compounds that block K_v11.1.