Variability in the measurement of hERG potassium channel inhibition: effects of temperature and stimulus pattern.

ABSTRACT

INTRODUCTION: In vitro evaluation of drug effects on hERG K(+) channels is a valuable tool for identifying potential proarrhythmic side effects in drug safety testing. Patch-clamp recording of hERG K(+) current in mammalian cells can accurately evaluate drug effects, but the methodology has not been standardized, and results vary widely. Our objective was to evaluate two potential sources of variability the temperature at which recordings are performed and the voltage pulse protocol used to activate hERG K(+) channels expressed in HEK293 cells. METHODS: A panel of 15 drugs that spanned a broad range of potency for hERG inhibition and pharmacological class was evaluated at both room and near-physiological temperatures using several patch-clamp voltage protocols. Concentration-response analysis was performed with three stimulus protocols 0.5- and 2-s step pulses, or a step-ramp pattern. RESULTS: Block by 2 of the 15 drugs tested, d,l-sotalol (antiarrhythmic) and erythromycin (antibiotic), was markedly temperature sensitive. hERG inhibition measured using a 2-s step-pulse protocol underestimated erythromycin potency compared with results obtained with a step-ramp protocol. Using conservative acceptance criteria and the step-ramp protocol, the IC(50) values for hERG block differed by less than twofold for 15 drugs. DISCUSSION: Data obtained at near-physiological temperatures using a step-ramp pattern are highly repeatable and provide a conservative safety evaluation of hERG inhibition.