Key result
High-throughput fluorescence assay analyzes 10,000 hERG data points daily with no false-positives.
Why the study?
Cardiac side effects such as QT prolongation caused by hERG potassium channel inhibition lead to late-phase drug development failures, necessitating early screening methods for compound interactions with hERG channels.
A high-throughput fluorescence assay offers a rapid, robust, and semiautomated method for early screening of hERG potassium channel inhibition to prevent drug-induced QT prolongation.
May accelerate early hERG screening to mitigate QT risk; leaves open whether it supplants patch-clamp in regulatory pipelines.
The number of projects in drug development that fail in late phases because of cardiac side effects such as QT prolongation can impede drug discovery and development of projects. The molecular target responsible for QT prolongation by a wide range of pharmaceutical agents is the myocardial hERG potassium channel. It is therefore desirable to screen for compound interactions with the hERG channel at an early stage of drug development. Here, the authors report a cell-based fluorescence assay using membrane potential-sensitive fluorescent dyes and stably transfected hERG channels from CHO cells. The assay allows semiautomated screening of compounds for hERG activity on 384-well plates and is sufficiently rapid for testing a large number of compounds. The assay is robust as indicated by a Z' factor larger than 0.6. The throughput is in the range of 10,000 data points per day, which is significantly higher than any other method presently available for hERG. The data obtained with the fluorescence assay were in qualitative agreement with those from patch-clamp electrophysiological analysis. There were no false-positive hits, and the rate of false-negative compounds is currently 12% but might be further reduced by testing compounds at higher concentration. Quantitative differences between fluorescence and electrophysiological methods may be due to the use- or voltage-dependent activity of the antagonists.
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Dorn et al. (2005) studied QT prolongation. Cell-based fluorescence assay vs. Patch-clamp electrophysiological analysis was evaluated on hERG potassium channel inhibition. A high-throughput cell-based fluorescence assay for hERG channel inhibition achieved a Z' factor >0.6 and throughput of 10,000 data points/day, with 0% false-positives and 12% false-negatives.
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