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Journal of Biomolecular Screening
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Validation of a Medium-Throughput Electrophysiological Assay for KCNQ2/3 Channel Enhancers Using IonWorks HT

Flora Jow

Discovery Neuroscience, Wyeth Research, Princeton, New Jersey

Ru Shen

Discovery Neuroscience, Wyeth Research, Princeton, New Jersey

Pranab Chanda

Discovery Neuroscience, Wyeth Research, Princeton, New Jersey

Eugene Tseng

Discovery Neuroscience, Wyeth Research, Princeton, New Jersey

Howard Zhang

Discovery Neuroscience, Wyeth Research, Princeton, New Jersey

Jeffrey Kennedy

Discovery Neuroscience, Wyeth Research, Princeton, New Jersey

John Dunlop

Discovery Neuroscience, Wyeth Research, Princeton, New Jersey

Mark R. Bowlby

Discovery Neuroscience, Wyeth Research, Princeton, New Jersey, bowlbym{at}wyeth.com

Enhancers of KCNQ channels are known to be effective in chronic pain models. To discover novel enhancers of KCNQ channels, the authors developed a medium-throughput electrophysiological assay by using the IonWorks platform. Screening of 20 CHO-K1 clones stably expressing KCNQ2/3 was performed on the IonWorks HT until the best clone (judged from seal rate, current level, and stability) was obtained. The KCNQ2/3 current amplitude in the cells was found to increase from 60 ± 15 pA to 473 ± 80 pA (at —10 mV), and the expression rate was increased by 56% when the cells were incubated at 27 °C overnight. The clone used for compound screening had a seal rate of greater than 90% and an overall success rate of greater than 70%. The voltage step protocol (hold cells at —80 mV and depolarize to —10 mV for 1 s) was designed to provide moderate current but still allow for pharmacological current enhancement. EC50s were generated from 8-point concentration-response curves with a control compound on each plate using compounds that were also tested with conventional patch clamp. The authors found that there was a very good correlation (R2 > 0.9) between the 2 assays, thus demonstrating the highly predictive nature of the IonWorks assay. (Journal of Biomolecular Screening 2007;1059-1067)

Key Words: ion channel • electrophysiology • opener • blocker • efflux

Journal of Biomolecular Screening, Vol. 12, No. 8, 1059-1067 (2007)
DOI: 10.1177/1087057107307448
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