title: Development and validation of a medium-throughput electrophysiological assay for KCNQ2/3 channel openers using QPatch HT.
abstract: The KCNQ2/3 channel has emerged as a drug target for a number of neurological disorders including pain and epilepsy. Known KCNQ2/3 openers have effects on two distinct biophysical properties of the channel: (1) a hyperpolarizing shift in the voltage dependence of channel activation (V(1/2)), and (2) an increase in channel open probability or peak whole-cell current. The current high-throughput screening assays for KCNQ2/3 openers measure changes of channel activity at sub-peak conductances and the output measure is a combination of effects on V(1/2) shift and peak current. Here, we describe a medium-throughput electrophysiological assay for screening KCNQ2/3 openers using the QPatch HT platform. We employed a double-pulse protocol that measures the shift in V(1/2) and the change in current amplitude at peak conductance voltage. Retigabine along with novel KCNQ2/3 openers were evaluated in this assay. Three classes of KCNQ2/3 openers were identified based on the hyperpolarizing shift in V(1/2) and the change in peak current. All three classes of compounds caused a hyperpolarizing shift in V(1/2), but they were differentiated by their respective effects on peak current amplitude (increase, decrease, or only modestly affecting peak current amplitude). KCNQ2/3 blockers were also identified with this assay. These compounds blocked currents without affecting voltage-dependent activation. In summary, we have developed a medium-throughput assay that can reliably detect changes in the biophysical properties of the KCNQ2/3 channel, V(1/2), and peak current amplitude, and therefore may serve as a reliable assay to evaluate KCNQ2/3 openers and blockers.
authors.affiliation: Neuroscience Research, Global Pharmaceutical Research and Development, Abbott Laboratories, Abbott Park, Illinois 60064-6125, USA. xufeng.zhang@abbott.com
authors.forename: Xu-Feng
authors.initials: XF
authors.lastname: Zhang
authors.forename: Di
authors.initials: D
authors.lastname: Zhang
authors.forename: Carol S
authors.initials: CS
authors.lastname: Surowy
authors.forename: Betty
authors.initials: B
authors.lastname: Yao
authors.forename: Michael F
authors.initials: MF
authors.lastname: Jarvis
authors.forename: Steve
authors.initials: S
authors.lastname: McGaraughty
authors.forename: Torben R
authors.initials: TR
authors.lastname: Neelands
mesh_terms: D002219:Carbamates / Q000737:chemistry / Q000378:metabolism; D055724:Electrophysiological Phenomena / Q000502:physiology; D057809:HEK293 Cells; D057166:High-Throughput Screening Assays / Q000379:methods / Q000592:standards*; D006801:Humans; D051658:KCNQ2 Potassium Channel / Q000145:classification / Q000378:metabolism*; D051659:KCNQ3 Potassium Channel / Q000145:classification / Q000378:metabolism*; D049990:Membrane Transport Modulators / Q000737:chemistry / Q000378:metabolism*; D018408:Patch-Clamp Techniques / Q000379:methods / Q000592:standards*; D010655:Phenylenediamines / Q000737:chemistry / Q000378:metabolism
chemical_list: D002219:Carbamates; D051658:KCNQ2 Potassium Channel; D051659:KCNQ3 Potassium Channel; C496916:KCNQ3 protein, human; D049990:Membrane Transport Modulators; D010655:Phenylenediamines; C101866:ezogabine