title: Novel neuroprotective K+ channel inhibitor identified by high-throughput screening in yeast.
abstract: Discovery of K+ channel modulators is limited by low-throughput capacity of existing K+ channel assays. To enable high-throughput screening for novel pharmacological modulators of K+ channels, we developed an assay based on growth of yeast that functionally expresses mammalian Kir2.1 channels. Screening of 10,000 small molecules from a combinatorial chemical library yielded 42 potential Kir2.1 inhibitors. One compound, 3-bicyclo[2.2.1]hept-2-yl-benzene-1,2-diol, was confirmed to inhibit K+ channels in patch-clamp measurements in mammalian cells with EC50 values of 60 and 1 microM for Kir2.1 and Kv2.1 channels, respectively. Inhibition of Kv2.1 channels decreased in the presence of the external pore blocker tetraethylammonium (TEA) and depended on a residue required for extracellular TEA action, suggesting that the identified compound targets the external mouth of the channel. Furthermore, at the nontoxic concentration of 3 microM, the identified compound completely abolished in vitro neuronal apoptosis mediated by Kv2.1 channels. Therefore, yeast-based screening has identified a novel uncharged neuroprotective mammalian K+ channel inhibitor.
authors.affiliation: Department of Pharmacology, University of Pittsburgh School of Medicine, Pittsburgh PA 15261, USA. elm59@pitt.edu
authors.forename: Elena
authors.initials: E
authors.lastname: Zaks-Makhina
authors.forename: Yonjung
authors.initials: Y
authors.lastname: Kim
authors.forename: Elias
authors.initials: E
authors.lastname: Aizenman
authors.forename: Edwin S
authors.initials: ES
authors.lastname: Levitan
mesh_terms: D000818:Animals; D001643:Bridged Bicyclo Compounds / Q000737:chemistry / Q000494:pharmacology*; D002478:Cells, Cultured; D051668:Delayed Rectifier Potassium Channels; D004353:Drug Evaluation, Preclinical / Q000379:methods*; D006801:Humans; D018696:Neuroprotective Agents / Q000302:isolation & purification* / Q000494:pharmacology; D026902:Potassium Channel Blockers / Q000302:isolation & purification* / Q000494:pharmacology; D015221:Potassium Channels / Q000187:drug effects / Q000378:metabolism*; D024642:Potassium Channels, Voltage-Gated; D051381:Rats; D017207:Rats, Sprague-Dawley; D012441:Saccharomyces cerevisiae / Q000187:drug effects / Q000378:metabolism*; D051669:Shab Potassium Channels; D019789:Tetraethylammonium / Q000494:pharmacology
chemical_list: C481601:3-bicyclo(2.2.1)hept-2-yl-benzene-1,2-diol; D001643:Bridged Bicyclo Compounds; D051668:Delayed Rectifier Potassium Channels; C496988:KCNB1 protein, human; C496991:Kcnb1 protein, rat; D018696:Neuroprotective Agents; D026902:Potassium Channel Blockers; D015221:Potassium Channels; D024642:Potassium Channels, Voltage-Gated; D051669:Shab Potassium Channels; D019789:Tetraethylammonium