Ion channels are membrane proteins that selectively conduct ions across cellular membranes, existing in both excitable cells and non-excitable cells. Ion channels play a critical role in cellular physiology, including electrical and cellular signaling, ion homeostasis, and hormone secretion. The objective of this proposal is to find small molecule chemical probes that activate or potentiate a disease-causing potassium channel - KCNQ1. Abnormality of this channel, through co-assembly with other accessory subunits in different tissues, is thought to be causal to both cardiac diseases and epithelial cell diseases, such as long QT syndrome and cystic fibrosis. We have developed cell lines, validated the cell lines in a fluorescence-based thallium surrogate flux assay, and perform feasibility trials of high throughput screening. This proposal outlines a specific plan to conduct a >300,000-compound screen using the KCNQ1 cell line to search for specific activators/potentiators. The active compounds will then be evaluated by automated patch-clamp recording of various cell lines expressing different combinations of KCNQ1/KCNE channels, including KCNE1, KCNE2, KCNE3, KCNE4 and KCNE5 b- subunits. Because different KCNE subunits co-assemble with KCNQ1 in different tissues, specific probes developed from this proposal will therefore be useful for tissue-specific investigations. In collaboration with Dr. Gordon Tomaselli in Division of Cardiology and Dr. Mark Donowitz in Hopkins Center for Epithelial Disorders of Johns Hopkins School of Medicine, our compound validation plan includes testing of the isolated compounds in native preparations. These active compounds, with further efforts in pharmacology and medicinal chemistry, may be exploited for therapeutic remedy of cardiac arrhythmia and water and salt imbalance in epithelial tissues.
Non-neuronal KCNQ1 potassium channels are critical functional components of heartbeat, hearing, and water/salt balance in lung and intestinal tissues. Dysfunction of these potassium channels results in serious arrhythmia, cardiac arrest, hearing loss, diarrhea, and cystic fibrosis. There is increasing evidence for their causality for long QT syndrome, familial atrial fibrillation, Jervell and Lange-Nielsen syndrome, Romano-Ward syndrome, short QT syndrome, and sudden infant death syndrome (SIDS). Discovery of small molecule probes through this project can therefore provide tools for better understanding of these channels, improve assessment of safety concerns in future pharmaceutical development, and develop therapeutics for the above mentioned diseases.
