Sharan R. Srinivasan, Haoran Huang, Wei-Chih Chang, Joshua A. Nasburg, Hai M. Nguyen, Tim Strassmaier, Heike Wulff and Vikram G. Shakkottai
Molecular Pharmacology 2022
Publication Date: July 1, 2022
DOI: 10.1124/molpharm.121.000478
Impaired cerebellar Purkinje neuron firing resulting from reduced expression of large-conductance calcium-activated potassium (BK) channels is a consistent feature in models of inherited neurodegenerative spinocerebellar ataxia (SCA). Restoring BK channel expression improves motor function and delays cerebellar degeneration, indicating that BK channels are an attractive therapeutic target. Current BK channel activators lack specificity and potency and are therefore poor templates for future drug development. We implemented an automated patch clamp platform for high-throughput drug discovery of BK channel activators using the Nanion SyncroPatch 384PE system. We screened over 15,000 compounds for their ability to increase BK channel current amplitude under conditions of lower intracellular calcium that is present in disease. We identified several novel BK channel activators that were then retested on the SyncroPatch 384PE to generate concentration-response curves (CRCs). Compounds with favorable CRCs were subsequently tested for their ability to improve irregular cerebellar Purkinje neuron spiking, characteristic of BK channel dysfunction in SCA1 mice. We identified a novel BK channel activator, 4-chloro-N-(5-chloro-2-cyanophenyl)-3-(trifluoromethyl)benzene-1-sulfonamide (herein renamed BK-20), that exhibited a more potent half-maximal activation of BK current (pAC50 = 4.64) than NS-1619 (pAC50 = 3.7) at a free internal calcium concentration of 270 nM in a heterologous expression system and improved spiking regularity in SCA1 Purkinje neurons. BK-20 had no activity on small-conductance calcium-activated potassium (SK)1–3 channels but interestingly was a potent blocker of the T-type calcium channel, Cav3.1 (IC50 = 1.05 μM). Our work describes both a novel compound for further drug development in disorders with irregular Purkinje spiking and a unique platform for drug discovery in degenerative ataxias.
Read the full paper in Molecular Pharmacology.