Kinetic Analysis of Membrane Potential Dye Response to NaV1.7 Channel Activation Identifies Antagonists with Pharmacological Selectivity against NaV1.5. (June 2016)

• Record Type: Journal Article
• Authors: Finley, Michael; Cassaday, Jason; Kreamer, Tony; Li, Xinnian; Solly, Kelli; O'Donnell, Greg; Clements, Michelle; Converso, Antonella; Cook, Sean; Daley, Chris; Kraus, Richard; Lai, Ming-Tain; Layton, Mark; Lemaire, Wei; Staas, Donnette; Wang, Jixin
• Is Part Of: Journal of biomolecular screening. Volume 21:Number 5(2016)
• Page Start: 480
• Page End: 489
• Publication Date: 2016-06
• Subjects: sodium channel -- high-throughput screen -- membrane potential; Drugs -- Analysis -- Periodicals; Drugs -- Testing -- Periodicals; Biomolecules -- Analysis -- Periodicals; 572.36
• DOI: 10.1177/1087057116629669
• Languages: English
• ISSNs: 1087-0571

Abstract

The NaV 1.7 voltage-gated sodium channel is a highly valued target for the treatment of neuropathic pain due to its expression in pain-sensing neurons and human genetic mutations in the gene encoding NaV 1.7, resulting in either loss-of-function (e.g., congenital analgesia) or gain-of-function (e.g., paroxysmal extreme pain disorder) pain phenotypes. We exploited existing technologies in a novel manner to identify selective antagonists of NaV 1.7. A full-deck high-throughput screen was developed for both NaV 1.7 and cardiac NaV 1.5 channels using a cell-based membrane potential dye FLIPR assay. In assay development, known local anesthetic site inhibitors produced a decrease in maximal response; however, a subset of compounds exhibited a concentration-dependent delay in the onset of the response with little change in the peak of the response at any concentration. Therefore, two methods of analysis were employed for the screen: one to measure peak response and another to measure area under the curve, which would capture the delay-to-onset phenotype. Although a number of compounds were identified by a selective reduction in peak response in NaV 1.7 relative to 1.5, the AUC measurement and a subsequent refinement of this measurement were able to differentiate compounds with NaV 1.7 pharmacological selectivity over NaV 1.5 as confirmed in electrophysiology.