Structure-based identification and characterisation of novel inhibitors of KNa1.1 potassium channels, a stratified target for KCNT1-related epilepsy

Several types of drug-resistant epileptic encephalopathies of infancy have been associated with mutations in the KCNT1 gene, which encodes the sodium-activated potassium channel subunit KNa1.1. These mutations are commonly gain-of-function, increasing channel activity, therefore inhibition by drugs is proposed as a stratified approach to treat disorders. To date, quinidine therapy has been trialled with several patients, but mostly with unsuccessful outcomes, which has been linked to its low potency and lack of specificity. Here we describe the use of a cryo-electron microscopy-derived KNa1.1 structure and mutational analysis to identify the quinidine biding site and identified novel inhibitors that target this site using computational methods. We describe six compounds that inhibit KNa1.1 channels with low- and sub-micromolar potencies, likely through binding in the intracellular pore vestibule. In preliminary hERG inhibition and cytotoxicity assays, two compounds showed little effect. These compounds may provide starting points for the development of novel pharmacophores for KNa1.1 inhibition, with the view to treating KCNT1-associated epilepsy and, with their potencies higher than quinidine, could become key tool compounds to further study this channel. Furthermore, this study illustrates the potential for utilising cryo-electron microscopy in ion channel drug discovery.