The cardiac late Na+ current (late INa) generates persistent inward currents throughout the plateau phase of the ventricular action potential and is an important determinant of repolarisation rate, EADs and arrythmia risk¹. As inhibition of late INa can offset drug effects on hERG and other repolarising K⁺conductances, it is one of the key cardiac channels in the Comprehensive in vitro Pro-arrythmia Assay (CiPA) panel being developed by the FDA to improve human clinical arrythmia risk assessment²̛ ³.
The standard CiPA late INa assay uses the anemone toxin ATX-II to pharmacologically inhibit inactivation and produce persistent openings of wildtype (WT) Nav1.5 channels, but this method is variable and non-physiological. In contrast, several mutations in the SCN5A gene cause a form of hereditary long QT syndrome (LQT3) by promoting late openings⁴. The ΔKPQ mutation deletes residues Lys 1505, Pro 1506 and Gln 1507 and results in a sustained, non-inactivating current during long depolarizations which causes prolongation of the action potential and can result in fatal ventricular arrhythmias such as Torsade de Pointes (TdP)⁵̛ ⁶. Here we utilised a stable Nav1.5 LQT3 mutant cell line to optimise and validate a high throughput automated patch clamp late INa assay on the SyncroPatch 384i platform. High quality gigaseal recordings were obtained with a high success rate, enabling the efficient and accurate determination of relevant biophysical and pharmacological properties of this CiPA-compliant late Nav1.5 assay. The combination of the SyncroPatch 384i automated patch clamp system and Nav1.5 ΔKPQ cell line created a reliable high throughput cardiac safety screening assay without the need for openers like ATX-II toxin.