Enhancing cardiac safety evaluation: Assessing drug interactions with the late Nav1.5 current

Late Na + current (INa,L) contributes to cardiac action potential (AP) and helps maintain Na + homeostasis. Contrary to the effect on Nav1.5, hERG and Cav1.2 channels, the effect of novel drugs on INa,L is not routinely studied in safety studies. Yet, INa,L inhibitors can counterbalance hERG blockade and be associated with anti-arrhythmic potential. Hence, evaluating the effects of drugs against Nav1.5, hERG and Cav1.2 channels provides valuable cardiotoxic insights, but does not fully predict changes in the electrophysiological and contractile properties of cardiomyocytes. To address this deficiency, we evaluated the effects of selective and non-selective INa,L inhibitors on cardiomyocyte function. Compounds known to inhibit INa,L (GS-967 specific for INa,L; ranolazine specific for both hERG and INa,L; loperamide which is a hERG, Nav1.5 and Cav1.2 inhibitor) and four preclinical compounds were tested for their effects on cardiac ion channels (peak Nav1.5, hERG, Cav1.2 and INa,L) with automated patch-clamp and multi-electrode array (MEA) in hiPSC-derived cardiomyocytes for electrophysiological properties, and contractility in human primary cardiomyocytes from consented donor hearts with MyoBLAZER™. Each compound was tested separately at multiple concentrations in the presence of ATX-II, a selective enhancer of INa,L. GS-967 and ranolazine reversed ATX-II-induced increases in contractility and field potential duration (FPD) in a concentration-dependent manner providing evidence of a functional INa,L in both hiPSC-derived cardiomyocytes and adult cardiomyocytes. Next, we evaluated the effects of four preclinical compounds. Two out of the four compounds showed similar behavior to GS-967 and ranolazine. For example, compound A inhibited ion channels (hERG, Nav1.5, Cav1.2 and INa,L with IC50 values of 7.9 mM, 12.4 mM, 0.8 mM, respectively) and reversed ATX-II changes on contractility and FPD with IC50 values of 0.99 mM and 2.3 mM, respectively. Here, we developed a protocol for assessing drug interactions with INaL on cardiomyocytes. This assay enhances our ability to predict cardiotoxicity potential and its incorporation into the traditional compound derisking strategy strengthens confidence in advancing molecules into clinical development.