Correlation electrical repolarization and membrane hERG abundance in iPSC cardiomyocytes induced by chronic drugs actions on channel trafficking

The acute effects of drugs (~1 h) on the electrical activity, particularly those related to hERG block have been extensively studied in iPSC-derived cardiomyocytes (iPSC -CMs). However, long-term (>12 h) drug exposure can also lead to slowly developing actions on hERG channels that result in QT prolongation, the most common are drugs that inhibit hERG trafficking to the surface membrane and cause an acquired long QT phenotype. The aim of the study is to assess the ability of iPSC-CM based assays to study the action of drugs known to inhibit hERG trafficking over a longer period using serum-free solutions. The following functional cellular parameters were measured over 72 h at 24 h intervals: (i) action potential duration (ii) contractile kinetics (iii) iPSC -CM monolayer integrity (iv) plasmalemma integrity (iv) membrane hERG expression. We used iCell2 (FujiFilm-CDI) iPSC -CMs, which we plated on a 96well plate and incubated in a serum free media. We performed automated image and signal analysis using a proprietary analysis platform (CellOPTIQ®- Clyde Biosciences). We tested 6 concentrations of Pentamidine and Arsenic Trioxide (ATO) which are both known to affect hERG trafficking and assessed chronic toxicity, electrophysiological, metabolic dysfunction. Pentamidine showed obvious prolongation of APD90 in the clinical range (1-3 μM) after 72 h and after 24 h at higher concentration (3-10 μM). ATO induced APD prolongation at 1-3 μM but shortening at 10 μM indicating mixed ion channel trafficking or metabolic effects. Western blot analysis of the membrane fraction showed significant downregulation of hERG expression of cells exposed to 48 h of 1 μM pentamidine. This work demonstrates the utility of chronic studies of IPSC-CMs to study the medium-long term actions of drugs on the hERG activity. The associated biochemical assays can confirm the extent to which reduction of hERG expression in the plasmalemmal membrane is responsible for the action.