Correlation of IKr Block with Action Potential Changes in Human Stem Cell Derived Cardiomyocytes

Abstract

Drugs that prolong the cardiac action potential (AP) often act through block of IKr, the native current mediated by HERG. HERG block in heterologous expression systems (e.g., HEK cells) is a primary assay for determining FDA safety screening. However, the relationship between IKr block and AP prolongation is complex and may vary from cell to cell depending on ion channel expression. Additionally, use dependence may differ significantly in heterologous expression systems. We used cardiomyocytes derived from human pluripotent stem cells (hiPSCDCMs) (Cellular Dynamics, WI) to measure drug effects on IKr and the cardiac AP in the same cell. Cells were cultured according to the manufacturer's directions and used 7-25 days after plating. Standard electrophysiology voltage clamp (whole-cell rupture or Amphotericin B perforated patch), was used. hiPSCDCMs lack endogenous IK1, so virtual IK1 was added via a Cybercyte System (Cytocybernetics, NY) to obtain stable electrical behavior. IKr was recorded using a 4 s voltage step from a holding potential of −80 mV to +30 mV (P1) followed by a single voltage step to −50 mV (P2) for 6 s (30 s inter-sweep interval). Following IKr measurements, cardiac APs were triggered by a 1.5-2.0 nA stimulus for 1.5 ms at 0.5 Hz. These IKr and AP measurement protocols were then repeated with dofetilide or loratadine. Dofetilide (300 nM) completely blocked IKr and loratidine (6 μM) blocked IKr 36±1%. After observing IKr block, both dofetilide and loratadine prolonged the cardiac AP. These results demonstrate a direct correlation between IKr block and effects on the cardiac AP. In accordance with CiPA, this approach may provide a more robust and comprehensive cardiac safety screen as compared to the standard HERG assay.