Evaluation of an in silico cardiac safety assay: using ion channel screening data to predict QT interval changes in the rabbit ventricular wedge

Abstract

Introduction: Drugs that prolong the QT interval on the electrocardiogram present a major safety concern for pharmaceutical companies and regulatory agencies. Despite a range of assays performed to assess compound effects on the QT interval, QT prolongation remains a major cause of attrition during compound development. In silico assays could alleviate such problems. In this study we evaluated an in silico method of predicting the results of a rabbit left-ventricular wedge assay.

Methods: Concentration-effect data were acquired from either: the high-throughput IonWorks/FLIPR; the medium-throughput PatchXpress ion channel assays; or QSAR, a statistical IC50 value prediction model, for hERG, fast sodium, L-type calcium and KCNQ1/minK channels. Drug block of channels was incorporated into a mathematical differential equation model of rabbit ventricular myocyte electrophysiology through modification of the maximal conductance of each channel by a factor dependent on the IC50 value, Hill coefficient and concentration of each compound tested. Simulations were performed and agreement with experimental results, based upon input data from the different assays, was evaluated.

Results: The assay was found to be 78% accurate, 72% sensitive and 81% specific when predicting QT prolongation (>10%) using PatchXpress assay data (77 compounds). Similar levels of predictivity were demonstrated using IonWorks/FLIPR data (121 compounds) with 78% accuracy, 73% sensitivity and 80% specificity. QT shortening (<-10%) was predicted with 77% accuracy, 33% sensitivity and 90% specificity using PatchXpress data and 71% accuracy, 42% sensitivity and 81% specificity using IonWorks/FLIPR data. Strong quantitative agreement between simulation and experimental results was also evident.

Discussion: The in silico action potential assay demonstrates good predictive ability, and is suitable for very high-throughput use in early drug development. Adoption of such an assay into cardiovascular safety assessment, integrating ion channel data from routine screens to infer results of animal-based tests, could provide a cost- and time-effective cardiac safety screen.

Keywords: AP (D); Action Potential (Duration); Action potential; Cardiac safety; Compound screening; Concentration for 50% Inhibition; ECG; ECVAM; European Centre for the Validation of Alternative Methods; FLIPR; FLuorescence Imaging Plate Reader; GSK; GlaxoSmithKline; I(Kr); I(Ks); IC(50); ICH; International Conference for Harmonization; Ion channels; Mathematical model; Methods; QSAR; QT interval; Quantitative Structure Activity Relationship; Rabbit ventricular wedge; TdP; Torsades de Pointes; electrocardiogram; hERG; human-Ether-a-go-go Related Gene; minus log(10) of IC(50); pIC(50); rapid delayed rectifier potassium current; slow delayed rectifier potassium current.

Fig. 1

a) An action potential generated from a single cell simulation and b) a pseudo-ECG from a one-dimensional tissue simulation at a range of concentrations for compound 2659 (see Supplementary data). Simulations were performed using PatchXpress data, parameterising the drug block model with the IC₅₀ value and Hill coefficient in the single cell simulation and with just the IC₅₀ value (and assuming the Hill coefficient is 1) in the one-dimensional simulation. The intervals used for calculation of the APD90 value and QT interval from the simulated control result are indicated in a) and b). Arrows indicate the effect observed in the APD90 value/QT interval with increasing compound concentration. In c) the percent change in APD90/QT interval determined from the simulation results and experimental results (both individual preparation results and the average of these) in the rabbit ventricular wedge assay is plotted for comparison.

Fig. 2

Plot of concentrations (EC10 values) at which a 10% change in APD90 value/QT interval is expected, as interpolated from simulation and experimental results, for each compound. The results from case 6, using PatchXpress data, are presented. Compounds exhibiting more than 10% change in their experimental or simulation results after drug administration, as compared to the control measurement, at at least one test concentration were included. This enabled interpolation of the EC10 values. Points plotted with an asterisk (*) are compounds for which more than a 10% change in the QT interval length (as compared to the control measurement) is exhibited in both simulation and experimental results. These points are plotted in one of four quadrants according to the classification of the simulation and experimental results: both experimental and simulation results show prolongation (quadrant 1), both show shortening (quadrant 3), experimental results show shortening and simulation results show prolongation (quadrant 2), simulation results show shortening and experimental results show prolongation (quadrant 4). Points plotted with a circle (○) are compounds for which only one of the experimental and simulation results show more than a 10% change in the QT interval as compared to the control. Interpolation is used to determine the EC10 value from this result. For the remaining experimental or simulation result, the percent change in QT interval is between −10% and 10% (showing “no effect”) at all concentrations tested. This result is assumed to have an EC10 value corresponding to the maximum concentration tested in the rabbit wedge assay amongst all the compounds which show “no effect” (with the percent change in APD90 or QT interval between −10% and 10%), which has a value of 500 μM. Half log unit lines which correspond to the error commonly associated with the ion channel assays are included as an indication of the range of accepted error.