Development and evaluation of high throughput functional assay methods for HERG potassium channel

Area of Science:

• Pharmacology
• Biophysics
• Drug Discovery

Background:

• The hERG channel is crucial for cardiac repolarization.
• Assessing hERG channel inhibition is vital for drug safety to prevent cardiotoxicity.
• Existing assay methods present challenges in terms of cost, sensitivity, and accuracy.

Purpose of the Study:

• To develop and evaluate three distinct functional assay methods for assessing hERG channel activity.
• To compare the performance of these assays using known hERG channel inhibitors.
• To identify the most reliable and efficient assay for drug screening.

Main Methods:

• Development and testing of three functional hERG channel assay methods.
• Evaluation against five known hERG channel inhibitors: dofetilide, terfenadine, sertindole, astemizole, and cisapride.
• Comparison of DiBAC4(3)-based assays, membrane potential dye assays, and nonradioactive Rb+ efflux assays.

Main Results:

• DiBAC4(3)-based assays were economical but yielded high false-hit rates due to dye-compound interactions.
• Membrane potential dye assays had fewer color-quenching issues but were expensive and still produced false hits.
• The nonradioactive Rb+ efflux assay demonstrated the highest sensitivity and the lowest false-hit rate.

Conclusions:

• The nonradioactive Rb+ efflux assay is the most sensitive and reliable method for hERG channel inhibition screening.
• This assay minimizes false positives, improving the accuracy of drug safety assessments.
• The findings support the adoption of the Rb+ efflux assay for robust preclinical cardiotoxicity evaluation.