High-throughput phenotyping of heteromeric human ether-à-go-go-related gene potassium channel variants can discriminate pathogenic from rare benign variants

Chai-Ann Ng; Matthew D Perry; Whitney Liang; Nicola J Smith; Brian Foo; Alvin Shrier; Gergely L Lukacs; Adam P Hill; Jamie I Vandenberg

doi:10.1016/j.hrthm.2019.09.020

High-throughput phenotyping of heteromeric human ether-à-go-go-related gene potassium channel variants can discriminate pathogenic from rare benign variants

Heart Rhythm. 2020 Mar;17(3):492-500. doi: 10.1016/j.hrthm.2019.09.020. Epub 2019 Sep 23.

Authors

Chai-Ann Ng¹, Matthew D Perry¹, Whitney Liang², Nicola J Smith¹, Brian Foo³, Alvin Shrier³, Gergely L Lukacs⁴, Adam P Hill¹, Jamie I Vandenberg⁵

Affiliations

¹ Molecular Cardiology and Biophysics Division, Victor Chang Cardiac Research Institute, Darlinghurst, New South Wales, Australia; St Vincent's Clinical School, UNSW Sydney, Darlinghurst, New South Wales, Australia.
² Molecular Cardiology and Biophysics Division, Victor Chang Cardiac Research Institute, Darlinghurst, New South Wales, Australia.
³ Department of Physiology, McGill University, Montreal, Quebec, Canada.
⁴ Department of Physiology, McGill University, Montreal, Quebec, Canada; Department of Biochemistry, McGill University, Montreal, Quebec, Canada.
⁵ Molecular Cardiology and Biophysics Division, Victor Chang Cardiac Research Institute, Darlinghurst, New South Wales, Australia; St Vincent's Clinical School, UNSW Sydney, Darlinghurst, New South Wales, Australia. Electronic address: j.vandenberg@victorchang.edu.au.

PMID: 31557540
DOI: 10.1016/j.hrthm.2019.09.020

Abstract

Background: KCNH2 encodes the human ether-à-go-go-related gene potassium channel, which passes the rapid delayed rectifier potassium current. Loss-of-function variants in KCNH2 cause long QT syndrome type 2, which is associated with a markedly increased risk of cardiac arrhythmias. The majority of rare KCNH2 variants, however, are likely to be benign.

Objective: The purpose of this study was to develop a high-throughput assay for discriminating pathogenic from benign KCNH2 variants.

Methods: Nonsynonymous homozygous KCNH2 variants stably expressed in Flp-In human embryonic kidney 293 cell lines were phenotyped using an automated patch-clamp platform and a cell surface enzyme-linked immunosorbent assay. Functional phenotyping of heterozygous KCNH2 variants stably expressed in Flp-In human embryonic kidney 293 cell lines using a bicistronic vector was performed using an automated patch-clamp platform.

Results: In homozygous KCNH2 variant cell lines, discrepancies between current density and cell surface expression levels measured using an enzyme-linked immunosorbent assay can be explained by changes in gating properties of the variant channels. For the 30 heterozygous KCNH2 variant cell lines studied, the assay correctly predicted the ClinVar ascribed classification for 17/17 pathogenic/likely pathogenic/benign variants. Of the 13 pore-domain variants studied, 11 had a dominant-negative expression defect while the remaining 2 had enhanced inactivation gating, resulting in a dominant-negative phenotype.

Conclusion: High-throughput electrophysiological phenotyping of heterozygous KCNH2 variants can accurately distinguish between dominant-negative, haploinsufficient loss-of-function, and benign variants. This assay will help with future classification of KCNH2 variants.

Keywords: Automated patch-clamp; Electrophysiology; Ion channels; Kv11.1; Long QT syndrome; Phenotyping; hERG.

Publication types

Research Support, Non-U.S. Gov't

MeSH terms

Arrhythmias, Cardiac / genetics*
Arrhythmias, Cardiac / metabolism
Arrhythmias, Cardiac / pathology
DNA / genetics*
DNA / metabolism
DNA Mutational Analysis
Enzyme-Linked Immunosorbent Assay
Ether-A-Go-Go Potassium Channels / genetics*
Ether-A-Go-Go Potassium Channels / metabolism
HEK293 Cells
Humans
Mutation, Missense*
Patch-Clamp Techniques
Phenotype

Substances

Ether-A-Go-Go Potassium Channels
DNA