Targeted Enhancement of hERG K+ Channel Activity with scFv Antibody Fragments

Abstract

Cardiac IKr is a repolarizing current comprising subunits encoded by the human ether-a-go-go related gene (hERG). A defining feature of hERG and related channel types is an N-terminal PAS domain that, through its effects on gating, reduces current amplitude. To probe the functional consequences of disrupting this natural channel modulation, we screened for single chain fragment variable (scFv) antibodies against the PAS domain using phage display. We assessed the binding properties to PAS and functional consequences of these antibodies on the hERG channel. Two isolates affecting hERG 1a current properties were shown by GST pull-down assays to interact with different regions of the PAS domain with submicromolar affinities. In HEK-293 cells stably expressing hERG 1a, one antibody slowed inactivation onset and the other accelerated recovery from inactivation. Both significantly increased repolarizing current during a voltage protocol mimicking a ventricular action potential. Because native IKr channels are heteromers of the original hERG 1a isolate and hERG 1b, an isoform from which the PAS domain is transcriptionally omitted, we tested the effect of the scFv antibodies on cardiomyocytes derived from human induced pluripotent stem cells. Similar to the HEK-293 findings, the antibodies diminished channel rectification and significantly increased the repolarizing charge during a ventricular action potential voltage clamp. Here, we demonstrate the versatility of scFv antibodies as a site-specific probe of ion channel function. Given their ability to increase IKr, we propose the PAS-channel interface as a therapeutic target and the use of scFv antibodies as a novel treatment for diseases of excitability where action potential duration is prolonged.