In vivo KCNH2-suppression-replacement gene therapy attenuates the pathogenic phenotype in transgenic rabbits with short QT syndrome type 1

Abstract

Abstract Background Short QT syndrome type 1 (SQT1) is a genetic channelopathy characterized by gain-of-function variants in the KCNH2-encoded potassium channel underlying the IKr current, leading to a pathologically shortened cardiac action potential duration (APD). Patients with SQT1 manifest shortening of the QT duration on 12-lead ECG and are prone to cardiac arrhythmias that can culminate in sudden cardiac death. Purpose To investigate the effect of a KCNH2-specific suppression-replacement (KCNH2-SupRep) gene therapy on the pathogenic phenotype in transgenic SQT1 rabbits. Methods A novel dual-component KCNH2-SupRep gene therapy was developed by combining a custom-designed KCNH2 shRNA (suppression) and an shRNA-immune (shIMM) KCNH2 cDNA (replacement) into AAV9. In vivo AAV9-mediated KCNH2-SupRep / sham treatment was performed by intra-aortic root injection of the KCNH2-SupRep AAV9 (1E10 vg/kg) / NaCl 0.9% during balloon occlusion using a Swan Ganz catheter. Three weeks after administration, 12-lead ECGs were assessed in adult transgenic SQT1 (KCNH2-N588K, gain of IKr due to impaired inactivation) and wild-type (WT) rabbits to determine the effect of KCNH2-SupRep / sham treatment on the rabbit’s QTc duration and QT index. Patch-clamp recordings were performed in isolated ventricular cardiomyocytes (CMs) to evaluate the effect of KCNH2-SupRep / sham treatment on the CM’s action potential duration at 90% repolarization (APD90). Results After in vivo AAV9-mediated KCNH2-SupRep gene therapy injection (1E10 vg/kg) in 7 SQT1 rabbits, QTc duration and QT index significantly increased in all SQT1 rabbits (QTc: untreated(UT)-SQT1: 153.2±3.8ms vs treated-SQT1: 166.5±4.6ms, p=0.0002, N=7 each; QTi: UT-SQT1: 92.5±1.9% vs treated-SQT1: 99.7±2.7%, p=0.0007, N=7 each), reaching a QT index of ~100% typically observed in WT rabbits. No such changes were observed in the sham-SQT1 rabbits. Furthermore, cardiomyocytes isolated from 7 SQT1 rabbits that received in vivo SupRep gene therapy demonstrated APD90 prolongation closer to WT rabbits compared to UT-SQT1 and sham-SQT1 rabbits (APD90: UT-SQT1: 314.2±11.9ms, WT: 404.5±13.8ms, treated-SQT1: 362.9±10.6ms; UT-SQT1 vs WT: p<0.0001; UT-SQT1 vs treated-SQT1: p=0.019; WT vs Treated-SQT1: p=0.031). In line with these observations, first IKr measurements demonstrate smaller IKr currents in SupRep-SQT1 than in sham-SQT1 CMs. Conclusion In vivo KCNH2-SupRep gene therapy normalizes the pathologically shortened QTc duration and QT index in transgenic SQT1 rabbits. Moreover, KCNH2-SupRep treatment prolongs APD90 in SQT1 compared to UT-SQT1 and sham-SQT1 reaching levels closer to WT.