Elsevier

Heart Rhythm

Volume 20, Issue 5, May 2023, Pages 709-717
Heart Rhythm

Experimental
Functional characterization and identification of a therapeutic for a novel SCN5A-F1760C variant causing type 3 long QT syndrome refractory to all guideline-directed therapies

https://doi.org/10.1016/j.hrthm.2023.01.032Get rights and content

Background

Pathogenic variants in the SCN5A-encoded Nav1.5 sodium channel cause type 3 long QT syndrome (LQT3). We present the case of an infant with severe LQT3 who was refractory to multiple pharmacologic therapies as well as bilateral stellate ganglionectomy. The patient’s novel variant, p.F1760C-SCN5A, involves a critical residue of the Nav1.5’s local anesthetic binding domain.

Objective

The purpose of this study was to characterize functionally the p.F1760C-SCN5A variant using TSA-201 and patient-specific induced pluripotent stem cell–derived cardiomyocytes (iPSC-CMs).

Methods

Whole-cell patch clamp was used to assess p.F1760C-SCN5A associated sodium currents with/without lidocaine (Lido), flecainide, and phenytoin (PHT) in TSA-201 cells. p.F1760C-SCN5A and CRISPR-Cas9 variant-corrected isogenic control (IC) iPSC-CMs were generated. FluoVolt voltage dye was used to measure the action potential duration (APD) with/without mexiletine or PHT.

Results

V1/2 of inactivation was right-shifted significantly in F1760C cells (–72.2 ± 0.7 mV) compared to wild-type (WT) cells (–86.3 ± 0.9 mV; P <.0001) resulting in a marked increase in window current. F1760C increased sodium late current 2-fold from 0.18% ± 0.04% of peak in WT to 0.49% ± 0.07% of peak in F1760C (P = .0005). Baseline APD to 90% repolarization (APD90) was increased markedly in F1760C iPSC-CMs (601 ± 4 ms) compared to IC iPSC-CMs (423 ± 15 ms; P <.0001). However, 4-hour treatment with 10 μM mexiletine failed to shorten APD90, and treatment with 5μM PHT significantly decreased APD90 of F1760C iPSC-CMs (453 ± 6 ms; P <.0001).

Conclusion

PHT rescued electrophysiological phenotype and APD of a novel p.F1760C-SCN5A variant. The antiepileptic drug PHT may be an effective alternative therapeutic for the treatment of LQT3, especially for variants that disrupt the Lido/mexiletine binding site.

Introduction

Congenital long QT syndrome (LQTS) is an inherited autosomal genetic heart disease clinically characterized by a prolonged QT interval on 12-lead electrocardiogram.1 Patients with LQTS can present with arrhythmic syncope/seizures, sudden cardiac arrest, or sudden cardiac death often after an abrupt emotional or physical stress trigger.2 LQTS has an estimated prevalence of ∼1 in 2000 individuals and for high-risk subjects who are untreated, the estimated 10-year mortality rate is approximately 50%.3

Gain-of-function mutations in the SCN5A-encoded α-subunit of the Nav1.5 voltage-gated cardiac sodium channel are associated with type 3 long QT syndrome (LQT3), which accounts for approximately 5%–10% of all LQTS cases.4,5 LQT3-associated pathogenic variants in SCN5A can disrupt the fast inactivation of the channel, resulting in an increased late sodium current (INa,L), and/or alter the biophysical property of the channel, causing delayed inactivation and an increase in “window current,” both of which may lead to pathologic prolongation of the cardiac action potential duration (APD).4

Patients with LQT3 often present with resting bradycardia and QT-interval prolongation that is more pronounced during slow heart rates.4 Current clinical management options for patients with LQT3 include drug therapy (β-blockers; sodium channel blockers, particularly intravenous lidocaine [Lido] or oral mexiletine), denervation therapy (left cardiac sympathetic denervation surgery), and/or implantable cardioverter-defibrillator.6 Despite these treatment strategies, the risk of death during cardiac events among LQT3 patients is significantly higher than in patients with either type 1 or type 2 LQTS.7

Sodium channel blockade is a direct pharmacologic approach for the clinical management of LQT3. Various sodium channel blocking agents including mexiletine, Lido, and flecainide (Flec) can attenuate the QT interval.8, 9, 10, 11, 12 However, the efficacy of sodium channel blocker treatment is highly dependent on the location of the mutation.13 These sodium channel inhibitors are thought to bind to a local anesthetic site on Nav1.5 involving amino acids F1760 and Y1767.14,15 Mutations located near this binding site could disrupt the binding of these specific drugs. Phenytoin (PHT), an anticonvulsant drug commonly used to prevent and control seizures, is another sodium channel blocker that historically had been suggested as an alternative therapy for patients with severe LQT3.16

Here we present the case of an infant with severe LQT3 who was refractory to bilateral stellate ganglionectomy and pharmacologic treatments with either Lido or mexiletine. The decedent had a novel variant, p.F1760C, involving a critical residue of the Nav1.5’s local anesthetic (ie, Lido/mexiletine) binding domain.14,15 In the present study, we characterized functionally the novel variant p.F1760C-SCN5A in TSA-201 cells as well as the infant’s specific induced pluripotent stem cell–derived cardiomyocytes (iPSC-CMs). Disease modeling studies were performed alongside CRISPR/Cas9-gene variant-corrected, isogenic control (IC) iPSC-CMs.

Section snippets

Patient sample

A female infant with severe LQT3 who subsequently died at 4 months of age was referred to the Mayo Clinic Windland Smith Rice Sudden Death Genomics Laboratory after genetic testing for LQTS was positive for a novel SCN5A genetic variant (c.5279T>G; p.F1760C-SCN5A). After obtaining informed written consent for this study, which was approved by the Mayo Clinic Institutional Review Board (IRB number 1216-97 and IRB number 09-006465), a blood sample was collected for the generation of

Clinical description

The female infant was diagnosed in utero with documented 3:1 heart block. She was delivered at a site having a level 2 newborn intensive care unit, and she had sudden cardiac arrest shortly after birth. The infant underwent 8 minutes of cardiopulmonary resuscitation and subsequently was transferred to a center with a cardiac surgery department, where she underwent placement of a dual-chamber pacemaker. She experienced intermittent torsades de pointes that usually terminated spontaneously, but

Discussion

We report a now-deceased infant with severe LQT3 who was refractory to bilateral stellate ganglionectomy and pharmacologic treatments. Commercial genetic testing for LQTS identified a novel, de novo p.F1760C-SCN5A variant. Interestingly, p.F1760 is one of the critical amino acid residues that constitute the local anesthetic binding site within the sodium channel pore that is crucial for interaction with Lido.14,15 Mechanistic studies using cystine substitution and methanethiosulfonate reagent

Conclusion

We present the APD shortening effect of PHT in a patient-specific iPSC-CM model of LQT3. This study provides pharmacologic and functional evidence suggesting that PHT may be a therapeutic option for some patients with LQT3, particularly those who are refractory to all guideline-directed conventional therapies.

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  • Funding Sources: This work was supported by the Mayo Clinic Windland Smith Rice Comprehensive Sudden Cardiac Death Program.

    Disclosures: Dr Ackerman is a consultant for Abbott, Boston Scientific, Bristol Myers Squibb, Daiichi Sankyo, Invitae, Medtronic, Tenaya Therapeutics, and UpToDate. Dr Ackerman and Mayo Clinic are involved in an equity/royalty relationship with AliveCor, Anumana, ARMGO Pharma, Pfizer, and Thryv Therapeutics; however, none of these entities have contributed to this study in any manner. All other authors have reported that they have no relationships relevant to the contents of this paper to disclose.

    1

    Ms Marissa J. Stutzman and Mr Xiaozhi Gao contributed equally to this work.

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