IS IT POSSIBLE TO MODEL JERVELL AND LANGE-NIELSEN SYNDROME (JLNS) USING PATIENT-SPECIFIC INDUCED PLURIPOTENT STEM CELLS (IPSC)?

Abstract

<h3>Background</h3> JLNS is a rare channelopathy caused by mutations in the KCNQ1 and KCNE1 genes that encodes the α and β subunits of the IKS current channel. Patients with JLNS have congenital bilateral deafness and prolongation of the QT interval on electrocardiogram (ECG), which can lead to arrhythmias, syncope, and a high risk of sudden death. Induced pluripotent stem cells-derived cardiomyocytes (iPSC-CM) can be a useful tool to recapitulate the disease phenotype in vitro enabling the comprehension of its molecular and cellular mechanisms. <h3>Methods</h3> The DNA of a proband with clinical symptoms of JVLNS was analyzed by next-generation sequencing. Sanger was done on samples from family members. iPSC were generated from the proband (Pac6), a cousin (Pac53) suspect of JLNS, the grandfather (Pac25), his sister (Pac24), and his mother (Pac8). iPSC were evaluated for euploidy and characterized for pluripotency by PCR, flow cytometry, and immunohistochemistry. iPSC were differentiated into CMs and its efficiency was assessed by flow cytometry. Electrophysiology was evaluated by intracellular microelectrode (PA) and microelectrode array (MEA). <h3>Results</h3> A homozygous variant c.1394-2A>G was identified in the KCNQ1 gene of Pac6 and Pac53. Both have congenital bilateral deafness and a QTc interval of 670 and 660ms respectively. Pac25 and Pac24 have no mutation and Pac8 have the variant in heterozygosis (Long QT syndrome type 1 - LQTS1). iPSC showed pluripotency markers in PCR and Flow Cytometry (OCT3/4, SOX2, NANOG, etc) and the capacity to differentiate in 3 embryonic layers (shown by immunohistochemistry and PCR). iPSC karyotype was normal, except for Pac53, in which there‘s a translocation between chromosomes 14 and 22. The iPSC-CM had differentiation efficiency above 70%. Surprisingly, electrophysiology functional analysis of action potential duration at 90% of repolarization (APD90) showed no significant differences between Pac6 (APD90 = 255,67 ± 39,19) and Pac53 (APD90 = 315,45 ± 74,71) compared to Pac24 (APD90 = 262,22 ± 62,81) and Pac25 (APD90 = 215,43 ± 73,83). Similar results were observed when analyzed by MEA. <h3>Conclusion</h3> This work is unique to modeling a non-described mutation in the KCNQ1 gene present in a family with two JLNS and one asymptomatic LQTS1. The complete understanding of why the CM-iPSC failed to mimetics in vitro disease demands further investigation challenging the cells with pro-arrhythmic drugs and iKS inhibitors.