Aims SCN5A mutations are associated with various cardiac phenotypes, including long QT syndrome type 3 (LQT3), Brugada syndrome (BrS), and cardiac conduction disease (CCD). Certain mutations, such... Show moreAims SCN5A mutations are associated with various cardiac phenotypes, including long QT syndrome type 3 (LQT3), Brugada syndrome (BrS), and cardiac conduction disease (CCD). Certain mutations, such as SCN5A-1795insD, lead to an overlap syndrome, with patients exhibiting both features of BrS/CCD [decreased sodium current (I-Na)] and LQT3 (increased late I-Na). The sodium channel blocker mexiletine may acutely decrease LQT3-associated late I-Na and chronically increase peak I-Na associated with SCN5A loss-of-function mutations. However, most studies have so far employed heterologous expression systems and high mexiletine concentrations. We here investigated the effects of a therapeutic dose of mexiletine on the mixed phenotype associated with the SCN5A-1795insD mutation in HEK293A cells and human-induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs). Methods and results To assess only the chronic effects on trafficking, HEK293A cells transfected with wild-type (WT) SCN5A or SCN5A-1795insD were incubated for 48 h with 10 & mu;m mexiletine followed by wash-out, which resulted in an increased peak I-Na for both SCN5A-WT and SCN5A-1795insD and an increased late I-Na for SCN5A-1795insD. Acute re-exposure of HEK293A cells to 10 & mu;m mexiletine did not impact on peak I-Na but significantly decreased SCN5A-1795insD late I-Na. Chronic incubation of SCN5A-1795insD hiPSC-CMs with mexiletine followed by wash-out increased peak I-Na, action potential (AP) upstroke velocity, and AP duration. Acute re-exposure did not impact on peak I-Na or AP upstroke velocity, but significantly decreased AP duration. Conclusion These findings demonstrate for the first time the therapeutic benefit of mexiletine in a human cardiomyocyte model of SCN5A overlap syndrome. Show less
The aim of this thesis was to work towards pre-clinical proof-of-concept for NOTCH3 cysteine corrective exon skipping as a rational therapeutic approach for CADASIL. To address all aspects required... Show moreThe aim of this thesis was to work towards pre-clinical proof-of-concept for NOTCH3 cysteine corrective exon skipping as a rational therapeutic approach for CADASIL. To address all aspects required for therapeutic development, the work performed for this thesis included not only in vitro testing of NOTCH3 exon skipping in CADASIL patient derived vascular smooth muscle cells and studies into the function of the cysteine corrected proteins, but also the generation of a relevant humanized in vivo model, pre-clinical biomarker development, and studies defining prevalence, spectrum and characteristics of NOTCH3 mutations worldwide. Show less
