Transcriptional Dysregulation Underlies Both Monogenic Arrhythmia Syndrome and Common Modifiers of Cardiac Repolarization.

BACKGROUND: Brugada syndrome (BrS) is an inherited arrhythmia syndrome caused by loss-of-function variants in the cardiac sodium channel gene SCN5A (sodium voltage-gated channel alpha subunit 5) in ≈20% of subjects. We identified a family with 4 individuals diagnosed with BrS harboring the rare G145R missense variant in the cardiac transcription factor TBX5 (T-box transcription factor 5) and no SCN5A variant. METHODS: We generated induced pluripotent stem cells (iPSCs) from 2 members of a family carrying TBX5-G145R and diagnosed with Brugada syndrome. After differentiation to iPSC-derived cardiomyocytes (iPSC-CMs), electrophysiologic characteristics were assessed by voltage- and current-clamp experiments (n=9 to 21 cells per group) and transcriptional differences by RNA sequencing (n=3 samples per group), and compared with iPSC-CMs in which G145R was corrected by CRISPR/Cas9 approaches. The role of platelet-derived growth factor (PDGF)/phosphoinositide 3-kinase (PI3K) pathway was elucidated by small molecule perturbation. The rate-corrected QT (QTc) interval association with serum PDGF was tested in the Framingham Heart Study cohort (n=1893 individuals). RESULTS: TBX5-G145R reduced transcriptional activity and caused multiple electrophysiologic abnormalities, including decreased peak and enhanced "late" cardiac sodium current (INa), which were entirely corrected by editing G145R to wild-type. Transcriptional profiling and functional assays in genome-unedited and -edited iPSC-CMs showed direct SCN5A down-regulation caused decreased peak INa, and that reduced PDGF receptor (PDGFRA [platelet-derived growth factor receptor α]) expression and blunted signal transduction to PI3K was implicated in enhanced late INa. Tbx5 regulation of the PDGF axis increased arrhythmia risk due to disruption of PDGF signaling and was conserved in murine model systems. PDGF receptor blockade markedly prolonged normal iPSC-CM action potentials and plasma levels of PDGF in the Framingham Heart Study were inversely correlated with the QTc interval (P<0.001). CONCLUSIONS: These results not only establish decreased SCN5A transcription by the TBX5 variant as a cause of BrS, but also reveal a new general transcriptional mechanism of arrhythmogenesis of enhanced late sodium current caused by reduced PDGF receptor-mediated PI3K signaling.

Cardiomyopathy with lethal arrhythmias associated with inactivation of KLHL24.

Hypertrophic cardiomyopathy (HCM) is the most common inherited cardiovascular disorder, yet the genetic cause of up to 50% of cases remains unknown. Here, we show that mutations in KLHL24 cause HCM in humans. Using genome-wide linkage analysis and exome sequencing, we identified homozygous mutations in KLHL24 in two consanguineous families with HCM. Of the 11 young affected adults identified, 3 died suddenly and 1 had a cardiac transplant due to heart failure. KLHL24 is a member of the Kelch-like protein family, which acts as substrate-specific adaptors to Cullin E3 ubiquitin ligases. Endomyocardial and skeletal muscle biopsies from affected individuals of both families demonstrated characteristic alterations, including accumulation of desmin intermediate filaments. Knock-down of the zebrafish homologue klhl24a results in heart defects similar to that described for other HCM-linked genes providing additional support for KLHL24 as a HCM-associated gene. Our findings reveal a crucial role for KLHL24 in cardiac development and function.

Coagulation Gene Expression Profiling in Infants With Necrotizing Enterocolitis.

OBJECTIVES: Coagulopathy and mesenteric thrombosis are common in premature neonates with necrotizing enterocolitis (NEC). This pilot study aimed to investigate the hypothesis that there are changes in the gene expression related to the coagulation and anticoagulation systems in NEC. METHODS: Consecutive neonates (n = 11) with NEC (Bell stages 2-3) were recruited. Two comparison groups, matched for birth weight and corrected gestational age, were selected based on the absence of inflammation and coagulopathy (healthy control, n = 10), or the presence of a confirmed blood infection (sepsis control, n = 12). A pathway-specific quantitative polymerase chain reaction array was used to determine the expression of 94 genes involved in human blood coagulation and anticoagulation cascade. RESULTS: Twelve genes of the coagulation and anticoagulation systems were significantly altered in the patients with NEC compared with healthy controls. In particular, neutrophil elastase, CD63, PROS1, HGF, and F12 were significantly upregulated (mean fold changes [FCs] +2.74, P < 0.05) with an overall procoagulant effect; MFGE8, factor II (thrombin) receptor-like 1 (F2RL1), FGL2, PLAT, PROCR, SERPIND1, and HNF4A were significantly downregulated (mean FCs -2.45, P < 0.05) with a reduction in fibrinolysis and endothelial regeneration. In the comparison between NEC and sepsis, we did observe a significant difference in expression of F2RL1 (FC -2.50, P = 0.01). CONCLUSIONS: We have identified potential biomarkers associated with coagulopathy and disease progression in NEC. In particular, the overall procoagulant status, at the transcriptional level, should be further investigated to unveil molecular mechanisms leading to intestinal necrosis, multiorgan failure, and death.

The genetics of pro-arrhythmic adverse drug reactions.

Ventricular arrhythmia induced by drugs (pro-arrythmia) is an uncommon event, whose occurrence is unpredictable but potentially fatal. The ability of a variety of medications to induce these arrhythmias is a significant problem facing the pharmaceutical industry. Genetic variants have been shown to play a role in adverse events and are also known to influence an individual's optimal drug dose. This review provides an overview of the current understanding of the role of genetic variants in modulating the risk of drug induced arrhythmias.

Digenic inheritance of mutations in the cardiac troponin (TNNT2) and cardiac beta myosin heavy chain (MYH7) as the cause of severe dilated cardiomyopathy.

Familial dilated cardiomyopathy (DCM) is characterized by ventricular dilation and depressed myocardial performance. It is a genetically heterogeneous disorder associated with mutations in over 60 genes. We carried out whole exome sequencing in combination with cardiomyopathy-related gene-filtering on two affected family members to identify the possible causative mutation in a consanguineous Iranian family with DCM. Two novel variants in cardiomyopathy-related genes were identified: c.247 A > C; p.N83H in the Troponin T Type 2 gene (TNNT2) and c.2863G > A; p.D955N in the Myosin Heavy Polypeptide 7 gene (MYH7). Sanger sequencing and co-segregation analysis in the remaining family members supported the coexistence of these digenic mutations in affected members of the family. Carriers of either variant alone were asymptomatic. In summary, we find that digenic inheritance of two novel variants in DCM related genes is associated with a severe form of DCM. Exome sequencing has been shown to be very useful in identifying pathogenic mutations in cardiomyopathy families, and this report emphasizes the importance of comprehensive screening of DCM related genes, even after the identification of a single disease-causing mutation.

Role of common and rare variants in SCN10A: results from the Brugada syndrome QRS locus gene discovery collaborative study.

AIMS: Brugada syndrome (BrS) remains genetically heterogeneous and is associated with slowed cardiac conduction. We aimed to identify genetic variation in BrS cases at loci associated with QRS duration. METHODS AND RESULTS: A multi-centre study sequenced seven candidate genes (SCN10A, HAND1, PLN, CASQ2, TKT, TBX3, and TBX5) in 156 Caucasian SCN5A mutation-negative BrS patients (80% male; mean age 48) with symptoms (64%) and/or a family history of sudden death (47%) or BrS (18%). Forty-nine variants were identified: 18 were rare (MAF <1%) and non-synonymous; and 11/18 (61.1%), mostly in SCN10A, were predicted as pathogenic using multiple bioinformatics tools. Allele frequencies were compared with the Exome Sequencing and UK10K Projects. SKAT methods tested rare variation in SCN10A finding no statistically significant difference between cases and controls. Co-segregation analysis was possible for four of seven probands carrying a novel pathogenic variant. Only one pedigree (I671V/G1299A in SCN10A) showed co-segregation. The SCN10A SNP V1073 was, however, associated strongly with BrS [66.9 vs. 40.1% (UK10K) OR (95% CI) = 3.02 (2.35-3.87), P = 8.07 × 10-19]. Voltage-clamp experiments for NaV1.8 were performed for SCN10A common variants V1073, A1073, and rare variants of interest: A200V and I671V. V1073, A200V and I671V, demonstrated significant reductions in peak INa compared with ancestral allele A1073 (rs6795970). CONCLUSION: Rare variants in the screened QRS-associated genes (including SCN10A) are not responsible for a significant proportion of SCN5A mutation negative BrS. The common SNP SCN10A V1073 was strongly associated with BrS and demonstrated loss of NaV1.8 function, as did rare variants in isolated patients.