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The Brugada Syndrome (BrS) is characterized by ST-segment elevation in the right precordial leads and is associated with an increased risk of sudden cardiac death. The disorder was initially described as a monogenic primary cardiac electrical disease. However, mutations in SCN5A, encoding the cardiac sodium channel (NaV1.5), which is the major gene associated with the disorder are found in only around 20% of cases and are associated with low penetrance. Furthermore many cases did not display familial aggregation. Based on a previous GWAS conducted on 312 BrS patients and the discovery of the unexpected strong effect of 3 common variants, we proposed that the BrS may comprise a more complex inheritance model. We conducted a genome-wide association study on 2820 individuals with BrS and 10001 ancestry-matched controls to uncover additional genetic loci that modulate susceptibility to BrS, to characterize further the BrS genetic architecture and to uncover new molecular mechanisms. We identified 21 susceptibility variants that passed the genome-wide statistical significance threshold (P We identified 18 new susceptibility variants associated with BrS and uncovered a new pathophysiological molecular mechanism underlying BrS susceptibility. We provided further support for a complex genetic architecture underlying susceptibility for the disorder. Funding Acknowledgement Type of funding source: Public Institution(s). Main funding source(s): H2020 - Marie Sklodowska Curie IF grant, Rising star grant from the Pays de la Loire regional council

Background Genetics of sinus node dysfunction (SND) remains poorly understood with few genes identified (HCN4, SCN5A, GJA5, KCNQ1, MYH6 and ANK2) and further genetic heterogeneity. Purpose We report two families with SND segregating with an intergenic deletion associated with long-range cis-regulatory elements (CREs) of PITX2. Methods We applied 30x whole genome sequencing (WGS) to two French families in which exome sequencing did not allow to detect the causing genes of SND and analyzed them bioinformatically. Results We first applied WGS to a 4-generation family presenting 16 patients with SND, atrial fibrillation and/or repolarization abnormalities and identified a 15-Kb deletion within a 16.5-cM linkage interval (Zmax = 5.9, θ = 0) on chromosome 4q25. We also applied WGS on a second family presenting 5 patients with SND and identified a 91-Kb deletion overlapping the initial deletion. This intergenic deletion, which was located between PITX2 and ANK2 and contained a binding motif of CCCTC-binding factor (CTCF), was segregating in all patients and was not found in 855 French control genomes. CTCF functions as a genome organizer mediating genomic interactions between genes and CREs. To clarify a possible regulatory function of the 1.5-Mb intergenic region containing the deletion, we interrogated 3 epigenetic databases. High-throughput chromosome conformation capture (Hi-C) (1) revealed that this intergenic region was placed in a topologically associating domain (TAD) containing PITX2. Chromatin interaction analysis by paired-end tag (ChIA-PET) of CTCF (2) suggested several possible functional chromatin loops in the TAD. Of them, a CTCF-mediated chromatin loop corresponding to a 300-Kb genomic region was clarified as a possible CRE using histone modification data from Roadmap. The region ranging between the intra-deletion CTCF motif and another distal motif was repressed in H3K27me3 profiles of fetal heart, while the region was activated in H3K27ac profiles of H1 ESC-derived mesendoderm. Conclusion This CRE may regulate expression of PITX2 and/or ANK2 in pacemaker cell differentiation. Ongoing experiments on patient's iPSC and transgenic mouse will further characterize the disease mechanism.

[Background] Early life is a period of drastic epigenetic remodeling in which the epigenome is especially sensitive to extrinsic and intrinsic influence. However, the epigenome-wide dynamics of the DNA methylation changes that occur during this period have not been sufficiently characterized in longitudinal studies., [Methods] To this end, we studied the DNA methylation status of more than 750,000 CpG sites using Illumina MethylationEPIC arrays on 33 paired blood samples from 11 subjects at birth and at 5 and 10 years of age, then characterized the chromatin context associated with these loci by integrating our data with histone, chromatin-state and enhancer-element external datasets, and, finally, validated our results through bisulfite pyrosequencing in two independent longitudinal cohorts of 18 additional subjects., [Results] We found abundant DNA methylation changes (110,726 CpG sites) during the first lustrum of life, while far fewer alterations were observed in the subsequent 5 years (460 CpG sites). However, our analysis revealed persistent DNA methylation changes at 240 CpG sites, indicating that there are genomic locations of considerable epigenetic change beyond immediate birth. The chromatin context of hypermethylation changes was associated with repressive genomic locations and genes with developmental and cell signaling functions, while hypomethylation changes were linked to enhancer regions and genes with immunological and mRNA and protein metabolism functions. Significantly, our results show that genes that suffer simultaneous hyper- and hypomethylation are functionally distinct from exclusively hyper- or hypomethylated genes, and that enhancer-associated methylation is different in hyper- and hypomethylation scenarios, with hypomethylation being more associated to epigenetic changes at blood tissue-specific enhancer elements., [Conclusions] These data show that epigenetic remodeling is dramatically reduced after the first 5 years of life. However, there are certain loci which continue to manifest DNA methylation changes, pointing towards a possible functionality beyond early development. Furthermore, our results deepen the understanding of the genomic context associated to hyper- or hypomethylation alterations during time, suggesting that hypomethylation of blood tissue-specific enhancer elements could be of importance in the establishment of functional states in blood tissue during early-life., This work has been fnancially supported by: Ministerio de Ciencia, Innovación y Universidades (Grant Numbers PI14/01781 and PI11/00144), Instituto de Salud Carlos III and cofnanced by the European Regional Development Fund (to EL); the Plan Nacional de I+D+I 2013–2016/FEDER (PI15/00892 to MFF and AFF); IUOPA-ISPA-FINBA (to RGU, RFP and PS); AF Fernández is supported by a Miguel Servet II fellowship (contract CPII16/00007); JR Tejedor is supported by the Ministry of Economy and Competitiveness through a Juan de la Cierva postdoctoral fellowship (FJCI-2015-26965); the IUOPA is supported by the Obra Social Cajastur-Liberbank, Spain.

Obesity is a disorder that develops from the interaction between genotype and environment involving social, behavioral, cultural, and physiological factors. Obesity increases the risk for type 2 diabetes mellitus, hypertension, cardiovascular disease, cancer, musculoskeletal disorders, chronic kidney and pulmonary disease. Although obesity is clearly associated with an increased prevalence of hypertension, many obese individuals may not develop hypertension. Protecting factors may exist and it is important to understand why obesity is not always related to hypertension. The aim of this review is to highlight the knowledge gap for the association between obesity, hypertension, and potential genetic and racial differences or environmental factors that may protect obese patients against the development of hypertension and other co-morbidities. Specific mutations in the leptin and the melaninocortin receptor genes in animal models of obesity without hypertension, the actions of α-melanocyte stimulating hormone, and SNS activity in obesity-related hypertension may promote recognition of protective and promoting factors for hypertension in obesity. Furthermore, gene-environment interactions may have the potential to modify gene expression and epigenetic mechanisms could also contribute to the heritability of obesity-induced hypertension. Finally, differences in nutrition, gut microbiota, exposure to sun light and exercise may play an important role in the presence or absence of hypertension in obesity.

[Background] Maternal smoking during pregnancy is a major risk factor for adverse health outcomes. The main objective of the study was to assess the impact of in utero tobacco exposure on DNA methylation in children born at term with appropriate weight at birth., [Methods] Twenty mother-newborn dyads, after uncomplicated pregnancies, in the absence of perinatal illness were included. All mothers were healthy with no cardiovascular risk factors, except for the associated risks among those mothers who smoked. Umbilical cord blood and maternal peripheral venous blood were collected and an epigenome-wide association study was performed using a 450 K epigenome-wide scan (Illumina Infinium HumanMethylation 450BeadChip) with adjustment to normalize the DNA methylation for data cell variability in whole blood., [Results] The maternal plasmatic cotinine levels ranged from 10.70-115.40 ng/ml in the exposed group to 0-0.59 ng/ml in the non-exposed group. After adjusting for multiple comparisons in 427102 probes, statistically significant differences for 31 CpG sites, associated to 25 genes were observed. There was a greater than expected proportion of statistically-significant loci located in CpG islands (Fisher’s exact test, p = 0.029) and of those CpG islands, 90.3% exhibit higher methylation levels in the exposed group. The most striking and significant CpG site, cg05727225, is located in the chromosome 11p15.4, within the adrenomedullin gene., [Conclusions] In utero tobacco exposure, even in the absence of fetal growth restriction, may alter the epigenome, contributing to global DNA hypomethylation. Therefore, DNA status can be used as a biomarker of prenatal insults. Considering the possibility to reverse epigenetic modifications, a window of opportunity exists to change the programmed chronic disease., The study was partially funded by grant number PI11/00144, Instituto de Salud Carlos III, Spain and CIBER Fisiopatología Obesidad y Nutrición (CB06/03), Instituto de Salud Carlos III, Spain.

2527 Background: Histone deacetylase inhibitors (HDIs) are epigenetic therapies in development. To exploit the unique activity in impairing DNA repair, HDIs have been combined with chemotherapy. Belinostat is a potent HDI combined with Cis and Etop based on enhanced DNA damage and apoptosis in small cell lung cancer (SCLC) cells. Methods: Patients with relapsed/refractory cancer or previously untreated advanced stage SCLC were eligible. Belinostat was administered by continuous infusion (CIV) over 48h, from 400 mg/m2/24h, in cohorts of 3. Cis was administered on day 1 and Etop daily X3. Belinostat pharmacokinetics (PK) and several pharmacodynamic (PD) measures were assessed, including lysine acetylation in peripheral blood mononuclear cells (PBMCs) and γH2Ax staining in PBMCs and in hair follicles. Results: Five dose levels were explored in 20 patients with solid tumors, including 5 patients with SCLC, two who had no prior therapy. At the first dose level, dose-limiting toxicities (DLT) of gr 4 ANC in 1, and gr 3 HTN in 1 were observed. Cis and Etop were reduced to 60 mg/m2 and 80 mg/m2, respectively, and the dose level repeated without DLT. At the next dose level, 800 mg/m2/24h belinostat, grade 3 HTN and grade 4 pneumonitis were observed. At the MTD of 600 mg/m2/24h belinostat, DLT was seen in 1 of 6 pts; however, all 6 pts required later dose reductions. We thus considered 500 mg/m2/24h in combination with Cis and Etop to be the recommended Phase II dose; confirmation ongoing. PKs show belinostat levels at 1 uM over the 48h infusion, decreasing rapidly to the 60h timepoint. In total 11 pts, 3 with SCLC, completed 6 cycles. PR was seen in 6 pts (3 with SCLC). PD studies confirmed γH2AX staining in PBMCs and hair follicles, peaking at 36h and 60h, respectively. Tubulin and lysine acetylation (Ac-K) in PBMCs peaked at 36h; Ac-K recovered more rapidly than tubulin, mirroring γH2AX. Conclusions: The MTD of belinostat over 48h by CIV was 600 mg/m2/24h, in combination with Cis 60 mg/m2 on day 1 and Etop 80 mg/m2 on days 1 - 3. PD endpoints indicate that belinostat is active in promoting both acetylation and DNA damage. The HDI combined with chemotherapy requires dose reduction and likely represents an on-target increase in DNA damage. Clinical trial information: NCT00926640.