Elevated levels of Low-density lipoprotein cholesterol (LDL-C) in plasma represent the foremost risk factor for atherosclerotic cardiovascular diseases (ASCVD), the leading cause of global mortality. Despite the effectiveness of current medications, there is a need for novel lipid-lowering therapies to further diminish ASCVD risk, especially in high-risk patients. Due to the high prevalence of metabolic syndrome, individuals at heightened ASCVD risk increasingly present with co-morbidities such as metabolic dysfunction-associated steatotic liver disease (MASLD). MASLD, for which there is currently no approved treatment, constitutes the primary cause of liver-related mortality and transplantation. Crucially, the pathophysiology of elevated plasma LDL-C and the risk of ASCVD and MASLD are closely intertwined. The ANTIDOTE project aims to identify a novel therapeutic target with beneficial effects, not only on dyslipidemia-related ASCVD, but also on MASLD. To achieve this dual objective, we employ a translational approach integrating, studies in patients cohorts, genomic methodologies, pre-clinical and in vitro experiments. In a broad-scale discovery genetic study, we identified a candidate gene in the 17q25 locus with potential protective effects on dyslipidemia-related ASCVD and MASLD. This gene is involved in lipid biosynthesis and in the regulation of mitochondrial dynamics and function. However, its role in cardiometabolic diseases has not been assessed thus far. Based on genetic evidences, along with the established link between mitochondrial function and cardiometabolic diseases, ANTIDOTE aims to evaluate this newly identified gene as a promising therapeutic target by: i) Refining genetic variants in the 17q25 locus and their associated phenotypic correlations in humans ii) Assessing the effects of deleting this candidate gene in murine models of cardiometabolic diseases iii) Defining the biological mechanisms impacted in advanced in vitro models.

More than 1,000,000 pacemakers have been implanted worldwide in 2009. Progressive Cardiac Conduction Disease (PCCD), also called Lenègre or Lev disease, represents the major cause of pacemaker implantation in developed countries. It is a slowly evolving lifespan disease that progressively affects cardiac conduction, leading ultimately to pacemaker implantation to prevent the risk of complete atrioventricular block and Stokes-Adams syncope. There is no available pharmacological treatment to prevent the progression of the disease. There is thus a need to understand perfectly the pathophysiological mechanisms of the disease and to identify molecular targets involved in its early phases. There is also a need to identify prognostic biomarkers because patients at risk of developing high degree atrioventricular block are uneasy to identify. We previously identified SCN5A as the first gene associated to PCCD and shown that mutations lead to a loss of function of SCN5A gene product, the cardiac sodium channel Nav1.5. To understand the pathophysiology of PCCD, we studied a Scn5a heterozygous knockout (Scn5a+/-) mouse model and showed that this model exhibits an age-related deterioration in conduction due to progressive occurrence of fibrosis. Scn5a+/- mice represent a unique model to elucidate PCCD pathophysiological mechanisms and identify molecular targets for developing selective preventive treatments, identify biomarkers for diagnosis and prognosis, and test preventive pharmacological treatments. In this context, the objectives of the PreventPCCD project are to: 1. identify the molecular and cellular mechanisms of fibrotic remodelling in Scn5a+/- mice in order to propose putative specific targets for developing a selective preventive therapy of PCCD; 2. evaluate the potential of Magnetic Resonance Imaging (MRI) to detect development of fibrosis in Scn5a+/- mice in order to propose it as a tool for early detection of fibrosis in PCCD patients; 3. discover early circulating biomarkers of the severity of fibrotic remodelling in Scn5a+/- mice that can be easily evaluated in a clinical context; 4. evaluate the potential of Renin-Angiotensin-Aldosterone System (RAAS) inhibitors to prevent fibrosis development. This project, which combines basic research in cell and mouse models, molecular, biochemical and functional analyses in vitro and in vivo, as well as complementary competences in electrophysiology, investigation of contractile activity, and imaging, should lead to rapid clinical applications. We expect to identify at least one putative target for developing a preventive therapy and one circulating biomarker. We also expect to validate MRI as a PCCD diagnostic tool.

White adipose tissue dysfunction is central to the development of obesity-associated complications such as type 2 diabetes, non-alcoholic fatty liver disease and cardiovascular diseases. Despite the importance of this tissue, little is known about the early molecular determinants contributing to the adipocyte dysfunction during obesity. Our hypothesis is that cholesterol is a major player in adipocyte dysfunction. Our preliminary results in cell and mouse models and in patients with obesity show the involvement of free cholesterol accumulation in the establishment of this dysfunction. They also suggest that cholesterol esterification with monounsaturated fatty acids could preserve adipose homeostasis. Using complementary approaches combining cellular and mouse preclinical models, as well as human samples, we propose to determine: 1/ how cholesterol is harmful to the adipocyte, 2/ the importance of its esterification in the maintenance of adipocyte homeostasis, and 3/ whether induction of its esterification by nutritional approaches preserves adipocyte function during obesity. This project will identify therapeutic targets and provide the proof of concept of the feasibility of a strategy to prevent the metabolic complications of obesity.

Rationale: In western countries, cardiac arrhythmias affect more than 3% of the general population, causing high levels of morbidity and sudden cardiac death. In the heart, several transcription factors are powerful regulators of cardiac cell fate and organogenesis. Less well studied are the roles of transcription factors in physiological functions, such as the functional specialization of the ventricular conduction system or the sequence of ventricular electrical activity. These functions are intimately linked to the proper regulation of more than 100 ion channel subunits. Changes in their expression or function in the context of congenital or acquired heart disease affect action potentials and are likely to be at the root of cardiac arrhythmias and sudden death. Objective: To understand role of Iroquois transcription factors (IRX) in the control of generation and propagation of cardiac electrical impulse. Methods: To test our hypothesis, we will take advantage of a newly described rare congenital disorder, the Hamamy syndrome. Importantly, Hamamy patients carry a mutation in IRX5 gene and are characterized by both cardiac structural and electrophysiological defects, suggesting that the origin of some life-threatening arrhythmias, that usually develop as a patient ages, lies in cardiac structural and electrophysiological development. Therefore, studying the pathophysiological mechanism of this disease and the role of IRX5 may bring information to the mechanism of arrhythmias in the adult. Studies have also suggested a role for IRX5 in another familial and more frequent arrhythmic disease, the Brugada syndrome, as it is associated with defects in ventricular repolarizing gradient which regulation is ensured by Irx5 in the mouse. However, this role has not been proven yet. To do so, we designed a multidisciplinary experimental plan using biochemical, molecular and in vivo analyses with the following specific aims: - Identifying the role of IRX5 transcription factor in human heart formation and function, through Hamamy syndrome modeling using iPS cells and transgenic mouse model. - Exploring the role of IRX5 in the pathophysiology of Brugada syndrome using iPS cells. Expected results: With this research project, using a combination of newly generated patient-derived iPS cells and transgenic mouse lines we will elucidate the functional role of IRX5 transcription factor in human cardiac physiology, and decipher its role in the abnormal cardiac phenotype of the Hamamy and Brugada syndromes.

Voltage-gated Na+ (Nav) channels are key determinants of myocardial excitability and defects in Nav channel functioning or regulation, associated with inherited and acquired cardiac disease, increase the risk of life-threatening arrhythmias. In heart failure, the inactivation gating properties of the main cardiac Nav channels, the Nav1.5 channels, are altered, resulting in decreased channel availability and increased late Na+ current. Among the major determinants suggested to cause these defects are the activation of specific kinases, primarily the Ca2+/Calmodulin Kinase II (CaMKII), and the direct phosphorylation of the Nav1.5 channel pore-forming subunit. Mass spectrometry (MS)-based phosphoproteomic analyses in the laboratory were undertaken to identify in situ the native phosphorylation sites on the Nav1.5 and associated/regulatory proteins purified from mouse cardiac ventricles in both basal (wild-type) and failing (CaMKIIdc-overexpressing) conditions. These experiments led to the identification of several native phosphorylation sites: nineteen were identified in Nav1.5 and one in Fibroblast Growth Factor 13 (FGF13), a key Nav channel associated/regulatory protein involved in channel inactivation. Of these newly-identified phosphorylation sites, interestingly, three in the C-terminus of Nav1.5 abut the binding sites for FGF13 and Ca2+/calmodulin, another key regulator of Nav channel inactivation, and conversely, the FGF13 phosphorylation site is in the Nav channel-binding site. We hypothesize that phosphorylation at one or several of these four sites regulates Nav1.5 channel inactivation gating by altering interaction with FGF13 and/or Ca2+/calmodulin. Specifically, we will test the hypotheses that (1) the regulation of cardiac Nav1.5 channel inactivation by FGF13 and Ca2+/calmodulin in basal physiological conditions depends on the phosphorylation of Nav1.5 at serine-1884 and of FGF13 at serine-218; and (2) the phosphorylation of Nav1.5 by CaMKII at serines-1933 and -1984 in heart failure adversely affects the interaction of Ca2+/calmodulin and FGF13 with the channel and the associated regulation of channel inactivation. By specifying mechanisms whereby phosphorylation regulates cardiac Nav1.5 channel function, our results will link cell signaling to membrane excitability and foster a better understanding of the role of Nav1.5 channels in health and disease, which is needed for improved prevention and treatment of arrhythmias.