Vascular remodeling in patients with rare genetic disorders

National Heart, Lung, And Blood Institute

Investigators

Linked publications & trials

Abstract

Monogenetic disorders grant the unique ability to understand more complex diseases due to a single defined genetic defect. Advanced sequencing technology accelerates gene candidate discovery but progress can be slowed by the inability to distinguish between functional disease related and non-disease related mutations. We use patient-specific in vitro disease modeling systems combined with genetic tools to identify disease related mutation, to study gene related disease mechanism and to perform drug screening. Further, comprehensive clinical evaluation of patients with these conditions further help elucidate the disease mechanism and the organ systems affected. Vascular calcification is a secondary complication to diseases such as atherosclerosis, diabetes mellitus type II and chronic kidney disease but its underlying mechanism is poorly understood. ACDC is a rare genetic disease, in which de novo vascular calcifications form in the lower extremity arteries and peri-articular calcifications in the joints of affected adults and is caused by mutations in the 5'-nucleotidase Ecto (NT5E) gene encode for CD73, an enzyme involved in the extracellular purine metabolic pathway. Cultured fibroblasts isolated from patients had complete CD73 enzyme activity loss and cells displayed a compensatory increased tissue non-specific alkaline phosphatase (TNAP) activity, driving increased calcification. Genetic rescue as well as treatment with adenosine reversed this effect, suggesting adenosine signaling in normal tissue inhibits ectopic vessel calcification. Clinical and radiological follow up of our ACDC patients helped characterize a novel form of inflammatory arthritis. We also used biopsied tissues to determine the crystal composition in joint peri-articular and vascular calcifications by microscopy/micro tomography. Further, our small non-randomized treatment protocol in seven patients with ACDC with etidronate was completed in 2021. We tested effectiveness of etidronate in attenuating the progression of lower extremity arterial and peri-articular calcification and vascular blood flow. Study results showed that etidronate treatment did not significantly change these parameters over time but may have slowed the rate disease progression. While on treatment, patients tolerated the medication well with no significant side effects. Autosomal dominant hyper-IgE syndrome (AD-HIES; Jobs syndrome) is a rare immunodeficiency due to mutations in the STAT3 gene. Patients suffer from multiple life-threatening infections starting during childhood as well as multiple abnormalities outside the immune system, such as aberrant healing. Vascular abnormalities include arterial tortuosity and abnormal dilatation and aneurysms of medium sized arteries, potentially leading to myocardial infarction and subarachnoid hemorrhage. In a wound healing assay, we found delayed granulation tissue formation and vascularization in AD-HIES patients compared to healthy subjects. Further, RNA-Seq analysis of patient cells identified deficiencies in angiogenesis, extracellular matrix metabolism, and wound healing signaling pathways mediated by dysregulation of HIF1 signaling. Coronary arteries of AD-HIES patients show abnormal extracellular matrix organization and decreased HIF1 expression. Reduced HIF1 expression was further functionally verified in cell culture and mouse models of angiogenesis: cell culture vessel tube formation assay, teratoma growth assay, a mouse hind-limb ischemia and skin wound healing models. Autoinflammatory diseases, such as SAVI, DADA2, NOMID and CANDLE, negatively impact blood vessels, resulting in severe tissue damage as well as fatal outcomes for those affected. These diseases are caused by genetic mutations and underlying mechanisms have not been well characterized. We investigate these diseases through clinical evaluation, genetic testing, high content screening and single cell sequencing as well as patient derived cells to investigate the disease mechanisms with in vitro and in vivo murine models. In particular, SAVI (STING-associated vasculopathy with onset in infancy) patients present with early-onset systemic inflammation, cutaneous vasculopathy, and often life-threatening complications of pulmonary fibrosis, the mechanism of which was largely unknown. Endothelial-to-mesenchymal transition (EndMT) is a major mechanism contributing to multiple organ fibrosis. During the reporting period, we identified that spontaneously developed SAVI iEC (induced pluripotent stem cell-derived endothelial cells) EndMT contributes to perivascular fibrosis in SAVI patient lung sections. We further determined that the STING/TBK1/STAT3/Slug pathway is the main mechanism mediating EndMT and identified potential treatments including STING inhibitor, TBK1 inhibitor and JAK/STAT inhibitor using a SAVI-iEC disease model. We also established an iEC/iSMC coculture in-vitro disease model for CANDLE (chronic atypical neutrophilic dermatosis with lipodystrophy and elevated temperatures), which is associated with systemic and pulmonary hypertension. This model will allow us to further explore underlying disease mechanisms and test potential therapeutic tools for CANDLE. Small vessel diseases are conditions with narrowing of small arteries leading to an imbalance of blood supply upon demand, resulting in progressive chronic hypoperfusion with detrimental outcomes. CADASIL is caused by NOTCH3 mutations with predominant clinical features including migraines, strokes, memory loss, and multiple psychiatric symptoms. Similarly, Degos disease is a poorly understood rare genetic disorder leading to small vessel occlusions in multiple organs. Benign Degos is specifically limited to skin lesions, while systemic Degos progresses to other organs (gastrointestinal tract, central nervous system, lungs and heart) and may quickly become fatal. We have developed a robust clinical research program to work on characterizing the etiology/natural history of CADASIL and Degos through comprehensive clinical and molecular evaluations as well as to develop patient-derived disease models to better understand the pathophysiology of these conditions as well as other rare genetic disorders affecting the vascular system. To compare disease progression in patients with rare diseases that have clinical presentation similar to that of more common diseases in the general population, we are incorporating analysis of the population data from the clinical trials and health records in our vascular diseases program. Large population data has recently become available through resources such as the NHLBI BioLINCC, dbGaP or UK Biobank. We are working to develop increased expertise in obtaining and analyzing these large datasets. Over the course of the last year, we also began collaborating with two NIH groups evaluating post-COVID19 populations with a focus on the long-term cardiac and vascular sequelae. When these patients visit the NIH clinical center, we assess cardiac and vascular health using a variety of research techniques including electrocardiograms, echocardiograms, cardiac MRIs, optical imaging studies, NIRS, exercise ABI/Digit BI and eye fluorescein angiography. These tests are performed at various time points post-infection.