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 disease, in which de novo vascular calcifications form in 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 encoder for CD73, an enzyme in the extracellular purine metabolic pathway. Our small non-randomized treatment protocol in seven patients with ACDC with etidronate that was completed in 2021 tested effectiveness of etidronate in attenuating the progression of lower extremity arterial/peri-articular calcification formation and improving vascular blood flow. Results showed that etidronate treatment did not significantly change these parameters over time but may have slowed the rate disease progression. Patients tolerated the medication well with no significant side effects. A manuscript summarizing this data is currently under review. 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 from 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. RNA-Seq analysis identified deficiencies in angiogenesis, extracellular matrix metabolism, and wound healing signaling pathways mediated by dysregulation of HIF1 signaling. Currently, we continue to look for therapeutic targets for AD-HIES patients within signaling pathways affected by deficient HIF1 signaling. Prolidase deficiency (PD) is an autosomal recessive genetic disease caused by mutations in the PEPD gene encoding the enzyme prolidase D, leading to defects in turnover of proline-containing proteins, such as collagen. PD is characterized by chronic severe skin ulcers, recurrent infections, unusual facial features, variable intellectual disability levels, and enlargement of the liver and spleen. To determine the wound healing phenotype in PD patients, we established PD patient-specific iPSC-derived endothelial cell (EC) lines and primary dermal fibroblasts lines. We plan to use single culture/co-culture systems in combination with in vitro scratch wound healing/angiogenesis assays to determine the role of vascular endothelial cells and dermal fibroblasts in PD wound healing as well as to test different kinds of treatment. Autoinflammatory diseases, such as SAVI, DADA2, NOMID, CANDLE and LYN GOF, 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 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/in vivo murine models. 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 mediates EndMT and identified potential treatments including STING, TBK1 and JAK/STAT inhibitors 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. Neutrophilic inflammation is a hallmark of many monogenic autoinflammatory diseases. The pathomechanisms that regulate extravasation of damaging immune cells into surrounding tissues are poorly understood. In the last year, we identified three unrelated patients with perinatal-onset of neutrophilic cutaneous small vessel vasculitis and systemic inflammation caused by constitutive activation of Lyn kinase. Functional studies revealed increased expression of ICAM-1 on iECs and of 2-integrins on patient neutrophils, implicated in increased neutrophil adhesion and vascular transendothelial migration (TEM). Treatment with TNF inhibition improved systemic inflammation and liver fibrosis resolved on treatment with the Src kinase inhibitor dasatinib. Our findings reveal a critical role for Lyn kinase in modulating inflammatory signals, regulating microvascular permeability and neutrophil recruitment, and in promoting hepatic fibrosis. Small vessel diseases involve narrowing of small arteries, leading to reduced blood supply and chronic hypoperfusion. CADASIL, caused by NOTCH3 genetic variants, primarily features migraines, strokes, memory loss, and psychiatric symptoms. Kohlmeier-Degos (K-D) disease K-D is a rare small vessel vasculopathy of unknown etiology leading to small blood vessel occlusions in multiple organs, including the skin, central nervous system, eye, gastrointestinal tract, lungs, and heart. K-D commonly presents as a benign cutaneous form with lesions that appear as erythematous papules and evolve to form a scar with an atrophic, porcelain-white center surrounded by a telangiectatic border. Progression to systemic K-D, or malignant atrophic papulosis, occurs in 2/3 of the patients, is often debilitating, and can be fatal (60-70% mortality rate within 2-5 years after diagnosis) due to GI perforations, brain infarcts, spinal lesions, cardiac or pulmonary failure, sepsis or cachexia. We have established a clinical research program to explore the causes and progression of CADASIL and K-D through detailed clinical and molecular studies and to develop patient-derived models for better understanding their mechanisms. Our CADASIL natural history studies have helped us identify disease features on MRI, eye exam/imaging and non-invasive vascular imaging tests that have not been previously described that could serve as endpoint measurements for testing of therapeutic agents once molecular targets are identified. For our K-D program, we investigate the mechanism of vasculopathy in K-D patients using iPSC-derived patient-specific in vitro and in vivo disease models that are well-established platforms in our team. We are generating iPSC patient cell lines that will be induced to endothelial cells (iECs) and vascular and smooth muscle cells (iVSMCs) for experiments for in vitro co-culture assays to investigate cellular interactions between different cell types and determine cell type-specific transcriptional profile on a single cell level. To compare disease progression in patients with rare diseases with similar clinical presentation to that of more common diseases, we have incorporated population data analysis from 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, as well as general healthcare electronic medical records. We are working to develop increased expertise in obtaining and analyzing these large datasets. As a result, we have analyzed data from the Atherosclerosis Risk in Communities (ARIC) study and electronic medical records from Leumit Healthcare services (Israel) to identify risk factors for chronic diseases (PMID: 37245480; PMID: 36599719) and are currently working to integrate this analysis of risk factors with insights related to gene-disease interactions from rare disease research. We also have on-going collaborations to evaluate post-COVID19 patients with a focus on long-term cardiac and vascular sequelae. We assess cardiac and vascular health of these patients with a variety of research techniques including electrocardiograms, echocardiograms, cardiac MRIs, optical imaging studies, NIRS, etc.