Customized Massive Parallel Sequencing Panel for Diagnosis of Pulmonary Arterial Hypertension

Pulmonary arterial hypertension is a very infrequent disease, with a variable etiology and clinical expressivity, making sometimes the clinical diagnosis a challenge. Current classification based on clinical features does not reflect the underlying molecular profiling of these groups. The advance in massive parallel sequencing in PAH has allowed for the describing of several new causative and susceptibility genes related to PAH, improving overall patient diagnosis. In order to address the molecular diagnosis of patients with PAH we designed, validated, and routinely applied a custom panel including 21 genes. Three hundred patients from the National Spanish PAH Registry (REHAP) were included in the analysis. A custom script was developed to annotate and filter the variants. Variant classification was performed according to the ACMG guidelines. Pathogenic and likely pathogenic variants have been found in 15% of the patients with 12% of variants of unknown significance (VUS). We have found variants in patients with connective tissue disease (CTD) and congenital heart disease (CHD). In addition, in a small proportion of patients (1.75%), we observed a possible digenic mode of inheritance. These results stand out the importance of the genetic testing of patients with associated forms of PAH (i.e., CHD and CTD) additionally to the classical IPAH and HPAH forms. Molecular confirmation of the clinical presumptive diagnosis is required in cases with a high clinical overlapping to carry out proper management and follow up of the individuals with the disease.

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Expanding the Evidence of a Semi-Dominant Inheritance in GDF2 Associated with Pulmonary Arterial Hypertension

Cells ◽

10.3390/cells10113178 ◽

2021 ◽

Vol 10 (11) ◽

pp. 3178

Author(s):

Natalia Gallego ◽

Alejandro Cruz-Utrilla ◽

Inmaculada Guillén ◽

Amparo Moya Bonora ◽

Nuria Ochoa ◽

...

Keyword(s):

Pulmonary Arterial Hypertension ◽

Arterial Hypertension ◽

Genetic Diagnosis ◽

Missense Variant ◽

Compound Heterozygous ◽

Incomplete Penetrance ◽

Presumptive Diagnosis ◽

Pathogenic Variants ◽

Pulmonary Arterial ◽

Next Generation Sequencing Ngs

Pulmonary arterial hypertension (PAH) sometimes co-exists with hereditary hemorrhagic telangiectasia (HHT). Despite being clinically diagnosable according to Curaçao criteria, HHT can be difficult to diagnose due to its clinically heterogenicity and highly overlapping with PAH. Genetic analysis of the associated genes ACVRL1, ENG, SMAD4 and GDF2 can help to confirm or discard the presumptive diagnosis. As part of the clinical routine and to establish a genetic diagnosis, we have analyzed a cohort of patients with PAH and overlapping HHT features through a customized Next Generation Sequencing (NGS) panel of 21 genes, designed and validated in-house. We detected a homozygous missense variant in GDF2 in a pediatric patient diagnosed with PAH associated with HHT and a missense variant along with a heterozygous deletion in another idiopathic PAH patient (compound heterozygous inheritance). In order to establish variant segregation, we analyzed all available family members. In both cases, parents were carriers for the variants, but neither was affected. Our results expand the clinical spectrum and the inheritance pattern associated with GDF2 pathogenic variants suggesting incomplete penetrance and/or variability of expressivity with a semi-dominant pattern of inheritance.

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Novel Genetic and Molecular Pathways in Pulmonary Arterial Hypertension Associated with Connective Tissue Disease

Cells ◽

10.3390/cells10061488 ◽

2021 ◽

Vol 10 (6) ◽

pp. 1488

Author(s):

Ignacio Hernandez-Gonzalez ◽

Jair Tenorio-Castano ◽

Nuria Ochoa-Parra ◽

Natalia Gallego ◽

Carmen Pérez-Olivares ◽

...

Keyword(s):

Pulmonary Arterial Hypertension ◽

Connective Tissue ◽

Arterial Hypertension ◽

Connective Tissue Disease ◽

Pathogenic Variant ◽

Pulmonary Vascular Disease ◽

Variants Of Unknown Significance ◽

Pathogenic Variants ◽

Unknown Significance ◽

Pulmonary Arterial

Pulmonary Arterial Hypertension (PAH) is a severe complication of Connective Tissue Disease (CTD), with remarkable morbidity and mortality. However, the molecular and genetic basis of CTD-PAH remains incompletely understood. This study aimed to screen for genetic defects in a cohort of patients with CTD-PAH, using a PAH-specific panel of 35 genes. During recruitment, 79 patients were studied, including 59 Systemic Sclerosis patients (SSc) and 69 females. Disease-associated variants were observed in nine patients: 4 pathogenic/likely pathogenic variants in 4 different genes (TBX4, ABCC8, KCNA5 and GDF2/BMP9) and 5 Variants of Unknown Significance (VUS) in 4 genes (ABCC8, NOTCH3, TOPBP1 and CTCFL). One patient with mixed CTD had a frameshift pathogenic variant in TBX4. Two patients with SSc-PAH carried variants in ABCC8. A patient diagnosed with Systemic Lupus Erythematous (SLE) presented a pathogenic nonsense variant in GDF2/BMP9. Another patient with SSc-PAH presented a pathogenic variant in KCNA5. Four patients with SSc-PAH carried a VUS in NOTCH1, CTCFL, CTCFL and TOPBP1, respectively. These findings suggest that genetic factors may contribute to Pulmonary Vascular Disease (PVD) in CTD patients.

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BMPR2 Promoter Variants Effect Gene Expression in Pulmonary Arterial Hypertension Patients

Genes ◽

10.3390/genes11101168 ◽

2020 ◽

Vol 11 (10) ◽

pp. 1168

Author(s):

Jie Song ◽

Katrin Hinderhofer ◽

Lilian T. Kaufmann ◽

Nicola Benjamin ◽

Christine Fischer ◽

...

Keyword(s):

Gene Expression ◽

Pulmonary Arterial Hypertension ◽

Arterial Hypertension ◽

Transcriptional Activity ◽

Disease Manifestation ◽

Pathogenic Variants ◽

Protein Receptor ◽

Transcriptional Effect ◽

Pulmonary Arterial ◽

Human Pulmonary Artery

Pathogenic variants have been identified in 85% of heritable pulmonary arterial hypertension (PAH) patients. These variants were mainly located in the bone morphogenetic protein receptor 2 (BMPR2) gene. However, the penetrance of BMPR2 variants was reduced leading to a disease manifestation in only 30% of carriers. In these PAH patients, further modifiers such as additional pathogenic BMPR2 promoter variants could contribute to disease manifestation. Therefore, the aim of this study was to identify BMPR2 promoter variants in PAH patients and to analyze their transcriptional effect on gene expression and disease manifestation. BMPR2 promoter variants were identified in PAH patients and cloned into plasmids. These were transfected into human pulmonary artery smooth muscle cells to determine their respective transcriptional activity. Nine different BMPR2 promoter variants were identified in seven PAH families and three idiopathic PAH patients. Seven of the variants (c.-575A>T, c.-586dupT, c.-910C>T, c.-930_-928dupGGC, c.-933_-928dupGGCGGC, c.-930_-928delGGC and c.-1141C>T) led to a significantly decreased transcriptional activity. This study identified novel BMPR2 promoter variants which may affect BMPR2 gene expression in PAH patients. They could contribute to disease manifestations at least in some families. Further studies are needed to investigate the frequency of BMPR2 promoter variants and their impact on penetrance and disease manifestation.

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Whole Exome Sequence Analysis Provides Novel Insights into the Genetic Framework of Childhood-Onset Pulmonary Arterial Hypertension

Genes ◽

10.3390/genes11111328 ◽

2020 ◽

Vol 11 (11) ◽

pp. 1328

Author(s):

Simone M. Gelinas ◽

Clare E. Benson ◽

Mohammed A. Khan ◽

Rolf M. F. Berger ◽

Richard C. Trembath ◽

...

Keyword(s):

Pulmonary Arterial Hypertension ◽

Arterial Hypertension ◽

Molecular Diagnosis ◽

Right Heart Failure ◽

Childhood Onset ◽

Risk Alleles ◽

Pathogenic Variants ◽

Whole Exome ◽

Paediatric Cohort ◽

Pulmonary Arterial

Pulmonary arterial hypertension (PAH) describes a rare, progressive vascular disease caused by the obstruction of pulmonary arterioles, typically resulting in right heart failure. Whilst PAH most often manifests in adulthood, paediatric disease is considered to be a distinct entity with increased morbidity and often an unexplained resistance to current therapies. Recent genetic studies have substantially increased our understanding of PAH pathogenesis, providing opportunities for molecular diagnosis and presymptomatic genetic testing in families. However, the genetic architecture of childhood-onset PAH remains relatively poorly characterised. We sought to investigate a previously unsolved paediatric cohort (n = 18) using whole exome sequencing to improve the molecular diagnosis of childhood-onset PAH. Through a targeted investigation of 26 candidate genes, we applied a rigorous variant filtering methodology to enrich for rare, likely pathogenic variants. This analysis led to the detection of novel PAH risk alleles in five genes, including the first identification of a heterozygous ATP13A3 mutation in childhood-onset disease. In addition, we provide the first independent validation of BMP10 and PDGFD as genetic risk factors for PAH. These data provide a molecular diagnosis in 28% of paediatric cases, reflecting the increased genetic burden in childhood-onset disease and highlighting the importance of next-generation sequencing approaches to diagnostic surveillance.

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