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BackgroundInflammation and dysregulated immunity are important in the development of pulmonary arterial hypertension (PAH). Compelling preclinical data supports the therapeutic blockade of interleukin-6 (IL-6) signalling.MethodsWe conducted a phase 2 open-label study of intravenous tocilizumab (8 mg·kg−1) over 6 months in patients with group 1 PAH. Co-primary end-points were safety, defined by incidence and severity of adverse events, and change in pulmonary vascular resistance. Separately, a mendelian randomisation study was undertaken on 11 744 individuals with European ancestry including 2085 patients with idiopathic/heritable disease for the IL-6 receptor (IL6R) variant (rs7529229), known to associate with circulating IL-6R levels.ResultsWe recruited 29 patients (male/female 10/19; mean±sd age 54.9±11.4 years). Of these, 19 had heritable/idiopathic PAH and 10 had connective tissue disease-associated PAH. Six were withdrawn prior to drug administration; 23 patients received at least one dose of tocilizumab. Tocilizumab was discontinued in four patients owing to serious adverse events. There were no deaths. Despite evidence of target engagement in plasma IL-6 and C-reactive protein levels, both intention-to-treat and modified intention-to-treat analyses demonstrated no change in pulmonary vascular resistance. Inflammatory markers did not predict treatment response. Mendelian randomisation did not support an effect of the lead IL6R variant on risk of PAH (OR 0.99, p=0.88).ConclusionAdverse events were consistent with the known safety profile of tocilizumab. Tocilizumab did not show any consistent treatment effect.

Background Diagnostic ionizing radiation is a risk factor for breast cancer (BC). BC risk increases with increased dose to the chest and decreases with increased age at exposure, with possible effect modification related to familial or genetic predisposition. While chest X-rays increase the BC risk of BRCA1/2 mutation carriers compared to non-carriers, little is known for women with a hereditary predisposition to BC but who tested negative for a BRCA1 or BRCA2 (BRCA1/2) mutation. Methods We evaluated the effect of chest X-rays from diagnostic medical procedures in a dataset composed of 1552 BC cases identified through French family cancer clinics and 1363 unrelated controls. Participants reported their history of X-ray exposures in a detailed questionnaire and were tested for 113 DNA repair genes. Logistic regression and multinomial logistic regression models were used to assess the association with BC. Results Chest X-ray exposure doubled BC risk. A 3% increased BC risk per additional exposure was observed. Being 20 years old or younger at first exposure or being exposed before first full-term pregnancy did not seem to modify this risk. Birth after 1960 or carrying a rare likely deleterious coding variant in a DNA repair gene other than BRCA1/2 modified the effect of chest X-ray exposure. Conclusion Ever/never chest X-ray exposure increases BC risk 2-fold regardless of age at first exposure and, by up to 5-fold when carrying 3 or more rare variants in a DNA repair gene. Further studies are needed to evaluate other DNA repair genes or variants to identify those which could modify radiation sensitivity. Identification of subpopulations that are more or less susceptible to ionizing radiation is important and potentially clinically relevant.

Background Heritable pulmonary arterial hypertension (PAH) is most commonly due to heterozygous mutations of the BMPR2 gene. Based on expert consensus, guidelines recommend annual screening echocardiography in asymptomatic BMPR2 mutation carriers. The main objectives of this study were to evaluate the characteristics of asymptomatic BMPR2 mutation carriers, assess their risk of occurrence of PAH and detect PAH at an early stage in this high-risk population. Methods Asymptomatic BMPR2 mutation carriers underwent screening at baseline and annually for a minimum of 2 years (DELPHI-2 study; ClinicalTrials.gov: NCT01600898). Annual screening included clinical assessment, ECG, pulmonary function tests, 6-min walk distance, cardiopulmonary exercise testing, chest radiography, echocardiography and brain natriuretic peptide (BNP) or N-terminal (NT)-proBNP level. Right heart catheterisation (RHC) was performed based on predefined criteria. An optional RHC at rest and exercise was proposed at baseline. Results 55 subjects (26 males; median age 37 years) were included. At baseline, no PAH was suspected based on echocardiography and NT-proBNP levels. All subjects accepted RHC at inclusion, which identified two mild PAH cases (3.6%) and 12 subjects with exercise pulmonary hypertension (21.8%). At long-term follow-up (118.8 patient-years of follow-up), three additional cases were diagnosed, yielding a PAH incidence of 2.3% per year (0.99% per year in males and 3.5% per year in females). All PAH cases remained at low-risk status on oral therapy at last follow-up. Conclusions Asymptomatic BMPR2 mutation carriers have a significant risk of developing incident PAH. International multicentre studies are needed to confirm that refined multimodal screening programmes with regular follow-up allow early detection of PAH., Asymptomatic BMPR2 mutation carriers have a 2.3% per year risk of developing PAH. DELPHI-2 provides the platform for future international multicentre studies to refine multimodal screening algorithms in BMPR2 mutation carriers. http://bit.ly/3oi2KJ1

Contains fulltext : 237895.pdf (Publisher’s version ) (Open Access) BACKGROUND: Ovarian cancer risk in BRCA1 and BRCA2 mutation carriers has been shown to decrease with longer duration of oral contraceptive use. Although the effects of using oral contraceptives in the general population are well established (approximately 50% risk reduction in ovarian cancer), the estimated risk reduction in mutation carriers is much less precise because of potential bias and small sample sizes. In addition, only a few studies on oral contraceptive use have examined the associations of duration of use, time since last use, starting age, and calendar year of start with risk of ovarian cancer. OBJECTIVE: This study aimed to investigate in more detail the associations of various characteristics of oral contraceptive use and risk of ovarian cancer, to provide healthcare providers and carriers with better risk estimates. STUDY DESIGN: In this international retrospective study, ovarian cancer risk associations were assessed using oral contraceptives data on 3989 BRCA1 and 2445 BRCA2 mutation carriers. Age-dependent-weighted Cox regression analyses were stratified by study and birth cohort and included breast cancer diagnosis as a covariate. To minimize survival bias, analyses were left truncated at 5 years before baseline questionnaire. Separate analyses were conducted for each aspect of oral contraceptive use and in a multivariate analysis, including all these aspects. In addition, the analysis of duration of oral contraceptive use was stratified by recency of use. RESULTS: Oral contraceptives were less often used by mutation carriers who were diagnosed with ovarian cancer (ever use: 58.6% for BRCA1 and 53.5% BRCA2) than by unaffected carriers (ever use: 88.9% for BRCA1 and 80.7% for BRCA2). The median duration of use was 7 years for both BRCA1 and BRCA2 carriers who developed ovarian cancer and 9 and 8 years for unaffected BRCA1 and BRCA2 carriers with ovarian cancer, respectively. For BRCA1 mutation carriers, univariate analyses have shown that both a longer duration of oral contraceptive use and more recent oral contraceptive use were associated with a reduction in the risk of ovarian cancer. However, in multivariate analyses, including duration of use, age at first use, and time since last use, duration of oral contraceptive use proved to be the prominent protective factor (compared with 10 years [hazard ratio, 0.37; 95% confidence interval, 0.19-0.73]; P(trend)=.008). The inverse association between duration of use and ovarian cancer risk persisted for more than 15 years (duration of ≥10 years; BRCA1 15 years since last use [hazard ratio, 0.56; 95% confidence interval, 0.18-0.59]). Univariate results for BRCA2 mutation carriers were similar but were inconclusive because of limited sample size. CONCLUSION: For BRCA1 mutation carriers, longer duration of oral contraceptive use is associated with a greater reduction in ovarian cancer risk, and the protection is long term.

Background We report two cases of RASA1-related capillary malformation-arteriovenous malformation (CM-AVM1) syndrome mimicking hereditary haemorrhagic telangiectasia (HHT).Methods and results A 28-year-old man, previously embolised for cerebral arteriovenous malformations (AVMs), presented with epistaxis and typical nasal telangiectasias of HHT. CT scan revealed a large portocaval shunt. The second patient was a 9-year-old girl presenting with cyanosis and several mucocutaneous telangiectasias, similar to those observed in typical cases of HHT. CT scan revealed a huge and complex pulmonary AVM of the right lower lobe and a hepatic AVM within the left lobe. HHT diagnosis was considered possible according to the Curaçao criteria for the two patients, with at least two criteria for each. Genetic tests did not find any mutation in the three classic genes (Endoglin, Activin receptor-like kinase 1 or Mothers against decapentaplegic homolog 4), but identified in both cases an RASA1 mutation, known to cause CM-AVM1 syndrome.Conclusions Pulmonary AVM and portocaval shunt, usually encountered in HHT, have not yet been described in the CM-AVM1 syndrome. RASA1 screening may be considered in case of HHT suspicion, particularly when mutations are not found in the usually affected genes.

Introduction: Heritable pulmonary arterial hypertension (PAH) is most commonly due to heterozygous mutations of the BMPR2 gene (encoding Bone Morphogenetic Protein Receptor type II). Despite the lack of clear evidence, asymptomatic BMPR2 mutation carriers are recommended to undergo annual screening echocardiography for early detection of PAH. Objectives: To evaluate the utility of a comprehensive multimodal PAH screening program in asymptomatic adult BMPR2 mutation carriers. Methods: Asymptomatic BMPR2 mutation carriers underwent screening at baseline and annually for a minimum of two years (DELPHI-2 study, NCT01600898). Annual screening included clinical assessment, electrocardiogram, pulmonary function tests, 6-minute walk distance, cardiopulmonary exercise test (CPET), chest x-ray, echocardiography and N-terminal pro-brain natriuretic peptide level. Right heart catheterization (RHC) was performed based on predefined criteria. An optional RHC at rest and exercise was proposed at baseline. Results: Fifty-five subjects (26 males, median age 37 years) were included. At baseline, no PAH was suspected based on echocardiography and NT pro-BNP levels. All subjects accepted RHC at inclusion which identified two mild PAH cases (3.6%) and 12 subjects with exercise pulmonary hypertension of unknown significance (21.8%). At long term follow-up, three additional PAH cases were diagnosed, yielding a PAH incidence of 2.3%/year. All detected cases have remained at low risk status on oral PAH therapy at last follow-up. Conclusion: In adult BMPR2 mutation carriers, a multimodal screening program with regular follow-up allows early detection of PAH.

Pulmonary veno-occlusive disease (PVOD) occurs in humans either as a heritable form (hPVOD) due to biallelic inactivating mutations of EIF2AK4 (encoding GCN2) or as a sporadic form in older age (sPVOD). The chemotherapeutic agent mitomycin C (MMC) is a potent inducer of PVOD in humans and in rats (MMC-PVOD). Here, we compared human hPVOD and sPVOD, and MMC-PVOD pathophysiology at the histological, cellular, and molecular levels to unravel common altered pathomechanisms. MMC exposure in rats was associated primarily with arterial and microvessel remodeling, and secondarily by venous remodeling, when PVOD became symptomatic. In all forms of PVOD tested, there was convergent GCN2-dependent but eIF2α-independent pulmonary protein overexpression of HO-1 (heme oxygenase 1) and CHOP (CCAAT-enhancer-binding protein [C/EBP] homologous protein), two downstream effectors of GCN2 signaling and endoplasmic reticulum stress. In human PVOD samples, CHOP immunohistochemical staining mainly labeled endothelial cells in remodeled veins and arteries. Strong HO-1 staining was observed only within capillary hemangiomatosis foci, where intense microvascular proliferation occurs. HO-1 and CHOP stainings were not observed in control and pulmonary arterial hypertension lung tissues, supporting the specificity for CHOP and HO-1 involvement in PVOD pathobiology. In vivo loss of GCN2 (EIF2AK4 mutations carriers and Eif2ak4-/- rats) or in vitro GCN2 inhibition in cultured pulmonary artery endothelial cells using pharmacological and siRNA approaches demonstrated that GCN2 loss of function negatively regulates BMP (bone morphogenetic protein)-dependent SMAD1/5/9 signaling. Exogenous BMP9 was still able to reverse GCN2 inhibition-induced proliferation of pulmonary artery endothelial cells. In conclusion, we identified CHOP and HO-1 inhibition, and BMP9, as potential therapeutic options for PVOD. ispartof: AMERICAN JOURNAL OF RESPIRATORY CELL AND MOLECULAR BIOLOGY vol:63 issue:1 pages:118-131 ispartof: location:United States status: published

IntroductionTBX4 mutation causes small patella syndrome (SPS) and/or pulmonary arterial hypertension (PAH). The characteristics and outcomes of PAH associated with TBX4 mutations are largely unknown.MethodsWe report the clinical, functional, radiologic, histologic and haemodynamic characteristics and outcomes of heritable PAH patients carrying a TBX4 mutation from the French pulmonary hypertension (PH) network.Results20 patients were identified in 17 families. They were characterised by a median age at diagnosis of 29 years (0–76 years) and a female to male ratio of three. Most of the patients (70%) were in New York Heart Association (NYHA) functional class III or IV with a severe haemodynamic impairment (median pulmonary vascular resistance (PVR) of 13.6 (6.2–41.8) Wood units). Skeletal signs of SPS were present in 80% of cases. Half of the patients had mild restrictive or obstructive limitation and diffusing capacity of the lung for carbon monoxide (DLCO) was decreased in all patients. High-resolution computed tomography (HRCT) showed bronchial abnormalities, peri-bronchial cysts, mosaic distribution and mediastinal lymphadenopathies. PAH therapy was associated with significant clinical improvement. At follow-up (median 76 months), two patients had died and two had undergone lung transplantation. One-year, three-year and five-year event-free survival rates were 100%, 94% and 83%, respectively. Histologic examination of explanted lungs revealed alveolar growth abnormalities, major pulmonary vascular remodelling similar to that observed in idiopathic pulmonary arterial hypertension (IPAH) and accumulation of cholesterol crystals within the lung parenchyma.ConclusionPAH due to TBX4 mutations may occur with or without skeletal abnormalities across a broad age range from birth to late adulthood. PAH is usually severe and associated with bronchial and parenchymal abnormalities.

International audience; Rationale: Recently, rare heterozygous mutations in GDF2 were identified in patients with pulmonary arterial hypertension (PAH). GDF2 encodes the circulating BMP (bone morphogenetic protein) type 9, which is a ligand for the BMP2 receptor.Objectives: Here we determined the functional impact of GDF2 mutations and characterized plasma BMP9 and BMP10 levels in patients with idiopathic PAH.Methods: Missense BMP9 mutant proteins were expressed in vitro and the impact on BMP9 protein processing and secretion, endothelial signaling, and functional activity was assessed. Plasma BMP9 and BMP10 levels and activity were assayed in patients with PAH with GDF2 variants and in control subjects. Levels were also measured in a larger cohort of control subjects (n = 120) and patients with idiopathic PAH (n = 260).Measurements and Main Results: We identified a novel rare variation at the GDF2 and BMP10 loci, including copy number variation. In vitro, BMP9 missense proteins demonstrated impaired cellular processing and secretion. Patients with PAH who carried these mutations exhibited reduced plasma levels of BMP9 and reduced BMP activity. Unexpectedly, plasma BMP10 levels were also markedly reduced in these individuals. Although overall BMP9 and BMP10 levels did not differ between patients with PAH and control subjects, BMP10 levels were lower in PAH females. A subset of patients with PAH had markedly reduced plasma levels of BMP9 and BMP10 in the absence of GDF2 mutations.Conclusions: Our findings demonstrate that GDF2 mutations result in BMP9 loss of function and are likely causal. These mutations lead to reduced circulating levels of both BMP9 and BMP10. These findings support therapeutic strategies to enhance BMP9 or BMP10 signaling in PAH.

Pulmonary arterial hypertension (PAH) is a rare disease that leads to premature death from right heart failure. It is strongly associated with elevated red cell distribution width (RDW), a correlate of several iron status biomarkers. High RDW values can signal early-stage iron deficiency or iron deficiency anaemia. This study investigated whether elevated RDW is causally associated with PAH. A two-sample Mendelian randomisation (MR) approach was applied to investigate whether genetic predisposition to higher levels of RDW increases the odds of developing PAH. Primary and secondary MR analyses were performed using all available genome-wide significant RDW variants (n=179) and five genome-wide significant RDW variants that act via systemic iron status, respectively. We confirmed the observed association between RDW and PAH (OR 1.90, 95% CI 1.80–2.01) in a multicentre case–control study (cases n=642, disease controls n=15 889). The primary MR analysis was adequately powered to detect a causal effect (odds ratio) between 1.25 and 1.52 or greater based on estimates reported in the RDW genome-wide association study or from our own data. There was no evidence for a causal association between RDW and PAH in either the primary (ORcausal 1.07, 95% CI 0.92–1.24) or the secondary (ORcausal 1.09, 95% CI 0.77–1.54) MR analysis. The results suggest that at least some of the observed association of RDW with PAH is secondary to disease progression. Results of iron therapeutic trials in PAH should be interpreted with caution, as any improvements observed may not be mechanistically linked to the development of PAH., Mendelian randomisation using genetic data from the largest-to-date PAH cohort does not support red cell distribution width or iron deficiency as a cause of PAH, which is important when interpreting iron replacement trials in this condition http://bit.ly/2PPaa88

International audience; Background: Splenic artery aneurysm (SAA) is a rare but potentially fatal condition. Rupture results in 25% mortality up to 75% in pregnant women with 95% fetal mortality. Brief reports suggest an increased risk of developing SAA in patients with HHT.Methods: We analyzed enhanced multidetector CT data in 186 HHT patients matched (gender and ± 5 year old) with 186 controls. We screened for SAA and recorded diameter of splenic and hepatic arteries and hepatic, pancreatic and splenic parenchymal involvements. We determined by univariate and multivariate analysis, the relationship with age, sex, genetic status, cardiovascular risk factors (CVRF) and visceral involvement.Results: SAA concerned 24.7% of HHT patients and 5.4% of controls, p

OBJECTIVE To assess the spectrum of genetic anomalies in a cohort of children presenting at least one cerebral or spinal pial arteriovenous fistula (AVF), and to describe their clinical characteristics. METHODS From 1988 to 2016, all consecutive patients with at least one cerebral or spinal pial AVF were screened for genetic disease. All patients aged

Background: Pulmonary arterial hypertension (PAH) is a rare disease with an emerging genetic basis. Heterozygous mutations in the gene encoding the bone morphogenetic protein receptor type 2 ( BMPR2 ) are the commonest genetic cause of PAH, whereas biallelic mutations in the eukaryotic translation initiation factor 2 alpha kinase 4 gene ( EIF2AK4 ) are described in pulmonary veno-occlusive disease/pulmonary capillary hemangiomatosis. Here, we determine the frequency of these mutations and define the genotype-phenotype characteristics in a large cohort of patients diagnosed clinically with PAH. Methods: Whole-genome sequencing was performed on DNA from patients with idiopathic and heritable PAH and with pulmonary veno-occlusive disease/pulmonary capillary hemangiomatosis recruited to the National Institute of Health Research BioResource–Rare Diseases study. Heterozygous variants in BMPR2 and biallelic EIF2AK4 variants with a minor allele frequency of sorting intolerant from tolerant predictions) were identified as potentially causal. Phenotype data from the time of diagnosis were also captured. Results: Eight hundred sixty-four patients with idiopathic or heritable PAH and 16 with pulmonary veno-occlusive disease/pulmonary capillary hemangiomatosis were recruited. Mutations in BMPR2 were identified in 130 patients (14.8%). Biallelic mutations in EIF2AK4 were identified in 5 patients with a clinical diagnosis of pulmonary veno-occlusive disease/pulmonary capillary hemangiomatosis. Furthermore, 9 patients with a clinical diagnosis of PAH carried biallelic EIF2AK4 mutations. These patients had a reduced transfer coefficient for carbon monoxide (K co ; 33% [interquartile range, 30%–35%] predicted) and younger age at diagnosis (29 years; interquartile range, 23–38 years) and more interlobular septal thickening and mediastinal lymphadenopathy on computed tomography of the chest compared with patients with PAH without EIF2AK4 mutations. However, radiological assessment alone could not accurately identify biallelic EIF2AK4 mutation carriers. Patients with PAH with biallelic EIF2AK4 mutations had a shorter survival. Conclusions: Biallelic EIF2AK4 mutations are found in patients classified clinically as having idiopathic and heritable PAH. These patients cannot be identified reliably by computed tomography, but a low K co and a young age at diagnosis suggests the underlying molecular diagnosis. Genetic testing can identify these misclassified patients, allowing appropriate management and early referral for lung transplantation.

SummaryBackground Hereditary hemorrhagic telangiectasia (HHT) is an autosomal dominant genetic disorder that is caused by mutations in mainly two genes, that is ENG, encoding endoglin (HHT1), or ACVRL1, encoding activin receptor-like kinase 1 (ALK-1/HHT2). HHT is characterized by recurrent epistaxis, mucocutaneous telangiectasia, and vascular visceral dysplasia responsible for visceral arteriovenous malformations (AVM). Aim to report the experience of two university hospitals (Trousseau, Paris, and CHIC, Creteil) with screening children for HHT and pulmonary AVM (PAVM) using high resolution computed tomography (HRCT). Methods parents with confirmed HHT were offered to have their children screened for the mutation identified in their family, and informed consent was obtained. Children carrying the same mutation as their parents underwent HRCT of the chest without contrast. Results between 2008 and 2015, 99 children were screened for HHT mutations. Mutations were identified in 59 patients, that is 24 HHT1 and 35 HHT2. Radiologic and clinical screening was possible in 52 patients (21 HHT-1 and 31 HHT-2). Among those, PAVM was identified in 13 patients (25%; n = 8 HHT1; n = 5 HHT2), and four of them required embolization therapy. Conclusion This study highlights the usefulness of genetic screening in children with known HHT family. It also suggests that a non-invasive protocol such as HRTC is an efficient approach to detect non-symptomatic lesions that are present early on in children carrying the ENG (HHT1), but also the ACVRL1 mutations (HHT2). Pediatr Pulmonol. 2017;52:642–649. © 2017 Wiley Periodicals, Inc.

The impact of bone morphogenetic protein receptor 2 (BMPR2) gene mutations on vascular remodelling in pulmonary arterial hypertension (PAH) is unknown. We sought to identify a histological profile of BMPR2 mutation carriers.Clinical data and lung histology from 44 PAH patients were subjected to systematic analysis and morphometry.Bronchial artery hypertrophy/dilatation and bronchial angiogenesis, as well as muscular remodelling of septal veins were significantly increased in PAH lungs carrying BMPR2 mutations. We found that patients displaying increased bronchial artery remodelling and bronchial microvessel density, irrespective of the mutation status, were more likely to suffer from severe haemoptysis. History of substantial haemoptysis (>50 mL) was significantly more frequent in BMPR2 mutation carriers. 43.5% of BMPR2 mutation carriers, as opposed to 9.5% of noncarriers, displayed singular large fibrovascular lesions, which appear to be closely related to the systemic lung vasculature.Our analysis provides evidence for the involvement of the pulmonary systemic circulation in BMPR2 mutation-related PAH. We show that BMPR2 mutation carriers are more prone to haemoptysis and that haemoptysis is closely correlated to bronchial arterial remodelling and angiogenesis; in turn, pronounced changes in the systemic vasculature correlate with increased pulmonary venous remodelling, creating a distinctive profile in PAH patients harbouring a BMPR2 mutation.

Introduction: Pulmonary hypertension associated with neurofibromatosis type 1 (PH-NF1) is a rare but severe complication of NF1. The largest study about PH-NF1 described 8 patients and there are few data about outcomes on PH treatment. Methods: We report clinical, functional, radiologic, and hemodynamic characteristics and outcomes of patients with pre-capillary PH-NF1 identified within the French PH Registry. Computed tomography (CT) at PH diagnosis were analysed by a pneumologist and a radiologist. Results: We identified 39 NF1-PH cases, characterized by a female predominance (77%) and a median age at PH diagnosis of 63 [min-max 35-82] y-o. At diagnosis, 90% were in NYHA functional class III or IV. Median 6-min walk distance was 209 [50-460] m. Pulmonary function tests showed normal or mild abnormalities except low DLCO (29 [20-58] %). PH was severe with a mean pulmonary artery pressure of 45 [29-71] mmHg, a cardiac index of 2.3 [1.2-3.6] L/min/m2 and pulmonary vascular resistance of 9.8 [4.6-20.1] WU. CT showed pulmonary cysts (72%), ground glass opacities (67%) and emphysema (49%). 38 patients received PAH-specific therapy. Transplant-free survival rates at 1, 3 and 5 years were 92%, 64% and 44%, respectively. Four patients underwent lung transplantation. Conclusions: PH-NF1 was characterized by female predominance, advanced age, severe hemodynamic impairment and poor prognosis.

International audience; Background Heritable forms of pulmonary arterial hypertension (PAH) and pulmonary veno-occlusive disease/pulmonary capillary haemangiomatosis (PVOD/PCH) diverge by lung histopathological lesions, clinical and para-clinical presentation, their responsible genes, and mode of transmission. Since the identification of the BMPR2 gene in families affected by PAH, mutations in several other genes have been discovered for both forms. The mutation landscape in these new genes is not yet well known.Methods We set up a next-generation sequencing-based targeted sequencing gene panel allowing known genes for PAH and PVOD/PCH to be analysed simultaneously. Genetic analysis was prospectively performed on 263 PAH and PVOD/PCH patients (adult and paediatric cases).Results Pathogenic mutations were identified in 19.5% of sporadic PAH patients (n=180), 54.5% of familial PAH patients and 13.3% of PVOD/PCH patients. BMPR2 was the most frequently mutated gene, followed by TBX4 in both paediatric and adult PAH. BMP9 mutations were identified in 1.2% of adult PAH cases. EIF2AK4 biallelic mutations were restricted to PVOD/PCH. A truncating mutation and a predicted loss-of-function variant were also identified in BMP10 in two severely affected sporadic PAH female patients.Conclusion Our results confirm that mutations are found in genes beyond BMPR2 in heritable PAH, emphasise the role of TBX4 and BMP9, and designate BMP10 as a new PAH gene.

International audience; In 2017, we implemented CTNNA1 germline analysis in probands suspected of having hereditary diffuse gastric cancer. Here, we report the results from a retrospective series of 41 cases, including the identification of a new family with a CTNNA1 mutation and the first prophylactic total gastrectomy in an asymptomatic carrier after a normal upper endoscopy. Diffuse gastric cancer foci with loss of catenin alpha-1 expression were seen in the resected tissue, suggesting that CTNNA1 and CDH1 germline mutations behave in a similar manner. Life-changing prophylactic total gastrectomy should therefore also be considered in CTNNA1 mutation carriers.

Background: In pulmonary arterial hypertension (PAH), pathological changes in pulmonary arterioles progressively raise pulmonary artery pressure and increase pulmonary vascular resistance, leading to right heart failure and high mortality rates. Recently, the first potassium channelopathy in PAH, because of mutations in KCNK3 , was identified as a genetic cause and pharmacological target. Methods: Exome sequencing was performed to identify novel genes in a cohort of 99 pediatric and 134 adult-onset group I PAH patients. Novel rare variants in the gene identified were independently identified in a cohort of 680 adult-onset patients. Variants were expressed in COS cells and function assessed by patch-clamp and rubidium flux analysis. Results: We identified a de novo novel heterozygous predicted deleterious missense variant c.G2873A (p.R958H) in ABCC8 in a child with idiopathic PAH. We then evaluated all individuals in the original and a second cohort for rare or novel variants in ABCC8 and identified 11 additional heterozygous predicted damaging ABCC8 variants. ABCC8 encodes SUR1 (sulfonylurea receptor 1)—a regulatory subunit of the ATP-sensitive potassium channel. We observed loss of ATP-sensitive potassium channel function for all ABCC8 variants evaluated and pharmacological rescue of all channel currents in vitro by the SUR1 activator, diazoxide. Conclusions: Novel and rare missense variants in ABCC8 are associated with PAH. Identified ABCC8 mutations decreased ATP-sensitive potassium channel function, which was pharmacologically recovered.

Pulmonary veno-occlusive disease (PVOD) is a rare form of pulmonary hypertension (PH) characterised by preferential remodelling of the pulmonary venules. In the current PH classification, PVOD and pulmonary capillary haemangiomatosis (PCH) are considered to be a common entity and represent varied expressions of the same disease. The recent discovery of biallelic mutations in the EIF2AK4 gene as the cause of heritable PVOD/PCH represents a major milestone in our understanding of the molecular pathogenesis of PVOD. Although PVOD and pulmonary arterial hypertension (PAH) share a similar clinical presentation, with features of severe precapillary PH, it is important to differentiate these two conditions as PVOD carries a worse prognosis and life-threatening pulmonary oedema may occur following the initiation of PAH therapy. An accurate diagnosis of PVOD based on noninvasive investigations is possible utilising oxygen parameters, low diffusing capacity for carbon monoxide and characteristic signs on high-resolution computed tomography of the chest. No evidence-based medical therapy exists for PVOD at present and lung transplantation remains the preferred definitive therapy for eligible patients.

Rationale: Pulmonary arterial hypertension is characterized by vascular remodeling and neomuscularization. PW1 + progenitor cells can differentiate into smooth muscle cells (SMCs) in vitro. Objective: To determine the role of pulmonary PW1 + progenitor cells in vascular remodeling characteristic of pulmonary arterial hypertension. Methods and Results: We investigated their contribution during chronic hypoxia–induced vascular remodeling in Pw1 nLacZ+/− mouse expressing β-galactosidase in PW1 + cells and in differentiated cells derived from PW1 + cells. PW1 + progenitor cells are present in the perivascular zone in rodent and human control lungs. Using progenitor markers, 3 distinct myogenic PW1 + cell populations were isolated from the mouse lung of which 2 were significantly increased after 4 days of chronic hypoxia. The number of proliferating pulmonary PW1 + cells and the proportion of β-gal + vascular SMC were increased, indicating a recruitment of PW1 + cells and their differentiation into vascular SMC during early chronic hypoxia–induced neomuscularization. CXCR4 inhibition using AMD3100 prevented PW1 + cells differentiation into SMC but did not inhibit their proliferation. Bone marrow transplantation experiments showed that the newly formed β-gal + SMC were not derived from circulating bone marrow–derived PW1 + progenitor cells, confirming a resident origin of the recruited PW1 + cells. The number of pulmonary PW1 + cells was also increased in rats after monocrotaline injection. In lung from pulmonary arterial hypertension patients, PW1-expressing cells were observed in large numbers in remodeled vascular structures. Conclusions: These results demonstrate the existence of a novel population of resident SMC progenitor cells expressing PW1 and participating in pulmonary hypertension–associated vascular remodeling.

Genetic causes of pulmonary arterial hypertension (PAH) and pulmonary veno-occlusive disease (PVOD) have been identified, leading to a growing need for genetic counselling.Between 2003 and 2014, genetic counselling was offered to 529 PAH and 100 PVOD patients at the French Referral Centre for Pulmonary Hypertension.Mutations in PAH-predisposing genes were identified in 72 patients presenting as sporadic PAH (17% of cases; 62 mutations in BMPR2, nine in ACVRL1 (ALK1) and one in ENG) and in 94 patients with a PAH family history (89% of cases; 89 mutations in BMPR2, three in ACVRL1 (ALK1) and two in KCNK3). Bi-allelic mutations in EIF2AK4 were identified in all patients with a family history of PVOD (n=19) and in seven patients (8.6%) presenting as sporadic PVOD. Pre-symptomatic genetic diagnosis was offered to 272 relatives of heritable PAH patients, identifying mutations in 36.4% of them. A screening programme is now offered to asymptomatic mutation carriers to detect PAH in an early phase and to identify predictors of outcomes in asymptomatic BMPR2 mutation carriers. BMPR2 screening allowed us to offer pre-implantation diagnosis to two couples with a BMPR2 mutation.Genetic counselling can be implemented in pulmonary hypertension centres.

Background: Heritable pulmonary veno-occlusive disease (PVOD) is linked to mutations in the eukaryotic initiation factor 2 alpha kinase 4 (EIF2AK4) gene, leading to a loss of general control nonderepressible 2 (GCN2). The role of GCN2 expression in pulmonary vascular remodeling remains obscure. We sought to identify specific histologic and biologic features in heritable PVOD. Methods: Clinical data and lung histology of 24 PVOD patients (12 EIF2AK4 mutation carriers, 12 non-carriers) were submitted to systematic histologic analysis and semiautomated morphometry. GCN2 expression was quantified by Western blotting in 24 PVOD patients, 44 patients with pulmonary arterial hypertension (PAH; 23 bone morphogenetic protein receptor type II [. BMPR2] mutation carriers, 21 non-carriers), and 3 experimental pulmonary hypertension models. Results: PVOD patients showed a significant decrease of pulmonary arterial patency (p < 0.0001) compared with healthy controls. Histology of EIF2AK4 mutation carriers was distinctive from non-carriers regarding (1) arterial remodeling, with significantly more severe intimal fibrosis (p = 0.001), less severe medial hypertrophy (p = 0.001), and (2) stronger muscular hyperplasia of interlobular septal veins (p = 0.002). GCN2 expression was abolished in heritable PVOD (p < 0.0001), but also importantly decreased in sporadic PVOD (p = 0.03) as well as in heritable (p = 0.002) and idiopathic PAH (p = 0.003); moreover, GCN2 was abolished in 2 experimental pulmonary hypertension models and importantly decreased in 1 model (p < 0.0001 for all models). Conclusions: Pulmonary arterial remodeling in PVOD is present to an important extent. A significant decrease of GCN2 expression is a common denominator of all tested groups of PVOD and PAH, including their respective experimental models. Our results underline specific morphologic and biologic similarities between PAH and PVOD and let us consider both conditions rather in one large spectrum of disease than as two distinct and clear-cut entities.

BACKGROUND Mutations in BMPR2 encoding bone morphogenetic protein receptor type 2 (BMPRII) is the main genetic risk factor for heritable pulmonary arterial hypertension (PAH). The suspected mechanism is considered to be a defect of BMP signaling. The BMPRII receptor exists in a short isoform without a cytoplasmic tail, which has preserved BMP signaling. METHODS This cohort study compared age at PAH diagnosis and severity between patients carrying a BMPR2 mutation affecting the cytoplasmic tail of BMPRII and affected carriers of a mutation upstream of this domain. RESULTS We identified 171 carriers affected with PAH with a mutated BMPR2 . Twenty-three were carriers of a point mutation located on the cytoplasmic tail of BMPRII. This population was characterized by having an older age at diagnosis compared with other BMPR2 mutation carriers (43.2 ± 12.1 years and 35.7 ± 14.6 years, P = .040), a lower pulmonary vascular resistance (13.3 ± 3.5 and 17.4 ± 6.7, P = .023), and a higher proportion of acute vasodilator responders with a long-term response to calcium channel blockers (8.7% and 0%, P = .02). No statistically significant differences were observed in survival. An in vitro assay showed that mutations located in the cytoplasmic tail led to normal activation of the Smad pathway, whereas activation was abolished in the presence of mutations located in the kinase domain. CONCLUSIONS Patients carrying a mutation affecting the cytoplasmic tail of BMPRII were characterized by an older age at diagnosis compared with other BMPR2 mutation carriers, less severe hemodynamic characteristics, and a greater chance of being a long-term responder to calcium channel blockers. Further investigations are needed to better understand the consequences of these BMPR2 mutations in BMPRII signaling pathways and their possible role in pulmonary arterial remodeling.

RASA1-related disease is a rare autosomal dominant disease characterized by capillary malformations, arteriovenous malformations (AVMs), and/or arteriovenous fistulas (AFVs). Penetrance is nearly complete and vascular malformations may cause serious complications such as organ injury due to oxygenation disorder, brain abscess, hemorrhage, and stroke. Early diagnosis is useful in order to discuss optimal management, including AVMs/AVFs embolization or surgical procedures, and try to prevent some of the complications. In this context, molecular testing of RASA1 gene mutation in relatives may help to better manage the family. All arteriovenous malformations are however not accessible to such procedures. In addition, these therapeutic procedures may result in potential side effects and complications. A couple was referred to our genetics unit and asked us for prenatal genetic testing about a RASA1 mutation. Here, we discuss about arguments that led our team to accept prenatal testing. To the best of our knowledge, no molecular prenatal diagnosis was reported until now in RASA1-related diseases. This first report of prenatal diagnosis in RASA1-related diseases may also offer perspectives for a more general discussion in the field of inherited arteriovenous malformations.

International audience; BackgroundBi-allelic mutations of the EIF2AK4 gene cause heritable pulmonary veno-occlusive disease and/or pulmonary capillary haemangiomatosis (PVOD/PCH). We aimed to assess the eff ect of EIF2AK4 mutations on the clinical phenotypes and outcomes of PVOD/PCH.Methods We did a population-based study using clinical, functional, and haemodynamic data from the registry of the French Pulmonary Hypertension Network. We reviewed the clinical data and outcomes from all patients referred to the French Referral Centre (Pulmonary Department, Hospital Kremlin-Bicêtre, University Paris-Sud) with either confi rmed or highly probable PVOD/PCH with DNA available for mutation screening (excluding patients with other risk factors of pulmonary hypertension, such as chronic respiratory diseases). We sequenced the coding sequence and intronic junctions of the EIF2AK4 gene, and compared clinical characteristics and outcomes between EIF2AK4 mutation carriers and non-carriers. Medical therapies approved for pulmonary arterial hypertension (prostacyclin derivatives, endothelin receptor antagonists and phosphodiesterase type-5 inhibitors) were given to patients according to the clinical judgment and discretion of treating physicians. The primary outcome was the event-free survival (death or transplantation). Secondary outcomes included response to therapies for pulmonary arterial hypertension and survival after lung transplantation. A satisfactory clinical response to specifi c therapy for pulmonary arterial hypertension was defi ned by achieving New York Heart Association functional class I or II, a 6-min walk distance of more than 440 m, and a cardiac index greater than 2·5 L/min per m² at the fi rst reassessment after initiation of specifi c therapy for pulmonary arterial hypertension.Findings We obtained data from Jan 1, 2003, to June 1, 2016, and identifi ed 94 patients with sporadic or heritable PVOD/PCH (confi rmed or highly probable). 27 (29%) of these patients had bi-allelic EIF2AK4 mutations. PVOD/ PCH due to EIF2AK4 mutations occurred from birth to age 50 years, and these patients were younger at presentation than non-carriers (median 26·0 years [range 0–50.3] vs 60·0 years [6·7–81·4] years; p

Background Less than 20 % of familial breast cancer patients who undergo genetic testing for BRCA1 and BRCA2 carry a pathogenic mutation in one of these two genes. The GENESIS (GENE SISter) study was designed to identify new breast cancer susceptibility genes in women attending cancer genetics clinics and with no BRCA1/2 mutation. Methods The study involved the French national network of family cancer clinics. It was based on enrichment in genetic factors of the recruited population through case selection relying on familial criteria, but also on the consideration of environmental factors and endophenotypes like mammary density or tumor characteristics to assess potential genetic heterogeneity. One of the initial aims of GENESIS was to recruit affected sibpairs. Siblings were eligible when index cases and at least one affected sister were diagnosed with infiltrating mammary or ductal adenocarcinoma, with no BRCA1/2 mutation. In addition, unrelated controls and unaffected sisters were recruited. The enrolment of patients, their relatives and their controls, the collection of the clinical, epidemiological, familial and biological data were centralized by a coordinating center. Results Inclusion of participants started in February 2007 and ended in December 2013. A total of 1721 index cases, 826 affected sisters, 599 unaffected sisters and 1419 controls were included. 98 % of participants completed the epidemiological questionnaire, 97 % provided a blood sample, and 76 % were able to provide mammograms. Index cases were on average 59 years old at inclusion, were born in 1950, and were 49.7 years of age at breast cancer diagnosis. The mean age at diagnosis of affected sisters was slightly higher (51.4 years). The representativeness of the control group was verified. Conclusions The size of the study, the availability of biological specimens and the clinical data collection together with the detailed and complete epidemiological questionnaire make this a unique national resource for investigation of the missing heritability of breast cancer, by taking into account environmental and life style factors and stratifying data on endophenotypes to decrease genetic heterogeneity.

Background Mutations in activin receptor-like kinase-1 (ACVRL-1) or endoglin (ENG) are mostly identified in patients with hereditary haemorrhagic telangiectasia (HHT) associated with pulmonary hypertension (PH), but have not yet been studied in Chinese patients. Material and methods In this study, we investigated the clinical and molecular genetic features of Chinese patients with HHT-associated PH and analysed genotype/phenotype correlations in 14 probands and their relatives. Mutation analyses in ACVRL-1, bone morphogenetic protein receptor type 2 (BMPR2) and ENG were performed in 14 Chinese Han patients with HHT-associated PH. Results The overall mutation rate was 71·4%, including 8 ACVRL-1 mutations and 2 ENG mutations, 6 of which were novel. Six patients were identified with arteriovenous malformations (AVMs), including four patients with pulmonary AVMs and two patients with liver AVMs. Five of the patients with AVMs were identified with mutations. Most patients received targeted therapy for PH. Conclusions Our findings have revealed the clinical phenotype and molecular genetic features of HHT-associated PH in Chinese Han patients and indicate that mutations of ACVRL-1 and ENG are genetic predisposing factors in Chinese patients. Our data further addressed clinical management and have provided limited experience in treating this group of disorders.

Based on nationwide data from the French national cancer institute (INCa), we analyzed the evolution of cancer genetics consultations and testing over time, and the uptake of targeted tests in relatives of families with BRCA1/2 or MMR genes mutation. Genetic testing and consultations for familial high-risk individuals are exclusively funded and monitored by the INCa in France. All nationwide cancer genetics centers reported annually standardized parameters of activity from 2003 to 2011. The analysis included a total of 240,134 consultations and 134,652 genetic tests enabling to identify 32,494 mutation carriers. Referral for hereditary breast and ovarian cancer (HBOC) or colorectal cancer predisposition syndromes represented 59 % (141,639) and 23.2 % (55,698) consultations, respectively. From 2003 to 2011, we found a dramatic and steady increase of tests performed for BRCA1/2 (from 2,095 to 7,393 tests/year, P < 0.0001) but not for MMR genes (from 1,144 to 1,635/year, P = NS). The overall percentage of deleterious mutations identified in the probands tested was 13.8 and 20.9 % in HBOC and Lynch syndromes, respectively. Pooled analysis for BRCA1/2 and Lynch syndrome tests showed an inverse relationship between the percentage of mutation detected and the absolute number of tests performed over the time (overall Cochran-Armitage test for trend: P < 0.001). In families with BRCA1/2 or MMR identified mutations, there was an average number of 2.94 and 3.28 relatives performing targeted tests, respectively. This nationwide study shows a lack of referral and genetic testing in Lynch as compared to HBOC syndromes. Only a third of relatives of a proband with a predisposing mutation performed a targeted test. Enhanced information about benefit of genetic testing should be given to clinicians and patients for Lynch syndrome and relatives of a proband carrying an identified predisposing mutation.

Eur J Clin Invest 2012 ABSTRACT: Background The various aetiologies and risk factors for pulmonary arterial hypertension (PAH) lead to close phenotypes with small differences. Plasma microparticles have been shown to be increased in vascular pathologies including PAH. The aim of this study was to determine whether the levels of endothelial and platelet-derived microparticles could vary between different forms of PAH: idiopathic PAH (iPAH), heritable PAH associated with BMPR2 (Bone morphogenetic protein receptor, type II) mutation (hPAH) and PAH associated with connective tissue diseases (aPAH). Materials and methods Microparticles were analysed using flow cytometry in plasma from controls and iPAH, hPAH and aPAH patients. Platelet-derived MP (PMP) were defined as CD31(+) /CD41(+) and endothelial-derived MP (EMP) as CD31(+) /CD41(-) . Two populations of PMP were isolated according to their size, defining small PMP (0·3-0·5 μm) and large PMP (0·5-0·9 μm). BMPR2 genotype, clinical and biologic parameters were recorded. Results EMP and small PMP levels in iPAH, hPAH and aPAH were similar and were significantly increased as compared with controls. No differences in large PMP levels were observed. After adjusting for age, sex, proBNP and CRP, EMP and small PMP levels did not correlate with clinical parameters. Conclusions iPAH, hPAH and aPAH were characterized by increased levels of EMP and of small PMP, a new class of PMP which seems to be differentially produced than large PMP.

BACKGROUND: Bi-allelic EIF2AK4 mutations predispose to the development of pulmonary veno-occlusive disease (PVOD). AIMS OF THE STUDY: To describe phenotype and outcomes of PVOD patients according to EIF2AK4 mutations status. RESULTS: 85 PVOD patients (25 mutations carriers and 60 non-carriers) were identified in the French referral centre for pulmonary hypertension. At diagnosis, EIF2AK4 mutations carriers were significantly younger than non-carriers. PVOD in non-carriers was characterized by male predominance and organic solvents exposure. No difference was observed in NYHA functional class, 6-MWD and hemodynamic characteristics except for pulmonary vascular resistances. 77 PVOD patients (21 EIF2AK4 mutation carriers and 56 non-carriers) received specific PAH therapy and 13 patients (16.8%) experienced drug-induced pulmonary edema. In the 53 patients reassessed within 8 months, mild clinical, functional and hemodynamic improvements were observed. During follow-up, 29 patients were transplanted and 43 patients died. Event-free survival (death or transplantation) at 1, 2 and 3 years was 64%, 56% and 34% in EIF2AK4 mutations carriers and 78%, 47% and 36% in non-carriers. CONCLUSION: PVOD patients carrying bi-allelic EIF2AK4 mutations are characterized by a younger age, severe clinical, functional and hemodynamic impairment and poor outcomes.

Some 15–20% of multiple adenomatous polyposis have no genetic explanation and 20–30% of colorectal cancer (CRC) cases are thought to be due to inherited multifactorial causes. Accumulation of deleterious effects of low-frequency dominant and independently acting variants may be a partial explanation for such patients. The aim of this study was to type a selection of rare and low-frequency variants (

Activin A receptor type II-like kinase-1 (ACVRL1, also known as ALK1) mutation is a cause of hereditary hemorrhagic telangiectasia (HHT) and/or heritable pulmonary arterial hypertension (PAH).To describe the characteristics of patients with PAH carrying an ACVRL1 mutation.We reviewed clinical, functional, and hemodynamic characteristics of 32 patients with PAH carrying an ACVRL1 mutation, corresponding to 9 patients from the French PAH Network and 23 from literature analysis. These cases were compared with 370 patients from the French PAH Network (93 with a bone morphogenetic protein receptor type 2 [BMPR2] mutation and 277 considered as idiopathic cases without identified mutation). Distribution of mutations in the ACVRL1 gene in patients with PAH was compared with the HHT Mutation Database.At diagnosis, ACVRL1 mutation carriers were significantly younger (21.8 +/- 16.7 yr) than BMPR2 mutation carriers and noncarriers (35.7 +/- 14.9 and 47.6 +/- 16.3 yr, respectively; P0.0001). In seven of the nine patients from the French PAH Network, PAH diagnosis preceded manifestations of HHT. ACVRL1 mutation carriers had better hemodynamic status at diagnosis, but none responded to acute vasodilator challenge and they had shorter survival when compared with other patients with PAH despite similar use of specific therapies. ACVRL1 mutations in exon 10 were more frequently observed in patients with PAH, as compared with what was observed in the HHT Mutation Database (33.3 vs. 5%; P0.0001).ACVRL1 mutation carriers were characterized by a younger age at PAH diagnosis. Despite less severe initial hemodynamics and similar management, these patients had worse prognosis compared with other patients with PAH, suggesting more rapid disease progression.

Resume Objectif Au cours des fecondations in vitro (FIV) classiques, 5 % des tentatives se soldent par un echec de fecondation imprevisible, du fait d’une absence d’anomalies des parametres spermatiques. Dans 56 % des cas, aucune cause ovocytaire evidente n’est retrouvee. On observe, par contre, un defaut ou une absence de fixation des spermatozoides a la zone pellucide (ZP). Cette etude a pour objectif de preciser les causes moleculaires d’origine spermatique des echecs de FIV, qui sont a ce jour imprevisibles d’apres les donnees de la spermiologie classique. Patientes et methodes Le profil d’expression proteique des spermatozoides de sujets, presentant un echec complet de fecondation et un defaut de fixation des spermatozoides a la ZP, est compare a celui de temoins (hommes a sperme normal et fecondant, participant a une FIV d’indication tubaire). Pour cela, les echantillons de sperme sont analyses par la technologie d’electrophorese 2 Dimensional Electrophoresis-Differential In Gel Electrophoresis (2D-DIGE) avec au prealable une etape de pre-fractionnement des extraits proteiques selon leur point isoelectrique (fraction acide, intermediaire et basique). Resultats Sept proteines differentiellement exprimees chez tous les cas et chez tous les temoins ont ete mises en evidence dans la fraction acide et sept dans la fraction basique. Douze d’entre elles ont ete identifiees par spectrometrie de masse, dont deux (le recepteur a la laminine LR67 et la L-xylulose reductase) ont deja fait la preuve de leur implication dans l’interaction gametique. Discussion et conclusion Ces proteines meritent des investigations complementaires visant a determiner si certains echecs de fecondation en FIV sont causes par des anomalies au niveau de leur expression.

Objectives Sporadic colorectal cancer is influenced by numerous single nucleotide polymorphisms (SNPs), each with minor effects on the cancer risk. This study seeks to determine whether there is any association of the I1307K, E1317Q and D1822V variants within the Adenomatous polyposis coli gene (APC) and risk to develop colorectal cancer in Tunisian population. Methods Direct sequencing was used to investigate three SNPs in the APC in 48 Tunisian sporadic colorectal cancer cases and 63 controls. Results There was no statistically significant association between the I1307K, E1317Q and D1822V variants investigated and colorectal cancer risk. Conclusion The lack of association may show that these variants selected for this study are not involved in the colorectal carcinogenic process. Otherwise, the eventual biological effect is so little to go undetected, unless increasing the sample size.

BACKGROUND: The TP53 pathway, in which TP53 and its negative regulator MDM2 are the central elements, has an important role in carcinogenesis, particularly in BRCA1- and BRCA2-mediated carcinogenesis. A single nucleotide polymorphism (SNP) in the promoter region of MDM2 (309T > G, rs2279744) and a coding SNP of TP53 (Arg72Pro, rs1042522) have been shown to be of functional significance. METHODS: To investigate whether these SNPs modify breast cancer risk for BRCA1 and BRCA2 mutation carriers, we pooled genotype data on the TP53 Arg72Pro SNP in 7011 mutation carriers and on the MDM2 309T > G SNP in 2222 mutation carriers from the Consortium of Investigators of Modifiers of BRCA1/2 (CIMBA). Data were analysed using a Cox proportional hazards model within a retrospective likelihood framework. RESULTS: No association was found between these SNPs and breast cancer risk for BRCA1 (TP53: per-allele hazard ratio (HR) = 1.01, 95% confidence interval (CI): 0.93-1.10, P(trend) = 0.77; MDM2: HR = 0.96, 95% CI: 0.84-1.09, P(trend) = 0.54) or for BRCA2 mutation carriers (TP53: HR = 0.99, 95% CI: 0.87-1.12, P(trend) = 0.83; MDM2: HR = 0.98, 95% CI: 0.80-1.21, P(trend) = 0.88). We also evaluated the potential combined effects of both SNPs on breast cancer risk, however, none of their combined genotypes showed any evidence of association. CONCLUSION: There was no evidence that TP53 Arg72Pro or MDM2 309T > G, either singly or in combination, influence breast cancer risk in BRCA1 or BRCA2 mutation carriers. British Journal of Cancer (2009) 101, 1456-1460. doi: 10.1038/sj.bjc.6605279 www.bjcancer.com Published online 25 August 2009 (C) 2009 Cancer Research UK

Sporadic colorectal tumorigenesis is caused by alterations in the Wnt (APC, CTNNB1) and Ras pathways. Our objective was to analyze the occurrence of these genetic alterations in relation to tumor and patient characteristics. The prevalence of somatic alteration in the hot-spot regions of the APC, BRAF, and CTNNB1 genes was investigated in 48 unselected and unrelated Tunisian patients with sporadic colorectal cancer, and the association between the molecular features at these genes in relation to tumor and patient characteristics (age at diagnosis, sex, tumor localization, stage, and differentiation) was analyzed. Loss of heterozygosity was observed at the APC locus in 52% of the analyzed tumors. 6 novel mutations were detected by polymerase chain reaction sequencing in the mutation cluster region of the APC gene. No mutations were observed in the CTNNB1 gene in any tumor, but 8% of tumors harbored mutation in the BRAF gene. Clinicopathological analyses showed an association between APC point mutations and the earliest occurrence of sporadic colorectal cancer. The findings confirm the heterogeneity of APC gene alteration and also reveal a particular profile of this pathology among Tunisian patients that confirms the epidemiological data for this country.

Apelin has been identified as the endogenous ligand of the human orphan G protein–coupled receptor APJ. This peptide exerts a variety of cardiovascular effects and particularly acts as an activator of angiogenesis. Importantly, hypoxia has been reported to regulate apelin expression but the molecular mechanism underlying hypoxia-induced apelin expression and the relationship with the physiological response of the apelin/APJ system are still not established. Here, we demonstrate that apelin expression is induced by hypoxia in cultured endothelial and vascular smooth muscle cells as well as in lung from mice exposed to acute hypoxia. Transient transfection experiments show that hypoxia-inducible transcriptional activation of apelin requires an intact hypoxia-responsive element (+813/+826) located within the first intron of the human apelin gene. Chromatin immunoprecipitation assay reveals that hypoxia-inducible factor-1α binds to the endogenous hypoxia-responsive element site of the apelin gene. Moreover, overexpression of hypoxia-inducible factor-1α increases the transcriptional activity of a reporter construct containing this hypoxia-responsive element, whereas small interfering RNA–mediated hypoxia-inducible factor-1α knockdown abolishes hypoxia-induced apelin expression. Finally, microinterfering RNA-mediated apelin or APJ receptor knockdown inhibits both hypoxia-induced endothelial cell proliferation in vitro and hypoxia-induced vessel regeneration in the caudal fin regeneration of Fli-1 transgenic zebrafish. The hypoxia-induced apelin expression may, thus, provide a new mechanism involved in adaptive physiological and pathophysiological response of vascular cells to low oxygen level.

Germline mutations in the gene encoding for bone morphogenetic protein receptor 2 (BMPR2) are a cause of pulmonary arterial hypertension (PAH).We conducted a study to determine the influence, if any, of a BMPR2 mutation on clinical outcome.The French Network of Pulmonary Hypertension obtained data for 223 consecutive patients displaying idiopathic or familial PAH in whom point mutation and large size rearrangements of BMPR2 were screened for. Clinical, functional, and hemodynamic characteristics, as well as outcomes, were compared in BMPR2 mutation carriers and noncarriers.Sixty-eight BMPR2 mutation carriers (28 familial and 40 idiopathic PAH) were compared with 155 noncarriers (all displaying idiopathic PAH). As compared with noncarriers, BMPR2 mutation carriers were younger at diagnosis of PAH (36.5 +/- 14.5 vs. 46.0 +/- 16.1 yr, P0.0001), had higher mean pulmonary artery pressure (64 +/- 13 vs. 56 +/- 13 mm Hg, P0.0001), lower cardiac index (2.13 +/- 0.68 vs. 2.50 +/- 0.73 L/min/m(2), P = 0.0005), higher pulmonary vascular resistance (17.4 +/- 6.1 vs. 12.7 +/- 6.6 mm Hg/L/min/m(2), P0.0001), lower mixed venous oxygen saturation (59 +/- 9% vs. 63 +/- 9%, P = 0.02), shorter time to death or lung transplantation (P = 0.044), and younger age at death (P = 0.002), but similar overall survival (P = 0.51).BMPR2 mutation carriers with PAH present approximately 10 years earlier than noncarriers, with a more severe hemodynamic compromise at diagnosis.

Hereditary hemorrhagic telangiectasia (HHT) is an autosomal dominant disease characterized by widespread arteriovenous malformations and caused by mutations in two major genes: ENG and ACVRL1. Two decades ago, a French epidemiological study pointed out that its prevalence was higher than previously thought and that its distribution varied greatly from one area to another, one of the highest concentrations of patients being found in the Haut-Jura mountains. Although germline mutations are usually family specific, some of them have been reported in unrelated patients, especially for ACVRL1. We performed haplotype analysis of 116 French and Italian patients carrying 13 ACVRL1 different mutations. For five of these mutations, we estimated the age of the most recent common ancestors (MRCAs) using the ESTIAGE program. Most mutations were related to both recurrent mutational events and founder effects with age estimates ranging from 100 to 550 years. The c.1112dupG mutation, which is likely to be responsible for the very high concentration of HHT patients found in the former epidemiological study, probably occurred in one inhabitant of the Haut-Jura Mountains more than three centuries ago. The p.Arg374Gln mutation occurred independently in at least two distinct geographical areas, including the area with the second highest prevalence in the epidemiological study and where the MRCA is rather recent (about 100 years ago). Partially shared haplotypes between French and Italian patients were found for three mutations. This suggests a common origin and a possible diffusion of these mutations from Italy to France.

Background: Pulmonary veno-occlusive disease (PVOD) is an uncommon form of pulmonary hypertension (PH), defined by pathologic changes of the pulmonary veins and capillaries. Heritable PVOD is associated with bi-allelic mutations in EIF2AK4 gene. Aims of the Study: To review clinical, functional, hemodynamic characteristics and outcomes of PVOD patients carriers of bi-allelic mutations in EIF2AK4 gene follow-up in the French PAH Network. Results: We identified 24 PVOD patients carriers of EIF2AK4 mutations (age at diagnosis of 26.4±8.7yrs and sex ratio 1.2). Most of the patients were in NYHA FC III (n=17) or IV (n=5) and 6-minute walk distance was 365±168m. Pulmonary function tests were normal, except a low DLCO of 30±7%. Right heart catheterization revealed severe hemodynamic impairment at diagnosis (mPAP 51±12mmHg, cardiac index 2.53±0.89L/min/m2, pulmonary vascular resistance 11.3±6.2WU with pulmonary capillary wedge pressure 8±4mmHg ). Specific PH therapy was iniated in 21 patients. No significant clinical, functional or hemodynamic improvement was observed and 3 patients experienced drug-induced pulmonary edema. During follow-up, 4 patients died and 18 were transplanted. No recurrence of PVOD occured after transplantation. Event-free survival (death or transplantation) at 1, 2 and 3 years were 63%, 52% and 30% respectively. Conclusion: Heritable PVOD due to EIF2AK4 mutations were characterized by a young age at diagnosis and severe clinical, functional and hemodynamic impairment. This condition was associated with an absence of response to specific PH therapy and poor outcomes. Lung transplantation remains the treatment of choice.

Introduction: Recent reports have shown that mutations in the eukaryotic initiation factor 2 alpha kinase 4 (EIF2AK4) gene cause autosomal recessively inheritable pulmonary veno-occlusive disease (PVOD) in familial and a subset of sporadic cases. The pathophysiology evolving from EIF2AK4 mutations remains still to be elucidated. Aim and objective: We sought to differentiate histopathological characteristics between EIF2AK4 mutation-carriers and non-carriers with PVOD. Methods: Histology of lung samples obtained after transplantation of EIF2AK4 mutation-carriers (n=9) and non-carriers (n=9) displaying clinical PVOD were systematically analyzed and scored in a blinded fashion, using quantitative semi-automated morphometry. Results: EIF2AK4 mutation-carriers were significantly distinctive from non-carriers regarding several patterns: a) intimal fibrosis (more severe) and media hypertrophy (less severe) of pulmonary arteries; b) diffuse interstitial fibrosis (less frequent) and patchy hemangiomatosis-like foci (more frequent) within the parenchyma. A trend to heavier remodeling of interlobular septal veins was observed. Conclusion: Histology of bi-allelic EIF2AK4 mutation-carriers can be differentiated from non-carriers due to arterial and interstitial remodeling patterns. Whether the severe arterial remodeling is of primary or secondary nature in the setting of heritable PVOD requires further insight into EIF2AK4 biology.

Pulmonary veno-occlusive disease (PVOD) is a rare form of pulmonary hypertension characterised by predominant remodelling of pulmonary venules. Bi-allelic mutations in the eukaryotic translation initiation factor 2α kinase 4 (EIF2AK4) gene were recently described as the major cause of heritable PVOD, but risk factors associated with PVOD remain poorly understood. Occupational exposures have been proposed as a potential risk factor for PVOD, but epidemiological studies are lacking.A case–control study was conducted in consecutive PVOD (cases, n=33) and pulmonary arterial hypertension patients (controls, n=65). Occupational exposure was evaluated via questionnaire interview with blinded assessments using an expert consensus approach and a job exposure matrix (JEM).Using the expert consensus approach, PVOD was significantly associated with occupational exposure to organic solvents (adjusted OR 12.8, 95% CI 2.7–60.8), with trichloroethylene being the main agent implicated (adjusted OR 8.2, 95% CI 1.4–49.4). JEM analysis independently confirmed the association between PVOD and trichloroethylene exposure. Absence of significant trichloroethylene exposure was associated with a younger age of disease (54.8±21.4 years, p=0.037) and a high prevalence of harbouring bi-allelic EIF2AK4 mutations (41.7% versus 0%, p=0.015).Occupational exposure to organic solvents may represent a novel risk factor for PVOD. Genetic background and environmental exposure appear to influence the phenotypic expression of the disease.

Background. Chronic allograft nephropathy is the main cause of long-term kidney graft loss. The plasminogen activator inhibitor type 1 (PAI-1) is a potential fibrogenic molecule whose secretion is regulated by several metabolic, inflammatory, and genetic factors. We aimed to determine whether PAI-1 secretion in renal transplant patients is correlated with the decline in renal function after transplantation. Methods. Renal transplant patients (145 male/71 female) were included in the study 1-27 years after transplantation (median of follow-up: 7.35 years). At inclusion, routine clinical and biological data were collected, the 4G/5G polymorphism of the recipient PAI-1 gene was determined, and the PAI-1 plasma level was measured. Results. The mean rate of decline in renal function was -4.26±0.30 ml/min/year. By multiple linear regression analysis, the rate of decline in renal function was significantly correlated with proteinuria (P=0.0176), occurrence of late acute rejection episodes (P=0.0001), and PAI-1 plasma level (P=0.0051). In addition, PAI-1 plasma level was also significantly correlated with body mass index (P=0.038), insulin (P

To the Editor: Dyspnoea curtails daily-living activities in patients with pulmonary veno-occlusive disease (PVOD) [1, 2] and pulmonary arterial hypertension (PAH) [3–5]. It is a common clinical observation that PVOD patients may experience greater dyspnoea than PAH patients during daily activities [1, 2]. However, this clinical feature and its putative underlying mechanisms have not yet been explored. Cardiopulmonary exercise testing (CPET) is well suited for understanding mechanisms underlying dyspnoea during exercise both in research and clinical settings [4, 5]. In the present study, we hypothesised that the perceived clinical difference between PVOD and PAH regarding exertional dyspnoea would be reflected by a different physiological response to CPET. Building on recent advances obtained with CPET in PAH patients [4], we set out to evaluate the relationship between exertional dyspnoea and the physiological response to CPET in eight PVOD patients presenting with recessive mutations in EIF2AK4 , compared with 16 idiopathic or heritable PAH patients. We studied eight clinically stable patients referred to the French Reference Center for Pulmonary Hypertension (Le Kremlin-Bicetre, France) for management of PAH [6, 7]. Using whole-exome sequencing, we detected recessive mutations in the major gene linked to PVOD development, EIF2AK4 (also called GCN2 ), that co-segregated with PVOD in eight patients studied, as recently described [8]. 16 clinically stable patients with diagnosed idiopathic or heritable PAH [9] and without other concomitant diseases were also evaluated. Pulmonary function tests, cycle ergometer symptom-limited incremental CPET, and measurements at rest and at peak exercise of arterial oxygen tension ( P aO2) and arterial carbon dioxide tension ( P aCO2), the physiological dead space ( V D)/tidal volume ( V T) ratio, and the alveolar–arterial oxygen tension gradient ( P A–aO2) and …

BackgroundAngiotensin-converting enzyme (ACE) metabolizes many small peptides and plays a key role in blood pressure regulation. Elevated serum ACE is claimed to be associated with an increased risk for cardiovascular disease. Previously, two families with dramatically increased serum ACE were described, but no systematic survey of affected individuals was performed, and the molecular background of this trait is unknown.Methods and ResultsEight families were identified with autosomal dominant inheritance of a dramatic (5-fold) increase of serum ACE activity. Strikingly, no clinical abnormalities were apparent in the affected subjects. Isolated blood cells were used for genetic and biochemical analysis. The level of ACE expression on the blood leukocytes and dendritic cells and total cell-associated ACE of the affected individuals was similar to that in nonaffected relatives; however membrane-bound mutant ACE was much more efficiently clipped from the cell surface compared with its wild-type counterpart. A point mutation causing Pro1199Leu in the stalk region of the ACE molecule cosegregates with the increase in serum ACE (LOD score, 6.63).ConclusionsA point mutation in the stalk region of the ACE protein causes increased shedding, leading to increased serum ACE, whereas cell-bound ACE is unaltered, and affected individuals exhibit no clinical abnormalities. These findings qualify the importance of serum ACE and establish a new determinant of ACE solubilization.

To the Editor: Pulmonary arterial hypertension (PAH) has been the focus of major research in recent years [1]. Involvement of mutations in genes encoding for members of the transforming growth factor-β signalling pathway ( BMPR2 , ACVRL1 , ENG and SMAD8 ) has been demonstrated in the development of heritable PAH, allowing novel experimental and clinical approaches [2–4]. However, ∼30% of familial forms of PAH remain without any identification of genetic mutations. Recently, mutations of the KCNK3 gene (encoding K+ channel subfamily K member 3) have been reported in patients with familial and sporadic PAH [5]. KCNK3 belongs to a family of mammalian K+ channels, and are involved in the regulation of resting membrane potential, pulmonary vascular tone and in vascular remodelling. This result paves the way to the involvement of novel signalling pathways in the development of heritable PAH. Herein, we describe a novel association of PAH and a channelopathy due to mutation in ATP1A2 (encoding the α2-subunit of the Na+/K+-ATPase), a mutation known to cause familial hemiplegic migraine (FHM), a rare autosomal dominant disease [6]. A 24-year-old male was referred with a 1-year history of progressive exertional dyspnoea. Since the age of 8 years, he has reported recurrent episodes of hemiplegic migraine associated with muscle weakness and pain. The proband’s mother (II4) (fig. 1) and two of his brothers (III6 and III7) had recurrent hemiplegic migraine with aura. There was no familial history of PAH. On admission, the patient was in New York Heart Association (NYHA) functional class III. His 6-min …