Impact of heterozygous ALK1 mutations on the transcriptomic response to BMP9 and BMP10 in endothelial cells from hereditary hemorrhagic telangiectasia and pulmonary arterial hypertension donors.

Heterozygous activin receptor-like kinase 1 (ALK1) mutations are associated with two vascular diseases: hereditary hemorrhagic telangiectasia (HHT) and more rarely pulmonary arterial hypertension (PAH). Here, we aimed to understand the impact of ALK1 mutations on BMP9 and BMP10 transcriptomic responses in endothelial cells. Endothelial colony-forming cells (ECFCs) and microvascular endothelial cells (HMVECs) carrying loss of function ALK1 mutations were isolated from newborn HHT and adult PAH donors, respectively. RNA-sequencing was performed on each type of cells compared to controls following an 18 h stimulation with BMP9 or BMP10. In control ECFCs, BMP9 and BMP10 stimulations induced similar transcriptomic responses with around 800 differentially expressed genes (DEGs). ALK1-mutated ECFCs unexpectedly revealed highly similar transcriptomic profiles to controls, both at the baseline and upon stimulation, and normal activation of Smad1/5 that could not be explained by a compensation in cell-surface ALK1 level. Conversely, PAH HMVECs revealed strong transcriptional dysregulations compared to controls with > 1200 DEGs at the baseline. Consequently, because our study involved two variables, ALK1 genotype and BMP stimulation, we performed two-factor differential expression analysis and identified 44 BMP9-dysregulated genes in mutated HMVECs, but none in ECFCs. Yet, the impaired regulation of at least one hit, namely lunatic fringe (LFNG), was validated by RT-qPCR in three different ALK1-mutated endothelial models. In conclusion, ALK1 heterozygosity only modified the BMP9/BMP10 regulation of few genes, including LFNG involved in NOTCH signaling. Future studies will uncover whether dysregulations in such hits are enough to promote HHT/PAH pathogenesis, making them potential therapeutic targets, or if second hits are necessary.

The European Rare Disease Network for HHT Frameworks for management of hereditary haemorrhagic telangiectasia in general and speciality care.

Hereditary haemorrhagic telangiectasia (HHT) is a complex, multisystemic vascular dysplasia affecting approximately 85,000 European Citizens. In 2016, eight founding centres operating within 6 countries, set up a working group dedicated to HHT within what became the European Reference Network on Rare Multisystemic Vascular Diseases. By launch, combined experience exceeded 10,000 HHT patients, and Chairs representing 7 separate specialties provided a median of 24 years' experience in HHT. Integrated were expert patients who focused discussions on the patient experience. Following a 2016-2017 survey to capture priorities, and underpinned by more than 40 monthly meetings, and new data acquisitions, VASCERN HHT generated position statements that distinguish expert HHT care from non-expert HHT practice. Leadership was by specialists in the relevant sub-discipline(s), and 100% consensus was required amongst all clinicians before statements were published or disseminated. One major set of outputs targeted all healthcare professionals and their HHT patients, and include the new Orphanet definition; Do's and Don'ts for common situations; Outcome Measures suitable for all consultations; COVID-19; and anticoagulation. The second output set span aspects of vascular pathophysiology where greater understanding will assist organ-specific specialist clinicians to provide more informed care to HHT patients. These cover cerebral vascular malformations and screening; mucocutaneous telangiectasia and differential diagnosis; anti-angiogenic therapies; circulatory interplays between anaemia and arteriovenous malformations; and microbiological strategies to counteract loss of normal pulmonary capillary function. Overall, the integrated outputs, and documented current practices, provide frameworks for approaches that augment the health and safety of HHT patients in diverse health-care settings.

Safety of direct oral anticoagulants in patients with hereditary hemorrhagic telangiectasia.

BACKGROUND: Hereditary hemorrhagic telangiectasia (HHT) is a rare vascular dysplasia resulting in visceral arteriovenous malformations and smaller mucocutaneous telangiectasia. Most patients experience recurrent nosebleeds and become anemic without iron supplementation. However, thousands may require anticoagulation for conditions such as venous thromboembolism and/or atrial fibrillation. Over decades, tolerance data has been published for almost 200 HHT-affected users of warfarin and heparins, but there are no published data for the newer direct oral anticoagulants (DOACs) in HHT. METHODS: To provide such data, a retrospective audit was conducted across the eight HHT centres of the European Reference Network for Rare Multisystemic Vascular Diseases (VASCERN), in Denmark, France, Germany, Italy, the Netherlands and the UK. RESULTS: Although HHT Centres had not specifically recommended the use of DOACs, 32 treatment episodes had been initiated by other clinicians in 28 patients reviewed at the Centres, at median age 65 years (range 30-84). Indications were for atrial fibrillation (16 treatment episodes) and venous thromboembolism (16 episodes). The 32 treatment episodes used Apixaban (n = 15), Rivaroxaban (n = 14), and Dabigatran (n = 3). HHT nosebleeds increased in severity in 24/32 treatment episodes (75%), leading to treatment discontinuation in 11 (34.4%). Treatment discontinuation was required for 4/15 (26.7%) Apixaban episodes and 7/14 (50%) Rivaroxaban episodes. By a 4 point scale of increasing severity, there was a trend for Rivaroxaban to be associated with a greater bleeding risk both including and excluding patients who had used more than one agent (age-adjusted coefficients 0.61 (95% confidence intervals 0.11, 1.20) and 0.74 (95% confidence intervals 0.12, 1.36) respectively. Associations were maintained after adjustment for gender and treatment indication. Extreme hemorrhagic responses, worse than anything experienced previously, with individual nosebleeds lasting hours requiring hospital admissions, blood transfusions and in all cases treatment discontinuation, occurred in 5/14 (35.7%) Rivaroxaban episodes compared to 3/15 (20%) Apixaban episodes and published rates of ~ 5% for warfarin and heparin. CONCLUSIONS: Currently, conventional heparin and warfarin remain first choice anticoagulants in HHT. If newer anticoagulants are considered, although study numbers are small, at this stage Apixaban appears to be associated with lesser bleeding risk than Rivaroxaban.

Hereditary hemorrhagic telangiectasia: from molecular biology to patient care.

SUMMARY: Hereditary hemorrhagic telangiectasia (HHT) is an autosomal dominant vascular disorder characterized by severe and recurrent nosebleeds, mucocutaneous telangiectases, and, in some cases, life-threatening visceral arteriovenous malformations of various types, including pulmonary, hepatic, cerebral, and spinal. Gastrointestinal telangiectases are frequent and may cause severe bleeding. HHT type 1 results from mutations in ENG on chromosome 9 (coding for endoglin), and HHT type 2 results from mutations in ACVRL1 on chromosome 12 (coding for activin receptor-like kinase 1). Mutations of either of these two genes account for most clinical cases. In addition, mutations in MADH4 (encoding SMAD4), which cause a juvenile polyposis/HHT overlap syndrome, have been described, and recently, an HHT3 locus on chromosome 5 (5q31.3-5q32) has been reported. The mutated genes in HHT encode proteins that modulate transforming growth factor-beta superfamily signaling in vascular endothelial cells. Management of patients has changed considerably in the last 20 years, in terms of both treatment and the prevention of complications. The goal of this review was to describe the underlying molecular and cellular physiopathology, explore clinical and genetic diagnostic strategies for HHT, and present clinical management recommendations in order to treat symptomatic disease and to screen for vascular malformations.

The mutational spectrum of human malignant autosomal recessive osteopetrosis.

Human malignant infantile osteopetrosis (arOP; MIM 259700) is a genetically heterogeneous autosomal recessive disorder of bone metabolism, which, if untreated, has a fatal outcome. Our group, as well as others, have recently identified mutations in the ATP6i (TCIRG1) gene, encoding the a3 subunit of the vacuolar proton pump, which mediates the acidification of the bone/osteoclast interface, are responsible for a subset of this condition. By sequencing the ATP6i gene in arOP patients from 44 unrelated families with a worldwide distribution we have now established that ATP6i mutations are responsible for approximately 50% of patients affected by this disease. The vast majority of these mutations (40 out of 42 alleles, including seven deletions, two insertions, 10 nonsense substitutions and 21 mutations in splice sites) are predicted to cause severe abnormalities in the protein product and are likely to represent null alleles. In addition, we have also analysed nine unrelated arOP patients from Costa Rica, where this disease is apparently much more frequent than elsewhere. All nine Costa Rican patients bore either or both of two missense mutations (G405R and R444L) in amino acid residues which are evolutionarily conserved from yeast to humans. The identification of ATP6i gene mutations in two families allowed us for the first time to perform prenatal diagnosis: both fetuses were predicted not to be affected and two healthy babies were born. This study contributes to the determination of genetic heterogeneity of arOP and allows further delineation of the other genetic defects causing this severe condition.

X-linked anhidrotic ectodermal dysplasia with immunodeficiency is caused by impaired NF-kappaB signaling.

The molecular basis of X-linked recessive anhidrotic ectodermal dysplasia with immunodeficiency (EDA-ID) has remained elusive. Here we report hypomorphic mutations in the gene IKBKG in 12 males with EDA-ID from 8 kindreds, and 2 patients with a related and hitherto unrecognized syndrome of EDA-ID with osteopetrosis and lymphoedema (OL-EDA-ID). Mutations in the coding region of IKBKG are associated with EDA-ID, and stop codon mutations, with OL-EDA-ID. IKBKG encodes NEMO, the regulatory subunit of the IKK (IkappaB kinase) complex, which is essential for NF-kappaB signaling. Germline loss-of-function mutations in IKBKG are lethal in male fetuses. We show that IKBKG mutations causing OL-EDA-ID and EDA-ID impair but do not abolish NF-kappaB signaling. We also show that the ectodysplasin receptor, DL, triggers NF-kappaB through the NEMO protein, indicating that EDA results from impaired NF-kappaB signaling. Finally, we show that abnormal immunity in OL-EDA-ID patients results from impaired cell responses to lipopolysaccharide, interleukin (IL)-1beta, IL-18, TNFalpha and CD154. We thus report for the first time that impaired but not abolished NF-kappaB signaling in humans results in two related syndromes that associate specific developmental and immunological defects.

[High-dose chemotherapy in relapse of medulloblastoma in young children]. / Chimiothérapie intensive des rechutes du médulloblastome du jeune enfant.

Craniospinal irradiation is the gold standard treatment used in non metastatic medulloblastoma as prophylaxis against central nervous system (CNS) metastases. However, given the severe late effects caused by this procedure in children under 3 years of age, most pediatric oncologists are currently treating these patients with conventional chemotherapy in order to postpone or even avoid irradiation. In the French Society of Pediatric Oncology (SFOP) this attitude has been adopted since 1990. Among the patients treated without radiotherapy, 20 relapsed while on conventional chemotherapy and were entered in a study of high-dose chemotherapy (HDC) followed by autologous bone marrow transplantation (ABMT). Their median age at diagnosis was 23 months (range: 5-71 months) and the relapse occurred at a median time of 6.3 months after the initiation of chemotherapy. Complete surgical removal of the local relapse was the first treatment in 4/20 patients who were not evaluable for response. Sixteen of the 20 patients had measurable disease at the primary site (9 patients), or at metastatic sites (3 patients) or both (4 patients). The conditioning regimen consisted of combination busulfan 600 mg/m2 over 4 days and thiotepa 900 mg/m2 over 3 days. After recovery from aplasia, patients with a local relapse received local radiotherapy limited to posterior fossa. Among the 16 patients with measurable disease, 6 complete responses, 6 partial responses, 3 non response, were observed following HDC (response rate 75%). One patient was not evaluable. For the 20 patients, the event free survival (EFS) is 50%. Among the surviving patients, the median follow-up is 39.5 months post BMT (range: 21-92 months). Ten patients who developed a local relapse or local progression are alive with non evidence of disease (NED) without craniospinal irradiation. Among the 7 patients who developed a metastases or progression of metastases, only 1 is alive. Toxicity was high but manageable. One complication-related death occurred 1 month post BMT. With a 75% response rate, this HDC proved to be very efficient in relapsed medulloblastoma. A longer follow-up is necessary to demonstrate whether, after a local relapse, HDC could replace craniospinal irradiation as prophylaxis against CNS metastases.

Will high dose chemotherapy followed by autologous bone marrow transplantation supplant cranio-spinal irradiation in young children treated for medulloblastoma?

UNLABELLED: Cranio-spinal irradiation is the gold standard treatment used in non metastatic medulloblastoma as prophylaxis against central nervous system (CNS) metastases. However, given the severe late effects caused by this procedure in children under 3 years of age, most pediatric oncologists are currently treating these patients with conventional chemotherapy in order to postpone or even avoid irradiation. In the French Society of Pediatric Oncology (SFOP) this attitude has been adopted since 1987. Among the patients treated without radiotherapy, 20 relapsed while on conventional chemotherapy and were entered in a study of high-dose chemotherapy (HDC) followed by ABMT. Their median age at diagnosis was 23 months (R5-71) and the relapse occurred at a median time of 6.3 months after the initiation of chemotherapy. Complete surgical removal of the local relapse was the first treatment in 4/20 patients who were not evaluable for response. Sixteen of the twenty patients had measurable disease at the primary site (9 patients), or at metastatic sites (3 patients) or both (4 patients). The conditioning regimen consisted of combination Busulfan 600 mg/m2 over 4 days and Thiotepa 900 mg/m2 over three days. After recovery from aplasia, patients with a local relapse received local radiotherapy limited to posterior fossa. RESULTS: among the 16 patients with measurable disease, 6 CR, 6 PR, 3 NR, were observed following HDC (response rate 75%). One patient was not evaluable. For the 20 patients, the EFS is 50%. Among the surviving patients, the median follow up is 31 months post BMT (R12-82). Ten patients who developed a local relapse or local progression are alive with NED without craniospinal irradiation. Among the 7 patients who developed metastases or progression of metastases, only one is alive. Toxicity was high but manageable: the median duration of granulocytopenia < 0.5 x 109/l and thrombocytopenia < 50 x 10(9)/l was 13 and 41 days respectively. Bacteremia was documented in 4 cases. Grade > 2 mucositis and diarrhea were observed in 60% of patients. One complication-related death occurred 1 month post BMT. CONCLUSION: With a 75% response rate, this HDC proved to be very efficient in relapsed medulloblastoma. A longer follow up is necessary to demonstrate whether, after a local relapse, HDC could replace craniospinal irradiation as prophylaxis against CNS metastases.