SummaryHereditary haemorrhagic telangiectasia (HHT), or Rendu-Osler-Weber syndrome, is an autosomal dominant vascular disease. The clinical manifestations are epistaxis, mucocutaneous and gastrointestinal telangiectases, and arteriovenous malformations. There are two predominant types of HHT caused by mutations in Endoglin (ENG) and activin receptor-like kinase 1 (ALK1) (ACVRL1) genes, HHT1 and HHT2, respectively. No cure for HHT has been found and there is a current need to find new effective drug treatments for the disease. Some patients show severe epistaxis which interferes with their quality of life. We report preliminary results obtained with Raloxifene to treat epistaxis in postmenopausal HHT women diagnosed with osteoporosis. We tried to unravel the molecular mechanisms involved in the therapeutic effects of raloxifene. ENG and ACVRL1 genes code for proteins involved in the transforming growth factor β pathway and it is widely accepted that haploinsufficiency is the ori- Correspondence to:Luisa María Botella Centro de Investigaciones Biológicas CIB, CSIC Ramiro de Maeztu, 9 28040 Madrid, Spain E-mail: cibluisa@cib.csic.es gin for the pathogenicity of HHT. Therefore, identification of drugs able to increase the expression of those genes is essential to propose new therapies for HHT. In vitro results show that raloxifene increases the protein and mRNA expression of ENG and ALK1 in cultured endothelial cells. Raloxifene also stimulates the promoter activity of these genes, suggesting a transcriptional regulation of ENG and ALK1. Furthermore, Raloxifene improved endothelial cell functions like tubulogenesis and migration in agreement with the reported functional roles of Endoglin and ALK1. Our pilot study provides a further hint that oral administration of raloxifene may be beneficial for epistaxis treatment in HHT menopausal women. The molecular mechanisms of raloxifene involve counteracting the haploinsufficiency of ENG and ALK1.

BackgroundVon Hippel-Lindau (VHL) disease is a rare oncological disease with an incidence of 1:36,000, and is characterized by the growth of different types of tumors: hemangioblastomas in the central nervous system (CNS) and retina, renal carcinoma, pheochromocytomas, pancreatic serous cystadenoma, and endolymphatic sac tumors. These tumors do not express VHL protein (pVHL). pVHL ubiquitinates hypoxia inducible factor (HIF) protein for degradation by the proteasome; in the absence of VHL, HIF translocates to the nucleus to activate the expression of its target genes. Targeting VHL-derived tumors with drugs that have reduced side effects is urgent to avoid repeat CNS surgeries. Recent reports have shown that propranolol, a β-blocker used for the treatment of hypertension and other cardiac and neurological diseases, is the best option for infantile hemangioma (IH). Propranolol could be an efficient treatment to control hemangioblastoma growth in VHL disease because of its antiangiogenic effects demonstrated in IH and the hypothetical impact on HIF levels.MethodsHeLa 9X (HRE) hypoxia responsive element cell line and primary hemangioblastoma-derived cells were subjected to propranolol treatment and cell viability and apoptosis were evaluated. HIF1-α and Hif-2α expression after propranolol treatment was analyzed by western blotting. Quantitative PCR was performed to study the mRNA expression of HIF target genes. Vascular endothelial growth factor (VEGF) was measured in culture supernatants by immunoassay.ResultsPropranolol downregulated HIF-dependent transcription in HeLa 9XHRE cells. Under hypoxic conditions, propranolol decreased the expression of HIF target genes in hemangioblastoma cells, which stopped proliferating and died following long-term treatment. These results suggests that propranolol treatment promoted reduced HIF protein expression and corresponding downregulation of HIF target genes, and inhibited cell proliferation in parallel with induction of cell death by apoptosis.ConclusionsOur results suggest that propranolol could reduce the growth of HIF-dependent tumors and may thus be a promising treatment to delay surgery in VHL patients.

SummaryThe β-blocker propranolol, originally designed for cardiological indications (angina, cardiac arrhythmias and high blood pressure), is nowadays, considered the most efficient drug for the treatment in infantile haemangiomas (IH), a vascular tumour that affects 5–10% of all infants. However, its potential therapeutic benefits in other vascular anomalies remain to be explored. In the present work we have assessed the impact of propranolol in endothelial cell cultures to test if this drug could be used in the vascular disease hereditary haemorrhagic telangiectasia (HHT). This rare disease is the result of abnormal angiogenesis with epistaxis, mucocutaneous and gastrointestinal telangiectases, as well as arteriovenous malformations in several organs, as clinical manifestations. Mutations in Endoglin (ENG) and ACVLR1 (ALK1) genes, lead to HHT1 and HHT2, respectively. Endoglin and ALK1 are involved in the TGF-β1 signalling pathway and play a critical role for the proper development of the blood vessels. As HHT is due to a deregulation of key angiogenic factors, inhibitors of angiogenesis have been used to normalise the nasal vasculature eliminating epistaxis derived from telangiectases. Thus, the antiangiogenic properties of propranolol were tested in endothelial cells. The drug was able to decrease cellular migration and tube formation, concomitantly with reduced RNA and protein levels of ENG and ALK1. Moreover, the drug showed apoptotic effects which could explain cell death in IH. Interestingly, propranolol showed some profibrinolytic activity, decreasing PAI-1 levels. These results suggest that local administration of propranolol in the nose mucosa to control epistaxis might be a potential therapeutic approach in HHT.

BackgroundVon Hippel-Lindau (VHL) disease is a rare oncological disease with an incidence of 1:36,000, and is characterized by the growth of different types of tumours. Haemangioblastomas in the central nervous system (CNS) and retina, renal carcinoma and pheochromocytomas are the most common tumours. The absence of treatment for VHL leads to the need of repeated surgeries as the only option for these patients. Targeting VHL-derived tumours with drugs with reduced side effects is urgent to avoid repeated CNS surgeries. Recent reports have demonstrated that propranolol, a β-blocker used for the treatment of hypertension and other cardiac and neurological diseases, is the best option for infantile hemangioma (IH). Propranolol could be an efficient treatment to control haemangioblastoma growth in VHL disease given its antiangiogenic effects that were recently demonstrated by us. The main objective of the present study was the assessment of the efficacy and safety of propranolol on retinal haemangioblastoma in von Hippel-Lindau disease (VHL).Methods7 VHL patients, from different regions of Spain, affected from juxtapapillary or peripheral haemangioblastomas were administered 120 mg propranolol daily. Patients were evaluated every 3 months for 12 months, at Virgen de la Salud Hospital (Toledo). The patients had juxtapapillary or peripheral haemangioblastomas but had refused standard treatments.ResultsPropranolol was initiated with a progressive increase up to a final dose of 120 mg daily. All tumours remained stable, and no new tumours appeared. The reabsorption of retinal exudation was noted in the two patients having exudates. No adverse effects were recorded. VEGF and miRNA 210 levels were monitored in the plasma of patients as possible biomarkers of VHL. These levels decreased in all cases from the first month of treatment.ConclusionsAlthough more studies are necessary, the results of this work suggest that propranolol is a drug to be considered in the treatment of VHL patients with retinal haemangioblastomas. VEGF and miRNA 210 could be used as biomarkers of the VHL disease activity.Trial registrationThe study has a clinical trial design and was registered at EU Clinical Trials Register and Spanish Clinical Studies Registry, EudraCT Number: 2014–003671-30. Registered 2 September 2014.

Background: Hereditary Hemorrhagic Telangiectasia (HHT) is an autosomal dominant and agedependent vascular disorder characterised mainly by mutations in the Endoglin (ENG) or activin receptor-like kinase-1 (ALK1, ACVRL1) genes.

Hereditary hemorrhagic telangiectasia (HHT), or Rendu-OslerWeber syndrome, is an autosomal-dominant vascular disease. The clinical manifestations are epistaxis, mucocutaneous and gastrointestinal telangiectases, and arteriovenous malformations in internal organs. Patients show severe epistaxis, and/or gastrointestinal bleeding, both of which notably interfere with their quality of life. There are two predominant types of HHT caused by mutations in endoglin (ENG) and ACVRL1/activin receptor-like kinase 1 (ALK1) genes, named HHT1 and HHT2, respectively. ENG and ALK1 code for proteins involved in the transforming growth factor (TGF)-␤1 signaling pathway, and it is widely accepted that HHT pathogenicity results from haploinsufficiency. No cure for HHT has been found, so identification of drugs able to increase the expression of these genes is essential when proposing new therapies. We report the efficacy of tacrolimus (FK506) in increasing ENG and ALK1 expression. The rationale comes from a case report of a patient with HHT who received a liver transplantation after hepatic failure due to a liver arteriovenous malformation. The liver was transplanted, and the immunosuppressor FK506 was used to prevent the rejection. After the first month of FK506 treatment, the internal and external telangiectases, epistaxes, and anemia disappeared. Here, we find that the immunosuppressor FK506 increases the protein and mRNA expression of ENG and ALK1 in cultured endothelial cells and enhances the TGF-␤1/ALK1 signaling pathway and endothelial cell functions like tubulogenesis and migration. These results suggest that the mechanism of action of FK506 involves a partial correction of endoglin and ALK1 haploinsufficiency and may therefore be an interesting drug for use in patients with HHT who undergo transplantation.

Hereditary Haemorrhagic Telangiectasia (HHT) or Rendu-Osler-Weber syndrome, is an autosomal dominant rare disease characterized by localized angiodysplasia. This is manifested as epistaxis, mucocutaneous and gastrointestinal telangiectases, and arteriovenous malformations in the pulmonary, cerebral or hepatic circulation. The prevalence is between 1 in 5,000 to 8,000, although it is higher in some regions. The most frequent clinical manifestation of HHT is epistaxis, normally from light to moderate from the 4(th) decade of life. However, many patients show severe epistaxis which may interfere with their quality of life. The epistaxis is due to telangiectasia on the nasal mucosa. These are focally dilated postcapilar venules, which in advanced phases show many layers of smooth muscle cells without elastic fibers, and very frequently directly connect with dilated arterioles. As a consequence of these vascular alterations, telangiectases are very sensitive to slight trauma and even to the friction with the air when breathing, which gives rise to nose bleeds. Unfortunately, there is no optimal pharmacological treatment for the epistaxis in HHT. The use of antifibrinolytic agents for the treatment of HHT has been studied recently by our group as an effective relief for nasal and gastric haemorrhages. This work represents a systematic review and the beginning of a systematic laboratory work we are now conducting in our lab to screen for "orphan drugs" as therapeutic agents in HHT. In this context, the use of hormones, immunosuppresants and anti-angiogenic agents are under preclinical study in our laboratory.

The diagnosis of hereditary hemorrhagic telangiectasia (HHT) is based on the Curaçao criteria: epistaxis, telangiectases, arteriovenous malformations in internal organs, and family history. Genetically speaking, more than 90% of HHT patients show mutations in ENG or ACVRL1/ALK1 genes, both belonging to the TGF-β/BMP9 signaling pathway. Despite clear knowledge of the symptoms and genes of the disease, we still lack a definite cure for HHT, having just palliative measures and pharmacological trials. Among the former, two strategies are: intervention at “ground zero” to minimize by iron and blood transfusions in order to counteract anemia. Among the later, along the last 15 years, three different strategies have been tested: (1) To favor coagulation with antifibrinolytic agents (tranexamic acid); (2) to increase transcription of ENG and ALK1 with specific estrogen-receptor modulators (bazedoxifene or raloxifene), antioxidants (N-acetylcysteine, resveratrol), or immunosuppressants (tacrolimus); and (3) to impair the abnormal angiogenic process with antibodies (bevacizumab) or blocking drugs like etamsylate, and propranolol. This manuscript reviews the main strategies and sums up the clinical trials developed with drugs alleviating HHT.