Roberto Zarrabeitia

Estrogen therapy for hereditary haemorrhagic telangiectasia (HHT): Effects of raloxifene, on Endoglin and ALK1 expression in endothelial cells

Hereditary 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 beta pathway and it is widely accepted that haploinsufficiency is the origin 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.

Therapeutic action of tranexamic acid in hereditary haemorrhagic telangiectasia (HHT): Regulation of ALK-1/endoglin pathway in endothelial cells

Recurrent epistaxis is the most frequent clinical manifestation of hereditary haemorrhagic telangiectasia (HHT). Its treatment is difficult. Our objective was to assess the use of tranexamic acid (TA), an antifibrinolytic drug, for the treatment of epistaxis in HHT patients and to investigate in vitro the effects of TA over endoglin and ALK-1 expression and activity in endothelial cells. A prospective study was carried out on patients with epistaxis treated with oral TA in the HHT Unit of Sierrallana Hospital (Cantabria, Spain). Primary cultures of endothelial cells were treated with TA to measure the levels of endoglin and ALK-1 at the cell surface by flow cytometry. RNA levels were also measured by real-time PCR, and the transcriptional effects of TA on reporters for endoglin, ALK-1 and the endoglin/ALK-1 TGF-beta pathway were assessed. The results showed that the fourteen HHT patients treated orally with TA improved, and the frequency and severity of their epistaxis were decreased. No complications derived from the treatment were observed. Cultured endothelial cells incubated with TA exhibited increased levels of endoglin and ALK-1 at the protein and mRNA levels, enhanced TGF-beta signaling, and improved endothelial cell functions like tubulogenesis and migration. In summary, oral administration of TA proved beneficial for epistaxis treatment in selected patients with HHT. In addition to its already reported antifibrinolytic effects, TA stimulates the expression ofALK-1 and endoglin, as well as the activity of the ALK-1/endoglin pathway.

Propranolol as antiangiogenic candidate for the therapy of hereditary haemorrhagic telangiectasia

The 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.

Mutation study of Spanish patients with Hereditary Hemorrhagic Telangiectasia

BackgroundHereditary Hemorrhagic Telangiectasia (HHT) is an autosomal dominant and age-dependent vascular disorder characterised mainly by mutations in the Endoglin (ENG) or activin receptor-like kinase-1 (ALK1, ACVRL1) genes.MethodsHere, we have identified 22 ALK1 mutations and 15 ENG mutations, many of which had not previously been reported, in independent Spanish families afflicted with HHT.ResultsWe identified mutations in thirty-seven unrelated families. A detailed analysis of clinical symptoms was recorded for each patient analyzed, with a higher significant presence of pulmonary arteriovenous malformations (PAVM) in HHT1 patients over HHT2. Twenty-two mutations in ALK1 and fifteen in ENG genes were identified. Many of them, almost half, represented new mutations in ALK1 and in ENG. Missense mutations in ENG and ALK1 were localized in a tridimensional protein structure model.ConclusionOverall, ALK1 mutations (HHT2) were predominant over ENG mutations (HHT1) in our Spanish population, in agreement with previous data from our country and other Mediterranean countries (France, Italy), but different to Northern Europe or North America. There was a significant increase of PAVM associated with HHT1 over HHT2 in these families.

Local sclerotherapy with polydocanol (Aethoxysklerol®) for the treatment of Epistaxis in Rendu-Osler-Weber or Hereditary Hemorrhagic Telangiectasia (HHT): 15 years of experience.

Hereditary Haemorrhagic Telangiectasia or Rendu-Osler-Weber syndrome is a rare autosomal dominant vascular disease characterized by mucocutaneous and gastrointestinal telangiectases and localized arteriovenous malformations in lung, brain and liver. Epistaxis, due to rupture of telangiectases of the nasal mucosa, is the most frequent clinical manifestation, leading in many cases to severe impairment of the quality of life in the patients. Though several treatments have been used to reduce epistaxis, none have been completely effective, with the exception of polydocanol (Aethoxysklerol®) in submucosal or subpericondrial injections, which was first presented in 2000 with very good results. After fifteen years using polydocanol in submucosal injections on 45 patients and with nearly 300 injections, we have observed that in 95% of all cases, their nose bleeds improved with respect to frequency and quantity without any important side effects. There was just one case of septal perforation, another with increased septal perforation, and one patient who suffered from dizziness and blurred vision for a few minutes. In this paper the results obtained using this technique over a fifteen-year period will be presented and evaluated.

Tratamiento de las epistaxis en la telangiectasia hemorrágica hereditaria (enfermedad de Rendu-Osler-Weber) con ácido tranexámico

Introduccion Las epistaxis son la manifestacion clinica mas frecuente en los enfermos con telangiectasia hemorragica hereditaria (HHT). Su tratamiento es en ocasiones muy complejo, y no hay consenso sobre el protocolo terapeutico que se ha de aplicar. El objetivo de nuestro estudio fue valorar la utilidad del acido tranexamico oral en las epistaxis secundarias a HHT. Pacientes y metodo Se realizo un estudio prospectivo de pacientes con HHT tratados por epistaxis en la unidad de HHT de nuestro hospital mediante acido tranexamico durante 3 anos. Resultados Se trato a 10 pacientes con HHT mediante acido tranexamico oral durante el periodo de estudio. Todos ellos presentaron una disminucion tanto en la intensidad como la frecuencia de las epistaxis. En su mayoria estaban satisfechos con los resultados del tratamiento. Ninguno presento complicaciones relacionadas con el y 2 pacientes siguieron precisando otros tratamientos para control de las epistaxis. Conclusiones El acido tranexamico oral es util en el tratamiento de las epistaxis en algunos pacientes con HHT, pues reduce de forma significativa tanto la intensidad como la frecuencia. Sin embargo, en los que presentan epistaxis severas no impide la necesidad de seguir realizando otros tratamientos agresivos.

Treatment of Epistaxes in Hereditary Haemorrhagic Telangiectasia (Rendu-Osler- Weber Disease) With Tranexamic Acid

OBJECTIVE Recurrent epistaxis is the most frequent clinical manifestation of hereditary haemorrhagic telangiectasia (HHT). Its treatment occasionally presents difficulties as there is no consensus on the appropriate therapeutic protocol. Our objective was to explore the utility of oral tranexamic acid for the treatment of epistaxes in HHT patients. PATIENTS AND METHOD A 3-year prospective study was carried on HHT patients with epistaxis treated with oral tranexamic acid in the HHT unit at our hospital. RESULTS Ten patients with HHT were treated with oral tranexamic acid during the study. Most of them improved both the frequency and severity of their epistaxis and were satisfied with the treatment. No treatment-related complications were recorded. Two patients needed more aggressive treatments to control epistaxis. CONCLUSIONS Oral tranexamic acid is useful for achieving significant reductions in epistaxis frequency and intensity in selected patients with HHT. In those presenting severe epistaxis, however, it may need to be combined with more aggressive therapies.

Telangiectasia hemorràgica hereditaria

La telangiectasia hemorragica hereditaria (HHT), o sindrome de Rendu-Osler-Weber, es una enfermedad genetica con transmision autosomica dominante. Su prevalencia se estima en un paciente por 5-8.000 habitantes. Se han descrito mutaciones en 2 genes (gen de la endoglina y de ALK-1), y pueden distinguirse 2 tipos segun el gen mutado: HHT 1 y HHT 2, respectivamente. Se caracteriza por la aparicion de episodios espontaneos y repetidos de epistaxis, telangiectasias y malformaciones arteriovenosas viscerales (principalmente en el pulmon, el higado, el cerebro y el aparato digestivo), que son las responsables de sus manifestaciones clinicas y la base de los criterios de Curacao, con los que se realiza el diagnostico de la HHT. En este articulo se revisan la patogenia y las manifestaciones clinicas de esta rara enfermedad, asi como los protocolos de cribado de las lesions viscerales y las opciones terapeuticas de las que disponemos en la actualidad.: Hereditary Hemorrhagic Telangiectasia (HHT) or Rendu-Osler-Weber disease is a genetic disorder with a dominant autosomic transmission. Its prevalence is estimated in one in 5-8,000 individuals. Two different mutations have been described involving endoglin and ALK-1 genes, resulting in HHT type 1 and 2 respectively. It is characterized by the occurrence of spontaneous and recurrent episodes of epistaxis, telangiectasias and the presence of visceral arteriovenous malformations (mainly affecting lungs, liver, brain and digestive tract) which are responsible for the clinical manifestations and constitute a basic point in the diagnostic criteria of Curacao. The aim of this article is to review the pathogenesis, clinical aspects, screening procedures to disclose the visceral involvement and the therapeutic options of this rare disease.

MiR-205 is downregulated in hereditary hemorrhagic telangiectasia and impairs TGF-beta signaling pathways in endothelial cells

Hereditary hemorrhagic telangiectasia (HHT) is an autosomal dominant disorder characterized by arteriovenous malformations and hemorrhages. This vascular disease results mainly from mutations in 2 genes involved in the TGF-β pathway (ENG and ALK1) that are exclusively expressed by endothelial cells. The present study identified miR-27a and miR-205 as two circulating miRNAs differentially expressed in HHT patients. The plasma levels of miR-27a are elevated while those of miR-205 are reduced in both HHT1 and HHT2 patients compared to healthy controls. The role of miR-205 in endothelial cells was further investigated. Our data indicates that miR-205 expression displaces the TGF-β balance towards the anti-angiogenic side by targeting Smad1 and Smad4. In line, overexpression of miR-205 in endothelial cells reduces proliferation, migration and tube formation while its inhibition shows opposite effects. This study not only suggests that detection of circulating miRNA (miR-27a and miR-205) could help for the screening of HHT patients but also provides a functional link between the deregulated expression of miR-205 and the HHT phenotype.

Mutation affecting the proximal promoter of Endoglin as the origin of hereditary hemorrhagic telangiectasia type 1

BackgroundHereditary hemorrhagic telangiectasia (HHT) is a vascular multi-organ system disorder. Its diagnostic criteria include epistaxis, telangiectases in mucocutaneous sites, arteriovenous malformations (AVMs), and familial inheritance. HHT is transmitted as an autosomal dominant condition, caused in 85% of cases by mutations in either Endoglin (ENG) or Activin receptor-like kinase (ACVRL1/ACVRL1/ALK1) genes. Pathogenic mutations have been described in exons, splice junctions and, in a few cases with ENG mutations, in the proximal promoter, which creates a new ATG start site. However, no mutations affecting transcription regulation have been described to date in HHT, and this type of mutation is rarely identified in the literature on rare diseases.MethodsSequencing data from a family with HHT lead to single nucleotide change, c.-58G > A. The functionality and pathogenicity of this change was analyzed by in vitro mutagenesis, quantitative PCR and Gel shift assay. Student t test was used for statistical significance.ResultsA single nucleotide change, c.-58G > A, in the proximal ENG promoter co-segregated with HHT clinical features in an HHT family. This mutation was present in the proband and in 2 other symptomatic members, whereas 2 asymptomatic relatives did not harbor the mutation. Analysis of RNA from activated monocytes from the probands and the healthy brother revealed reduced ENG mRNA expression in the HHT patient (p = 0.005). Site-directed mutagenesis of the ENG promoter resulted in a three-fold decrease in luciferase activity of the mutant c.-58A allele compared to wild type (p = 0.005). Finally, gel shift assay identified a DNA-protein specific complex.ConclusionsThe novel ENG c.-58G > A substitution in the ENG promoter co-segregates with HHT symptoms in a family and appears to affect the transcriptional regulation of the gene, resulting in reduced ENG expression. ENG c.-58G > A may therefore be a pathogenic HHT mutation leading to haploinsufficiency of Endoglin and HHT symptoms. To the best of our knowledge, this is the first report of a pathogenic mutation in HHT involving the binding site for a transcription factor in the promoter of ENG.