Sánchez-Martínez et al. Orphanet Journal of Rare Diseases (2020) 15:138 https://doi.org/10.1186/s13023-020-01422-8 RESEARCH Open Access Current HHT genetic overview in Spain and its phenotypic correlation: data from RiHHTa registry Rosario Sánchez-Martínez1,2, Adriana Iriarte2,3, José María Mora-Luján2,3, José Luis Patier2,4, Daniel López-Wolf2,5, Ana Ojeda2,6, Miguel Angel Torralba2,7, María Coloma Juyol2,8, Ricardo Gil2,9, Sol Añón2,10, Joel Salazar-Mendiguchía11,12,13, Antoni Riera-Mestre2,3,14* and for the RiHHTa Investigators of the Rare Diseases Working Group from the Spanish Society of Internal Medicine Abstract Background: Hereditary hemorrhagic telangiectasia (HHT) is a rare vascular disease with autosomal dominant inheritance. Disease-causing variants in endoglin (ENG) and activin A receptor type II-like 1 (ACVRL1) genes are detected in more than 90% of cases submitted to molecular diagnosis. Methods: We used data from the RiHHTa (Computerized Registry of Hereditary Hemorrhagic Telangiectasia) registry to describe genetic variants and to assess their genotype-phenotype correlation among HHT patients in Spain. Results: By May 2019, 215 patients were included in the RiHHTa registry with a mean age of 52.5 ± 16.5 years and 136 (63.3%) were women. Definitive HHT diagnosis defined by the Curaçao criteria were met by 172 (80%) patients. Among 113 patients with genetic test, 77 (68.1%) showed a genetic variant in ACVRL1 and 36 (31.8%) in ENG gene. The identified genetic variants in ACVRL1 and ENG genes and their clinical significance are provided. ACVRL1 mutations were more frequently nonsense (50%) while ENG mutations were more frequently, frameshift (39.1%). ENG patients were significantly younger at diagnosis (36.9 vs 45.7 years) and had pulmonary arteriovenous malformations (AVMs) (71.4% vs 24.4%) and cerebral AVMs (17.6% vs 2%) more often than patients with ACVRL1 variants. Patients with ACVRL1 variants had a higher cardiac index (2.62 vs 3.46), higher levels of hepatic functional blood tests, and anemia (28.5% vs 56.7%) more often than ENG patients. Conclusions: ACVRL1 variants are more frequent than ENG in Spain. ACVRL1 patients developed symptomatic liver disease and anemia more often than ENG patients. Compared to ACVRL1, those with ENG variants are younger at diagnosis and show pulmonary and cerebral AVMs more frequently. Keywords: Hereditary hemorrhagic telangiectasia, Genetic test, Phenotype, Genotype, Rare diseases * Correspondence: ariera@bellvitgehospital.cat 2 Rare Diseases Working Group, Spanish Society of Internal Medicine, Madrid, Spain 3 Hereditary Hemorrhagic Telangiectasia Unit, Internal Medicine Department, Hospital Universitari de Bellvitge – IDIBELL, Feixa Llarga s/n. 08907 L’Hospitalet de Llobregat, Barcelona, Spain Full list of author information is available at the end of the article © The Author(s). 2020 Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons licence, and indicate if changes were made. The images or other third party material in this article are included in the article's Creative Commons licence, unless indicated otherwise in a credit line to the material. If material is not included in the article's Creative Commons licence and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this licence, visit http://creativecommons.org/licenses/by/4.0/. The Creative Commons Public Domain Dedication waiver (http://creativecommons.org/publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated in a credit line to the data. Sánchez-Martínez et al. Orphanet Journal of Rare Diseases (2020) 15:138 Page 2 of 8 Introduction developing clinical trials [19–21]. The aim of the present Hereditary hemorrhagic telangiectasia (HHT) or Rendu- study is to describe the genetic variants and their clinical Osler-Weber syndrome (ORPHA774; HHT 1: OMIM# correlation in HHT patients from the RiHHTa (Computer- 187300; HHT 2: OMIM# 600376) is a rare autosomal- ized Registry of Hereditary Hemorrhagic Telangiectasia) dominant multisystemic vascular disease characterized registry, comprehending a cohort of individuals followed- by telangiectasia and larger vascular malformations up at specialized HHT centers in Spain [14]. (VMs), with a prevalence of 1 in 6000 [1–4]. Although more than 800 pathogenic variants in more than 5 genes Material and methods have been reported, disease-causing variants in endoglin Study design (ENG; chromosome 9q) and activin A receptor type II- RiHHTa is a multicenter, prospective and observational like 1 (ACVRL1; chromosome 12q) genes are detected in registry, developed within the Rare Diseases Working more than 90% of cases submitted to molecular diagno- Group of the Spanish Society of Internal Medicine. sis and cause HHT1 and HHT2, respectively [5–7]. RiHHTa registry is currently formed by 42 researchers Mutations in SMAD4 (encoding the transcription factor from 29 Spanish hospitals. RiHHTa has an online design Smad4) have been described in less than 2% of the HHT (accessible from https://rihhta.healthincode.com) avail- population and cause juvenile polyposis/HHT overlap able in Spanish or English with individual encoded syndrome [8]. Endoglin (encoded by ENG) is an auxiliary access for each researcher. The rationale and method- co-receptor at the endothelial cell surface that promotes ology of RiHHTa have been published elsewhere [14]. BMP9 signaling through the activin receptor-like kinase Patients’ personal data included in the registry are in 1 (ALK1; encoded by ACVRL1). Both proteins contribute agree with Spanish personal data law (Ley 15/1999 de to the signaling hub formed by BMP9-Endoglin-ALK1- Protección de Datos de Carácter Personal). To safeguard Smad with a high impact in angiogenesis [9–11]. the identity of the included patients, the RiHHTa regis- HHT can be diagnosed either through molecular genetic try generate an encrypted identifier for each patient. All testing or using the Curaçao clinical criteria (recurrent epi- patients (or their relatives) provided written consent for staxis, cutaneous/mucosal telangiectasia, visceral involvement participation in the registry. The design of the RiHHTa and a first-degree relativewith HHT) [4, 5, 7, 12]. However, registry was approved by the Ethics Committee of the since HHT is an age-dependent disease, genetic study is a Hospital Universitari de Bellvitge (Barcelona, Spain; ethic very useful tool for diagnosis in young patients [4, 5, 7]. approval number PR241/16). The creation of RiHHTa Telangiectasia is the characteristic lesion in HHT and shows counted on the nonremunerated collaboration of the dilated postcapillary venules directly connected with dilated genetic studies company Health in Code (A Coruña, arterioles losing the capillary bed [7, 13]. Telangiectasia in Spain), which has broad experience in cardiomyopathy nasal mucosae causes spontaneous and recurrent epistaxis registries [22]. that are the most common and usually the earliest clinical The main objective of the present study was to describe manifestation of HHT [7, 12, 14]. Although endoglin and the type of genetic variants identified in HHT patients ALK1 are components of the same BMP9 receptor complex, included in RiHHTa registry and to analyse the genotype- they are structurally and functionally different proteins. Thus, phenotype correlation according to these mutations in pathogenic variants in their genes are related to different ENG or ACVRL1 genes. clinical phenotypes [5, 9]. Pulmonary arteriovenous malfor- mations (AVMs) and brain VMs are more common in Patient and variables patients with HHT1, while vascular hepatic malformations Patients included in the RiHHTa registry with a “defin- are more frequent in HHT2 [4, 7, 15]. ite” clinical diagnosis according to the Curaçao Criteria Of note, there is high inter-familial as well as intra- (≥ 3 criteria) or with a genetic diagnosis were considered familial vascular involvement and clinical manifestations for this study [12]. The inclusion period was from June variability [7, 16]. Therefore, genetic study is essential 2016 to June 2019. Demographic data, clinical character- for diagnostic confirmation and also for relatives’ genetic istics and complementary tests were collected according screening [5, 7, 17]. This low prevalence and clinical to mutations in ENG or ACVRL1 genes. The severity of variability makes HHT understanding challenging, espe- nosebleeds was measured according to the epistaxis cially in uncommon situations [18]. Development of severity score (ESS). ESS is an on-line tool that estimates registries that include clinical and genetic data may help the severity of epistaxis according to clinical data occur- to overcome these difficulties, and represent a priority in ring during the last 3 months. Epistaxis is considered the management of rare diseases [19, 20]. The use of moderate or severe if ESS results > 4 or > 7 points, quality registries reflects patients’ management in standard respectively [23]. clinical practice, helps reaching a consensus on diagnostic- For the screening of pulmonary visceral involve- therapeutic measures and also generates hypotheses for ment, a contrast transthoracic echocardiography Sánchez-Martínez et al. Orphanet Journal of Rare Diseases (2020) 15:138 Page 3 of 8 (TTE) was performed to establish the degree of Student t-test. We used two-tailed unpaired t-tests to right-left shunt (R-L) and the need to undergo a thoracic compare normally distributed continuous data between computed tomography (CT) angiography to objectively two groups, and we used the Mann-Whitney U test for confirm the presence of pulmonary arteriovenous malfor- non-normally distributed continuous data comparisons. mations (AVM) [7, 24]. In addition, an abdominal CT Normality was assessed using Shapiro-Wilk test. P values angiography, magnetic resonance imaging (MRI) or dop- of < 0.05 were considered statistically significant. Ana- pler ultrasound were performed to study hepatic and/or lyses were performed using SPSS, version 18 for the PC abdominal VMs. Hepatic involvement was defined accord- (SPSS, Inc. Chicago, IL, USA). ing to the three classical patterns of abnormal vascular communications: arteriovenous shunt (from hepatic artery Results to hepatic vein), portovenous shunt (from portal vein to Baseline characteristics hepatic vein) and arterioportal shunt (from hepatic artery During the study period, 215 patients from 29 Spanish to portal vein) [25]. Neurological involvement studies hospitals were included in the RiHHTa registry, 136 were carried out when neurological symptoms or family (63.3%) were women. Mean age at time of inclusion of history, by a cerebral CT and/or MRI. An endoscopic 52.5 ± 16.5 years old. Most (88.4%) patients were Cauca- digestive study was performed when there was dispropor- sian. The mean age at diagnosis of HHT was 42.2 ± 17.5 tionate anaemia to the degree of epistaxis or objectively years old. Epistaxis was the most frequent symptom at confirmed overt gastrointestinal (GI) bleeding [7]. disease onset which allowed the diagnosis of suspicion in 197 (91.6%) patients, while in another 7 (3.2%) Molecular analysis patients the diagnosis arose after a central nervous sys- All included patients underwent genetic testing. The se- tem event (including brain abscess in 4 (1.9%) patients, quencing technique depended on each treating centers’ and stroke, in 3 (1.4%). Family history were present in preference. Because RiHHTa includes patients from all 178 (82.8%) patients and in 167 (77.7%), muco- around Spain, different genetic techniques have been used cutaneous telangiectasia were observed at diagnosis or in the included patients. These techniques include Next during the follow-up. Definite HHT diagnosis defined by generation sequencing (NGS) technologies, Sanger sequen- the Curaçao criteria were met by 172 (80%) patients, cing and other techniques such Multiplex ligation- while the remaining were diagnosed through a positive dependent probe amplification (MLPA). In patients in genetic test, after a thorough familial evaluation. . which NGS was used, coding exons and intronic boundar- Epistaxis was also the most frequent symptom at diag- ies of 9 genes related to HHT disease or HHT-like pheno- nosis of HHT with a mean ESS of 3.65 ± 2.5. Anemia types (ACVRL1, BMP10, BMPR1A, BMPR2, ENG, GDF2, was present in 92 (42.8%) patients, 12 (5%) patients had RASA1, SMAD1, SMAD4) were captured using a custom a history of previous venous thromboembolic disease probe library (SureSelect Target Enrichment Kit for Illu- and 8 (3.7%) patients had atrial fibrillation. Medical at- mina paired-end multiplexed sequencing method; Agilent tention because of epistaxis at Emergency Department Technologies, Santa Clara, California, USA), and se- during follow-up was required by 69 (32.1%) patients quenced on the HiSeq 1500 platform following Illumina and red blood cell transfusion by 38 (17.7%) patients. protocols. To determine the pathogenicity of the included Pulmonary AVMs were detected in 48 (22.3%) patients variants in the RiHHTa registry, all mutations were and liver VMs identified by imaging test (CT angiog- checked again by Health in Code SL (A Coruña, Spain) raphy, MRI or doppler ultrasound) were observed in 58 and labeled according to the American College of Medical (27%) patients. Pancreatic involvement was detected in Genetics and Genomics (ACMG) guidelines [26]. When- seven (3.3%) subjects and other abdominal VMs in 16 ever a genetic variant was identified through NGS, Sanger (7.4%) patients, including splenic, renal and gastro- sequencing was the preferred used method for familial omental arterial aneurysms. Brain malformations were evaluation, unless technically not feasible. reported in five patients (2.3%) and GI telangiectasia were observed in 26 (12.1%) patients (Table 1). Statistical analysis A descriptive statistical analysis was performed for all Types of pathogenic variants categorical and continuous variables and expressed as A genetic test was performed in 113 HHT patients from proportions or means with standard deviations (SD), re- the RiHHTa registry (Table 2). For the mutation ana- spectively. The descriptive analysis was performed for lysis, Sanger sequencing was performed in 65 (57.5%) the entire sample. We used chi-square or Fisher’s exact cases, NGS in 38 (33.6%) and other techniques, such as test to compare categorical data between groups; Fisher’s MLPA, in 10 (8.8%) cases. A genetic variant was identi- exact test has been used when expected frequencies were fied in ACVRL1 gene in 77 (68.1%) patients, whereas 36 less than 5. Continuous variables were compared using (31.8%) patients harboured a genetic variant in ENG Sánchez-Martínez et al. Orphanet Journal of Rare Diseases (2020) 15:138 Page 4 of 8 Table 1 Baseline characteristics Table 2 Summary of the variants analysis n (%) or mean (SD) GENE mutation ENG, n (%) ACVRL1, n (%) Patients, n 215 Patients, n 36 77 Sex, n (%) Sequencing method, n (%) Male 79 (36.7) Sanger 20 (55.5) 45 (58.4) Female 136 (63.3) NGS 12 (33.3) 26 (33.8) Age at HHT diagnosis (years), mean ± SD 42.2 ± 17.5 Others 4 (11.1) 6 (7.8) Ethnicity Variant type, n (%) Caucasian 190 (88.4) Hispanic 2 (0.9) Nonsense 10 (27.7) 39 (50.6) Others 9 (4.2) Frameshift 13 (36.1) 20 (25.9) No clinical data 14 (6.5) Splice-site 4 (11.1) 3 (3.9) Underlying conditions, n (%) Missense 9 (25) 15 (19.4) Current smoker 26 (12.1) NGS Next-generation sequencing Hypertension 43 (20) Diabetes Mellitus 11 (5.1) gene. According to the type of mutation, ACVRL1 patients Dyslipidemia 33 (15.3) presented more frequently nonsense mutations (50.6%), Venous Thromboembolism 12 (5.5) while in ENG patients the most frequent type was frame- Atrial fibrillation 8 (3.7) shift (36.1%). Supplementary Table 1 shows those 52 vari- HHT criteria, n (%) ants we have complete genetic data, as well as their 4 HHT criteria 99 (46) frequencies in gnomAD and the bioinformatic predictor’s 3 HHT criteria 73 (34) evaluation. Supplementary Table 2 shows the ACMG clas- 2 or less HHT criteria 32 (14.9) sification of these variants. Among these variants, we iden- No clinical data 11 (5.1) tified 17 missense variants in ACVRL1, 41% of them were localized in the kinase domains of exons 7 and 8, and 29% Family history in exon 3. However, missense variants in ENG gene were Positive 178 (82.8) dispersed among the extracellular domain of the protein. Negative 11 (5.1) In both genes, the predominant variants were the ones No clinical data 26 (12.1) that caused loss-of-function mechanism (nonsense, frame- Symptoms at onset, n (%) shift and splice-site), which would almost certainly be Nosebleeds 197 (91.6) pathogenic in these genes. Figure 1 shows a schematic Central Nervous system event 7 (3.2) representation of ACVRL1 and ENG, as well as the exonic Cerebral abscess 4 (1.9) variants identified. Stroke 3 (1.4) Cutaneous telangiectases 1 (0.5) Genotype-phenotype correlation Dyspnea 1 (0.5) There were no significant differences between ENG and Anemia 1 (0.5) ACVRL1 patients in the age of inclusion, gender, num- No clinical data 8 (3.7) ber of Curaçao criteria, recurrent epistaxis, ESS, muco- Muco-cutaneous telangiectasia, n (%) 167 (77.7) cutaneous telangiectasia, family history, hepatic VMs or Baseline ESS 3.65 (2.5) GI involvement. Compared to patients with ACVRL1 mutations, ENG Gene mutation, n (%) patients were significantly younger at diagnosis (36.9 vs ENG 36 (16.7) 45.7 years) and usually had more pulmonary AVMs (71.4 ACVRL1 77 (35.8) vs 24.4%), as well as cerebral AVMs (17.6% vs 2%). No clinical data 102 (47.2) Those with ACVRL1 mutation had a higher cardiac Visceral involvement, n (%) index (3.46 L/min/m2 vs 2.62 L/min/m2), higher levels of Pulmonary AVM 48 (22.3) hepatic functional blood tests and more frequently Cerebral AVM 5 (2.3) anemia (56.7% vs 28.5%) than ENG patients (Table 3). Hepatic involvement 58 (27) Gastrointestinal involvementa 26 (12.1) Discussion HHT Hereditary hemorrhagic telangiectasia, SD Standard deviation, ESS Epistaxis The present study using data from the RiHHTa registry Severity Score, AVM Arteriovenous malformation a Gastrointestinal involvement detected by fibrogastroscopy and / or colonoscopy represents the largest series about genetic variants of HHT patients in Spain. A previous study reported 74 Sánchez-Martínez et al. Orphanet Journal of Rare Diseases (2020) 15:138 Page 5 of 8 Fig. 1 Schematic representation of the exons (boxes) of ACVRL1 and ENG genes. The identified exonic variants in our cohort are presented according to their location (colored boxes represent specific functional domains, each one of them is explained in the legends) clinically diagnosed Spanish patients, most of them with In our study, there were no statistically significant dif- ACVRL1 variants [27]. In our cohort, 77 out of 113 ferences in the presence of GI telangiectasia between (68.1%) patients also had variants in the ACVRL1 gene. ENG and ACVRL1 patients. Canzonieri et al. and van This finding is in agreement with previous data from Tuyl et al. systemically studied the extent of GI involve- other Mediterranean countries but different to Northern ment with gastroscopy, video capsule endoscopy, and Europe or North America, where pathogenetic variants colonoscopy in 22 and 35 HHT patients, respectively, in ENG are more frequent than in ACVRL1 [28, 29]. and found a higher prevalence of telangiectasia in pa- We have observed that patients with ENG gene vari- tients with ENG mutation [8, 35, 36]. In a recent study, ants were diagnosed at a younger age. This finding could Mora-Luján et al. reported that age, tobacco use, ENG be influenced because oral and nasal mucosal telangiec- mutation, and hemoglobin levels were associated with tases are present earlier in life in patients with HHT1 GI involvement in HHT comparing 67 patients with GI than with HHT2, so epistaxis also start earlier in these disease and 28 with negative GI study [37]. However, patients [30]. According to previous studies, we have ob- Berg et al., Sabbà et al. and Letteboer et al. assessing served a higher prevalence of pulmonary AVMs and genotype-phenotype relationship in HHT patients, re- cerebral VMs in patients with ENG mutations than in ported a similar incidence of GI telangiectasia in HHT1 those with ACVRL1 mutations (5–29%) [20, 23, 27, 31]. and HHT2 patients [38–40]. Hepatic involvement has been described more com- Similar to previous studies, the most frequent type of monly among ACVRL1 patients, with a wide range of pathogenic variants we have identified in both ENG and prevalence from 8 to 78% [29, 30, 32, 33]. In our series, ACVRL1 genes were those predicted to cause a loss-of- though vascular hepatic involvement was more frequent function (nonsense, frameshift and splice-site) [19, 24]. in ACVRL1 patients, no statistically significant differences Missense variants in ACVRL1 gene were predominantly were detected between ENG and ACVRL1 patients, prob- localized in the kinase domains of exons 7 and 8, while ably because different screening methods were used by missense variants in ENG gene were dispersed in the extra- RiHHTa investigators. However, patients with ACVRL1 cellular domain of the protein [27]. In a study conducted by mutation had a higher CI and levels of hepatic functional Sabbà et al., the correlation between phenotype and variant blood tests than ENG patients, suggesting a more severe type was analysed in a cohort of 77 HHT2 patients. hepatic involvement in patients with ACVRL1 mutations. ACVRL1 patients were divided into three subgroups In fact, in two studies including patients with genotype according to the type mutation: truncating mutations (20 data available, those who developed symptomatic liver patients), missense mutations (47 patients) and the c.626- disease had ACVRL1 mutation [15, 34]. 3C > G splicing mutation (10 patients). Patients with c.626- Sánchez-Martínez et al. Orphanet Journal of Rare Diseases (2020) 15:138 Page 6 of 8 Table 3 Genotype-phenotype correlation in ENG and ACVRL1 Table 3 Genotype-phenotype correlation in ENG and ACVRL1 patients patients (Continued) GENE Mutation ENG ACVRL1 p-value GENE Mutation ENG ACVRL1 p-value Patients, n 36 77 Cerebral AVM, n (%) 3 (8.3) 1 (1.3) < 0.005 Sex, n (%) Hepatic involvement, n (%) 9 (25) 33 (42.8) 0.075 Male 13 (36) 23 (29.9) 0.507 AV shunt 2 (5.5) 15 (19.4) 0.128 Female 23 (64) 54 (70.1) PV shunt 3 (8.3) 2 (2.6) 0.078 Age at diagnosis (years), 36.9 ± 17.9 45.7 ± 16.8 0.036 AP shunt 1 (2.7) 9 (11.7) 0.385 mean ± SD FNH 0 (0) 2 (2.6) 1 Ethnicity, n (%) NRH 0 (0) 1 (1.3) 1 Caucasian 36 (100) 67 (87) 0.218 Telangiectasia 4 (11.1) 20 (25.9) 0.413 Hispanic 0 (0) 5 (6.5) Liver function test (IU/L), Others/unknown 0 (0) 5 (6.5) mean ± SD Hypertension, n (%) 8 (22.2) 23 (29.9) 0.133 Aspartate aminotransferase 11.2 ± 10.4 19.7 ± 11.6 0.004 Diabetes Mellitus, n (%) 1 (2.7) 9 (11.7) 0.146 Alanine transaminase 11.5 ± 11.3 19.4 ± 12.5 0.010 Dyslipidaemia, n (%) 7 (19.4) 22 (28.5) 0.561 Gamma-glutamyl transferase 11.2 ± 12.8 43.4 ± 71.3 0.004 Venous Thromboembolism, 4 (11.1) 4 (5.2) 0.269 Alkaline phosphatase 35.4 ± 37.1 68.5 ± 69 0.026 n(%) Bilirubin 5.61 ± 8.7 5.19 ± 19.7 0.92 Atrial fibrillation, n (%) 2 (5.5) 6 (7.8) 0.588 Pancreatic involvement, 0 (0) 6 (7.8) 0.302 HHT criteria, n (%) 0.130 n (%) 4 HHT criteria 23 (63.8) 38 (49.3) Gastrointestinal involvement, n (%) 3 HHT criteria 9 (25) 27 (35.1) Gastroscopy 8 (22.2) 9 (11.7) 0.603 2 or less HHT criteria 4 (11.1) 12 (15.6) Colonoscopy 3 (8.3) 3 (3.9) 0.622 Family history, n (%) AV Arteriovenous (from hepatic artery to hepatic vein), PV Portovenous shunt Positive 34 (94.4) 73 (94.8) 0.626 (from portal vein to hepatic vein), AP Arterioportal (from hepatic artery to Negative 1 (2.7) 2 (2.6) portal vein). NRH Nodular regenerative hyperplasia, FNH Focal nodular hyperplasia, SD Standard deviation, R-L Right-left shunt Unknown 1 (2.7) 2 (2.6) Symptoms at onset, n (%) 0.461 3C > G mutation had a higher prevalence of pulmonary AVMs than the other two subgroups [29]. This data could Nosebleeds 32 (88.9) 65 (84.4) be of interest for the evaluation of variants of uncertain Cerebral abscess 1(2.7) 0 (0) significance and for defining useful prediction criteria based Others 2 (5.5) 5 (6.5) on the type of mutation. Genetic study, including the No clinical data 1 (2.7) 9 (11.7) description of new pathogenic variants, usually avoid un- Muco-cutaneous 29 (80.5) 62 (80.5) 0.645 necessary complementary tests, especially in patients with telangiectasia, n (%) no family history or those who meet less than 3 Curaçao Cerebral abscess, n (%) 2 (5.5) 0 (0) 0.132 criteria, as young patients [4, 5, 7, 33]. In fact, genetic study ESS basal, mean ± SD 3.45 ± 2.5 3.51 ± 2.3 0.94 has been found preferable to the clinical Curaçao criteria Anemia, n (%) 10 (27.7) 38 (49.3) 0.026 among patients between 0 and 21 years [41]. Our study has some limitations and strengths that Cardiac index (L/min/m2), 2.62 ± 0.7 3.46 ± 0.79 0.021 mean ± SD should be mentioned. First, it is an observational study from RiHHTa database. Thus, some non-measured vari- Contrast TTE (R-L shunt grade), < 0.005 n (%) ables could lead to a possible bias. Second, genetic and 0 2 (5.5) 28 (36.4) screening tests varied with local practice. Third, a variety of clinicians entered data into the registry, which may 1 12 (33.3) 17 (22.1) lend itself to potential inaccuracies in the data being re- 2 10 (27.7) 4 (5.2) ported. However the information obtained through 3 4 (11.1) 3 (3.9) RiHHTa registry is valuable and allows thorough under- 4 1 (2.7) 0 (0) standing the natural history of HHT. Moreover, patients Unknown 1 (2.7) 1 (1.3) in the RiHHTa registry were from different centers in Pulmonary AVM, n (%) 20 (55.5) 11 (14.3) < 0.005 Spain, which can better reflect epidemiology data of the disease in this country. Sánchez-Martínez et al. Orphanet Journal of Rare Diseases (2020) 15:138 Page 7 of 8 Conclusions Competing interests In conclusion, ACVRL1 variants are most frequent than The authors declare that they have no conflict of interest. ENG variants in Spain. A description of these identified Author details genetic variants and their clinical significance is pro- 1 Internal Medicine Department, Hospital General Universitario de Alicante - vided. Patients with ACVRL1 variants developed symp- ISABIAL, Alicante, Spain. 2Rare Diseases Working Group, Spanish Society of Internal Medicine, Madrid, Spain. 3Hereditary Hemorrhagic Telangiectasia tomatic liver disease and anemia more usually than ENG Unit, Internal Medicine Department, Hospital Universitari de Bellvitge – patients. Compared to ACVRL1, those with ENG vari- IDIBELL, Feixa Llarga s/n. 08907 L’Hospitalet de Llobregat, Barcelona, Spain. 4 ants were younger at diagnosis and show pulmonary and Department of Internal Medicine, Systemic and Orphan Diseases Unit, University Hospital Ramón y Cajal, University of Alcalá, IRYCIS, Madrid, Spain. cerebral AVMs more frequently. The RiHHTa registry 5 Internal Medicine Department, Hospital Universitario Fundación Alcorcón, can contribute to improve knowledge and management Madrid, Spain. 6Internal Medicine Department, Hospital Insular Universitario of HHT patients. de Gran Canaria, Gran Canaria, Spain. 7Internal Medicine Department, Hospital Clínico Universitario Lozano Blesa, Zaragoza, Spain. 8Internal Medicine Department, Hospital Universitario Miguel Servet, Zaragoza, Spain. 9 Supplementary information Internal Medicine Department, Hospital La Fe, Valencia, Spain. 10Internal Supplementary information accompanies this paper at https://doi.org/10. Medicine Department, Hospital Arnau de Vilanova, Valencia, Spain. 11Health 1186/s13023-020-01422-8. in Code, A Coruña, Spain. 12Clinical Genetics Program, Hospital Universitari de Bellvitge – IDIBELL, Barcelona, Spain. 13Genetics Department, Universitat Autònoma de Barcelona, Barcelona, Spain. 14Faculty of Medicine and Health Additional file 1: Table S1. Identified genetic variants in ACVRL1 and Sciences, Universitat de Barcelona, Barcelona, Spain. ENG genes. Table S2. Clinical significance of the identified variants according to ACMG criteria. Received: 19 February 2020 Accepted: 27 May 2020 Abbreviations ACVRL1: Activin A receptor type II-like 1; AVMs: Arteriovenous malformations; References CNS: Central nervous system; CT: Computed tomography; ENG: Endoglin; 1. The portal for rare diseases and orphan drugs. Available online: https:// ESS: Epistaxis severity score; GI: Gastrointestinal; HHT: Hereditary hemorrhagic www.orpha.net/consor/cgi-bin/index.php. Accessed 23 Nov 2019. telangiectasia; MLPA: Multiplex ligation-dependent probe amplification; 2. Kjeldsen AD, Vase P, Green A. Hereditary hemorrhagic telangiectasia (HHT): MRI: Magnetic resonance imaging; R-L: Right-left; SD: Standard deviations; a population-based study of prevalence and mortality in Danish HHT TTE: Transthoracic echocardiography; VMs: Vascular malformations patients. J Intern Med. 1999;245:31–9. 3. Donaldson JW, McKeever TM, Hall IP, Hubbard RB, Fogarty AW. The UK Acknowledgements prevalence of hereditary haemorrhagic telangiectasia and its association We express our gratitude to Health in Code (A Coruña, Spain) for supporting with sex, socioeconomic status and region of residence: a population-based this Registry. We also thank the patients enrolled in RiHHTa registry. study. Thorax. 2014;69:161–7. Coordinator of the RiHHTa Registry: Riera-Mestre A. 4. Shovlin CL, Buscarini E, Kjeldsen AD, Mager HJ, Sabba C, Droege F, et al. Members of the RiHHTa Registry of the Rare Diseases Working Group from European reference network for rare vascular diseases (VASCERN) outcome the Spanish Society of Internal Medicine: measures for hereditary Haemorrhagic telangiectasia (HHT). Orphanet J Rare Alonso-Cotoner C, Añón S, Beneyto M, MM Bermejo-Olano, Cerdà P, Dis. 2018;13:136. Cruellas F, De Los Santos A, Díez L, Fernández A, García-Morillo JS, Gil R, 5. McDonald J, Wooderchak-Donahue W, VanSant WC, Whitehead K, Gómez-Cerezo JF, Gómez del Olmo V, González-García A,Iriarte A, Iglesias Stevenson DA, Bayrak-Toydemir P. Hereditary hemorrhagic telangiectasia: P, Juyol MC, López-Osle N, López M, López-Wolf D, Mora-Luján JM, genetics and molecular diagnostics in a new era. Front Genet. 2015;6:1. Moreno M, Ojeda A, Patier JL, Pérez de León JA, Perez ML, Riera-Mestre 6. Prigoda NL, Savas S, Abdalla SA, Piovesan B, Rushlow D, Vandezande K, et al. A, Rivera S, Rodríguez S, Sánchez-Martínez R, Torralba MA, Zarrabeitia R. Hereditary haemorrhagic telangiectasia: mutation detection, test sensitivity and novel mutations. J Med Genet. 2006;43:722–8. 7. Faughnan ME, Palda VA, Garcia-Tsao G, Geisthoff UW, McDonald J, Proctor Authors’ contributions DD, et al. HHT Foundation International - Guidelines Working Group RSM, AI and ARM conceptualized and designed the study; all authors International guidelines for the diagnosis and management of hereditary acquired the data; RSM, AI, JSM and ARM analysed the data and interpreted haemorrhagic telangiectasia. J Med Genet. 2011;48:73–87. the findings; AI and ARM performed statistical analysis; JSM reviewed genetic 8. Gallione CJ, Repetto GM, Legius E, Rustgi AK, Schelley SL, Tejpar S, et al. A data; all authors contributed to the draft of the manuscript, read and combined syndrome of juvenile polyposis and hereditary haemorrhagic approved the final version of the manuscript. telangiectasia associated with mutations in MADH4 (SMAD4). Lancet. 2004; 363:852–9. Funding 9. Tillet E, Bailly S. Emerging roles of BMP9 and BMP10 in hereditary No funding was provided for this study. hemorrhagic telangiectasia. Front Genet. 2015;5:456. 10. Alsina-Sanchís E, García-Ibáñez Y, Figueiredo AM, Riera-Domingo C, Figueras Availability of data and materials A, Matias-Guiu X, et al. ALK1 loss results in vascular hyperplasia in mice and The datasets used and/or analysed during the current study are available humans through PI3K activation. Arterioscler Thromb Vasc Biol. 2018;38: from the corresponding author on reasonable request. 1216–29. 11. Iriarte A, Figueras A, Cerdà P, Mora JM, Jucglà A, Penín R, et al. PI3K (Phosphatidylinositol 3-Kinase) Activation and Endothelial Cell Proliferation Ethics approval and consent to participate in Patients with Hemorrhagic Hereditary Telangiectasia Type 1. Cells. 2019; The study was approved by the Clinical Research Ethics Committee of the 8(9):971. Hospital Universitari de Bellvitge (Barcelona, Spain; ethic approval number 12. Shovlin CL, Guttmacher AE, Buscarini E, Faughnan ME, Hyland RH, PR241/16). Westermann CJ, et al. Diagnostic criteria for hereditary hemorrhagic telangiectasia (Rendu- Osler-weber syndrome). Am J Med Genet. 2000; Consent for publication 91:66–7. This manuscript does not contain any individual person’s data in any form. 13. Braverman IM, Keh A, Jacobson BS. Ultrastructure and three-dimensional Each patient was identified by a unique alphanumeric identification code organization of the telangiectases of hereditary hemorrhagic telangiectasia. and all data were made anonymous and analysed as aggregates. J Invest Dermatol. 1990;95:422–7. Sánchez-Martínez et al. Orphanet Journal of Rare Diseases (2020) 15:138 Page 8 of 8 14. Riera-Mestre A, Mora Luján JM, Sanchez Martínez R, Torralba Cabeza MA, de hemorrhagic telangiectasia and correlation with their genotypes. Genet la Peña JL P, Juyol Rodrigo MC, et al. Computerized registry of patients with Med. 2014;16:3–10. hemorrhagic hereditary telangiectasia (RiHHTa Registry) in Spain: Objectives, 36. Van Tuyl SA, Letteboer TG, Rogge-Wolf C, Kuipers EJ, Snijder RJ, methods, and preliminary results. Rev Clin Esp. 2018;218:468–76. Westermann CJ, et al. Assessment of intestinal vascular malformations in 15. Lesca G, Olivieri C, Burnichon N, Pagella F, Carette M-F, Gilbert-Dussardier B, patients with hereditary hemorrhagic teleangiectasia and anemia. Eur J et al. Genotype-phenotype correlations in hereditary hemorrhagic Gastroenterol Hepatol. 2007;19:153–8. telangiectasia: data from the French-Italian HHT network. Genet Med. 2007; 37. Mora-Luján JM, Iriarte A, Alba E, Sánchez-Corral MA, Berrozpe A, Cerdà P, et al. 9:14–22. Gastrointestinal Bleeding in Patients with Hereditary Hemorrhagic 16. McDonald J, Bayrak-Toydemir P, Pyeritz RE. Hereditary hemorrhagic Telangiectasia: Risk Factors and Endoscopic Findings. J Clin Med. 2019;9(1):82. telangiectasia: an overview of diagnosis, management and pathogenesis. 38. Berg J, Porteous M, Reinhardt D, Gallione C, Holloway S, Umasunthar T, et al. Genet Med. 2011;13:607–16. Hereditary haemorrhagic telangiectasia: a questionnaire based study to 17. Van Gent MW, Velthuis S, Post MC, Snijder RJ, Westermann CJ, Letteboer TG, delineate the different phenotypes caused by endoglin and ALK1 et al. Hereditary hemorrhagic telangiectasia: how accurate are the clinical mutations. J Med Genet. 2003;40:585–90. criteria? Am J Med Genet A. 2013;161A:461–6. 39. Sabbà C, Pasculli G, Lenato GM, Suppressa P, Lastella P, Memeo M, et al. 18. Riera-Mestre A, Mora-Luján JM, Trujillo-Santos J, Del Toro J, Nieto JA, Hereditary hemorrhagic telangiectasia: clinical features in ENG and ALK1 Pedrajas JM, RIETE Investigators, et al. Natural history of patients with mutation carriers. J Thromb Haemost. 2007;5:1149–57. venous thromboembolism and hereditary hemorrhagic telangiectasia. 40. Letteboer TG, Mager JJ, Snijder RJ, Koeleman BP, Lindhout D, Ploos van Findings from the RIETE registry. Orphanet J Rare Dis. 2019;14:196. Amstel JK, et al. Genotype-phenotype relationship in hereditary 19. D’Agnolo HM, Kievit W, Andrade RJ, Karlsen TH, Wedemeyer H, Drenth JP. haemorrhagic telangiectasia. J Med Genet. 2006;43:371–7. Creating an effective clinical registry for rare diseases. Unit Eur Gastroenterol 41. Pahl KS, Choudhury A, Wusik K, Hammill A, White A, Henderson K, et al. J. 2016;4:333–8. Applicability of the Curaçao criteria for the diagnosis of hereditary hemorrhagic 20. EURORDIS-NORD-CORD Joint Declaration of 10 Key Principles for Rare telangiectasia in the pediatric population. J Pediatr. 2018;197:207–3. Disease Patient Registries. Available from: http://download.eurordis.org/ documents/pdf/EURORDIS_NORD_CORD_JointDec_Registries_FINAL.pdf. Accessed 15 Sept 2019. Publisher’s Note Springer Nature remains neutral with regard to jurisdictional claims in 21. Kodra Y, Posada de la Paz M, Coi A, Santoro M, Bianchi F, Ahmed F, et al. published maps and institutional affiliations. Data quality in rare diseases registries. Adv Exp Med Biol. 2017;1031:149–64. 22. Torrado M, Maneiro E, Trujillo-Quintero JP, Evangelista A, Mikhailov AT, Monserrat L. A novel heterozygous Intronic mutation in the FBN1 gene contributes to FBN1 RNA Missplicing events in the Marfan syndrome. Biomed Res Int. 2018;2018:3536495. 23. Hoag JB, Terry P, Mitchell S, Reh D, Merlo CA. An epistaxis severity score for hereditary hemorrhagic telangiectasia. Laryngoscope. 2010;120:838–43. 24. Barzilai B, Waggoner AD, Spessert C, Picus D, Goodenberger D. Two- dimensional contrast echocardiography in the detection and follow-up of congenital pulmonary arteriovenous malformations. Am J Cardiol. 1991;68: 1507–10. 25. Khalid SK, Garcia-Tsao G. Hepatic vascular malformations in hereditary hemorrhagic telangiectasia. Semin Liver Dis. 2008;28:247–58. 26. Richards S, Aziz N, Bale S, Bick D, Das S, Gastier-Foster J, et al. ACMG laboratory quality assurance committee. Standards and guidelines for the interpretation of sequence variants: a joint consensus recommendation of the American College of Medical Genetics and Genomics and the Association of Molecular Pathology. Genet Med. 2015;17:405–24. 27. Fontalba A, Fernandez-L A, García-Alegria E, Albiñana V, Garrido-Martin EM, Blanco FJ, et al. Mutation study of Spanish patients with hereditary hemorrhagic telangiectasia. BMC Med Genet. 2008;9:75. 28. Olivieri C, Pagella F, Semino L, Lanzarini L, Valacca C, Pilotto A, et al. Analysis of ENG and ACVRL1 genes in 137 HHT Italian families identifies 76 different mutations (24 novel). Comparison with other European studies. J Hum Genet. 2007;52:820–9. 29. Johnson DW, Berg JN, Baldwin MA, Gallione CJ, Marondel I, Yoon SJ, et al. Mutations in the activin receptor-like kinase 1 gene in hereditary haemorrhagic telangiectasia type 2. Nat Genet. 1996;13:189–95. 30. Letteboer TGW, Mager H-J, Snijder RJ, Lindhout D, Ploos van Amstel H-K, Zanen P, Westermann KJJ. Genotype–phenotype relationship for localization and age distribution of telangiectases in hereditary hemorrhagic telangiectasia. Am J Med Genet. 2008;146A:2733–9. 31. Bayrak-Toydemir P, McDonald J, Markewitz B, Lewin S, Miller F, Chou LS, et al. Genotype-phenotype correlation in hereditary hemorrhagic telangiectasia: mutations and manifestations. Am J Med Genet A. 2006;140:463–70. 32. Buscarini E, Gandolfi S, Alicante S, Londoni C, Manfredi G. Liver involvement in hereditary hemorrhagic telangiectasia. Abdom Radiol (NY). 2018;43:1920–30. 33. Riera-Mestre A, Ribas J, Castellote J. Medical management of haemorrhagic hereditary telangiectasia in adult patients. Med Clin (Barc). 2019;152:274–80. 34. Singh S, Swanson KL, Hathcock MA, Kremers WK, Pallanch JF, Krowka MJ, et al. Identifying the presence of clinically significant hepatic involvement in hereditary haemorrhagic telangiectasia using a simple clinical scoring index. J Hepatol. 2014;61:124–31. 35. Canzonieri C, Centenara L, Ornati F, Pagella F, Matti E, Alvisi C, et al. Endoscopic evaluation of gastrointestinal tract in patients with hereditary