Type 2 hereditary hemorrhagic telangiectasia (HHT) is a rare autosomal dominant disease and is associated with ALK1 gene mutations. Type 2 HHT patients primarily suffer from recurrent bleeding. There is currently no promising treatment.

A 5-year-old Chinese patient (III23) was admitted to Zhongshan Hospital for recurrent melena occurring over 2 mo. She had been experiencing epistaxis for years and had been diagnosed with idiopathic pulmonary hypertension 4 mo before presentation. Abdominal computed tomography examination showed hepatic arteriovenous malformation. Gene testing revealed a c.1121G>A mutation on the ALK1 gene. According to the international diagnostic criteria, this patient was diagnosed with HHT. In addition, 8 more family members exhibited HHT symptoms to varying degrees. Gene testing in 5 family members (2 with HHT symptoms and 3 without HHT symptoms) revealed the ALK1 c.1121G>A mutation in the 2 family members with HHT symptoms. This missense mutation results in the substitution of arginine for glutamine at amino acid position 374 (R374Q) in the conserved functional kinase domain of ALK1. Biological studies revealed that this mutation decreased the kinase activity of ALK1 and impeded the phosphorylation of its substrate Smad1. Moreover, the R374Q mutant downregulated the protein level of collagen-1, a fibrogenic factor, indicating abnormal fiber generation during vascular formation.

The R374Q mutant of ALK1 and its subsequent influence on fiber generation highly indicated its pathogenic role in this family with type 2 HHT. Detection of this gene mutation will facilitate early diagnosis of suspected type 2 HHT patients, and mechanistic studies will provide insights for future therapy.

Hereditary hemorrhagic telangiectasia (HHT), a rare autosomal dominant hereditary disorder, is characterized by abnormal bleeding and telangiectasia occurring predominantly in the mucosa of the nose, oral cavity, and the skin of the face and hands[1]. The underlying pathology of HHT patients is due to the lack of normal capillaries that link the artery and vein. Therefore, blood from the arteries flows directly into the veins at high pressure and ruptures the fragile sites in the shallow mucosa or deep visceral organs[2]. The common epistaxis and telangiectasia are easily discovered and treated, while the atypically silent lesions in the lung, liver, stomach, and bowel are often neglected, leading to delayed diagnosis and severe damage[1]. Constant silent bleeding can contribute to severe anemia, and abrupt massive bleeding in major organs can lead to death.

In these circumstances, early diagnosis of HHT is of extreme importance to educate patients about precautions against aberrant bleeding and prevention of detrimental outcomes. Since HHT is a dominant hereditary disease that is primarily caused by aberrant mutation of the ENG gene (type 1 HHT)[3] or the ALK1 gene (type 2 HHT)[4], gene sequencing of ENG and ALK1 is a useful method for HHT diagnosis. However, hundreds of mutations on ENG and ALK1 have been discovered[5], and mutations that are crucial for HHT occurrence have not been fully examined. Therefore, it is imperative to determine the pathogenic mutations that could be used for HHT diagnosis.

In this study, we analyzed a Chinese family with type 2 HHT and discovered a mutation, c.1121G>A (p.R374Q), in ALK1. Biological studies revealed that this mutation decreased the kinase activity of ALK1 and reduced expression of collagen-1, indicating its pathogenic role in dysregulating fiber generation during vascular formation. Hence, our study provided a potential genetic marker for type 2 HHT diagnosis, and the underlying mechanistic exploration provided the foundation of better treatments.

The patient was diagnosed with type 2 HHT according to the Curaçao criteria[9]. In addition, the patient was also diagnosed with pulmonary arterial hypertension, right heart failure, and severe iron anemia.

Omeprazole and hydrotalcite were administrated to prevent upper GI bleeding. The patient did not experience new active bleeding and was discharged.

The patient constantly experienced melena after discharge. One month later, she went to a local pulmonary hospital because of pulmonary arterial hypertension-associated shortness of breath. She experienced acute vomiting of blood as well as melena while receiving treatment at the pulmonary hospital. Scattered erythematous mucosa was identified in the stomach without typical ulcers (Figure 6). After hemostatic therapy, she was discharged. However, the patient consistently had symptoms of GI bleeding and died of heart failure 1 year later.

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Figure 6 Capsule endoscopy examination showed mucosal erosion in the stomach.

HHT is an autosomal dominant hereditary disease that occurs in 1 out of 5000 people[10]. However, the real incidence might be higher because many potential HHT patients are not diagnosed due to their varied presentation of symptoms. The HHT-related vascular malformation can occur in any organ, thereby contributing to the diversity of symptoms that vary from superficial skin telangiectasia to epistaxis to deeper organ issues, like pulmonary hypertension, GI bleeding, cerebral bleeding, etc. In addition, patients can have symptoms involving one organ or multiple organs concurrently. Patients with common epistaxis and telangiectasia are easily recognized, while those with atypically silent lesions in the lung, liver, and stomach are often neglected. In this case, the patient who was diagnosed with pulmonary hypertension was not screened for HHT until she presented with more indicative symptoms like GI bleeding.

The delayed diagnosis can be avoided by utilizing gene testing. HHT is an autosomal dominant hereditary disease[1] that is caused by gene mutations in ENG (type 1 HHT)[3], ALK1 (type 2 HHT)[4], SMAD4 (juvenile polyposis JP-HHT)[11], and BMP9 (HHT5)[12]. Over a thousand mutations have been reported within the ENG and ALK1 genes (http://arup.utah.edu/database/HHT/index.php). However, whether all the reported mutations are involved in causing HHT symptoms is unknown. Therefore, it is crucial to determine the pathogenic mutations of HHT. Identifying the diagnostic HHT pathogenic gene pool is of great value to enable patients with limited symptoms to be diagnosed in a timely manner.

Herein, we identified the mutation c.1121G>A in exon 8 of ALK1 in 3 patients in this family, which supported a diagnosis of type 2 HHT. The mutation, c.1121G>A, contributes to a substitution at amino acid position 374 (R374Q) that is located in the conserved functional kinase domain of ALK1. Typically the active loop (between amino acids 371 and 374) has a conformational change when activated and then facilitates its binding with substrates or ATP[13]. We investigated the influence of the R374Q mutation on ALK1 activity. According to our studies, the R374Q mutation did not affect protein expression or cellular localization, but it decreased the kinase activity of ALK1. It induced a lower gene product in the BRE4 luciferase reporter assay and impeded the phosphorylation of Smad1. Since the Smad1/Smad5 pathway is involved in TGF-β-mediated angiogenesis[8], the decreased level of phosphorylated Smad1 indicates that the R374Q mutant inhibited the kinase activity of ALK1 as well as downstream reactions, leading to abnormal angiogenesis. Moreover, telangiectasia is a pathological manifestation in type 2 HHT. The lack of protective fibers in vessel walls causes the abnormal dilatation. Exploration of the effect of the R374Q mutant on fibrogenesis revealed that the R374Q mutant reduced the protein level of collagen-1, a fibrogenic factor that can enhance fiber generation[14]. Hence, the R374Q mutant played a role in downregulating fibrogenesis and eventually causing vascular malformation.

According to these biomolecular studies, the R374Q mutant of ALK1 played a pathological role in HHT development in this family. The R374Q mutation was also reported in other HHT families[2,7,15-18], further demonstrating its prevalent occurrence and pathological effect in HHT. Intriguingly, most of the HHT patients with the R374Q mutation were reported experiencing hepatic AVMs and GI bleeding, suggesting a possible correlation between genotype to phenotype[2,7,15]. Besides, independent studies in different populations found that hepatic AVMs and GI bleeding were more common in patients with ALK1 mutations[19,20]. In this family, the proband suffered from GI bleeding and hepatic AVMs. Her elder brother experienced GI bleeding, while her son only experienced epistaxis. The diversity of phenotype in the son might be attributed to incomplete penetrance. Given the poor prognosis of the HHT patients carrying the R374Q mutation, this son is most likely to develop hepatic AVMs and GI bleeding when he gets older.

Unfortunately, there is no effective treatment to stop the progression of HHT. The son of the proband can only be educated on precautions against aberrant bleeding and to stop bleeding quickly to prevent detrimental outcomes. To develop novel therapeutic strategies, a transgenic mouse model is recommended to introduce specific mutations to HHT-associated genes, like c.1121G>A on the ALK1 gene, and monitor the phenotype of the HHT-mimic mice. Effective drug treatments that impede HHT progression and improve survival could be developed in this type of animal model.

Overall, we discovered a gene mutation c.1121G>A (p.R374Q) in ALK1 in a Chinese family with type 2 HHT. Detection of this mutation may facilitate early diagnosis of HHT, which would enable medical care to be implemented in a timely manner to prevent fatal damage. In addition, the study of the pathogenic mechanism of the R374Q mutant might provide insights for impeding HHT pathogenic progress and developing treatments.

We thank Ronggui Hu, Xingxing Xu, and Yonghui Tao (Institute of Biochemistry and Cell Biology, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences) for providing technical support. We thank the Yueguang Chen laboratory (Department of Biological Sciences and Bio-technology, Tsinghua University, Beijing, China) for providing the BRE4 reporter plasmid as a gift.