Identification of Alpha Thalassemia, RNF213 p.R4810K and PROC p.R189W among Children with Moyamoya Disease/Syndrome 

Lunliya Thampratankul1, Yusuke Okuno2, Patcharee Komvilaisak3, Duangrurdee Wattanasirichaigoon1 and Nongnuch Sirachainan1.

1 Department of Pediatrics, Faculty of Medicine Ramathibodi Hospital, Mahidol University, Bangkok, Thailand.
2 Department of Virology, Nagoya City University Graduate School of Medical Sciences, Nagoya, Japan.
3 Department of Pediatrics, Srinagarind Hospital, Faculty of Medicine, Khon Kaen University, Khon Kaen, Thailand.

Correspondence to: Professor Nongnuch Sirachainan, MD, Department of Pediatrics Faculty of Medicine, Ramathibodi Hospital, Mahidol University, Bangkok 10400 Thailand. Tel: +66 2 201 1749. Fax: +66 2 201 1748. E-mail: nongnuch.sir@mahidol.ac.th

Published: July 1, 2022
Received: March 31, 2022
Accepted: June 18, 2022
Mediterr J Hematol Infect Dis 2022, 14(1): e2022057 DOI 10.4084/MJHID.2022.057

This is an Open Access article distributed under the terms of the Creative Commons Attribution License
(https://creativecommons.org/licenses/by-nc/4.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

To the editor

Moyamoya is an occlusive vasculopathy characterized by progressive bilateral or unilateral stenosis/occlusion of the distal internal carotid artery and the abnormal development of a hazy network of basal collateral vessels.1 Moyamoya, classified into moyamoya syndrome and moyamoya disease, is more common in Asian than in Western countries. Moyamoya syndrome is associated with underlying conditions, such as hemoglobinopathy and post-radiation, while moyamoya disease is not.[1] The etiology of moyamoya remains unknown; however, genetic analyses identified the variable incidence rates of RNF213 p.R4810K of 23.1-90.1% in most East Asian populations.[2] The genetic variants of moyamoya reported from Southeast Asian countries are limited. Therefore, we reported the underlying disease of non-deletional α-thalassemia, antiphospholipid antibodies (APAs), and genetic variants of PROC p.R189W, and less frequent RNF213 p.R4810K in moyamoya disease/syndrome, using whole-exome sequencing (WES).
We describe a cross-sectional study involving 14 patients (7 with moyamoya syndrome and 7 with moyamoya disease), female: male 1.33:1, with a median age of 9.3 years (1.7-12.6) at diagnosis. Their median follow-up time was 6.3 years. The diagnosis was confirmed using magnetic resonance imaging and angiographic study. Recurrent hemiparesis and seizure were the most common presentations. All patients had no family history of moyamoya. Genomic DNA was extracted from peripheral blood samples and sent for WES, performed on a Novaseq 6000 System (Illumina, San Diego, CA, USA). WES data were analyzed using the commercial software, Sophia DDM, V4.4 (Sophia Genetics). This study was approved by the Ethics Committee of the Faculty of Medicine Ramathibodi Hospital (ID: COA. MURA2020/1788).   
Among the seven patients with moyamoya syndrome, three were diagnosed with non-deletional α-thalassemia disease [hemoglobin H/Constant Spring (--SEA/CS)], one with Williams syndrome, and one post cranial radiation. Two of the three α-thalassemia patients also had APAs. The WES study demonstrated a heterozygous variant of a prothrombotic gene of heterozygous PROC gene mutation (c.565C > T, p.R189W) in three patients. Two patients had low protein (PC) activities (42% and 61%). WES study also confirmed the genetic information of thalassemia [αCS (HBA2: c.427T > C)/ --SEA (NG_000006.1: g.26264_45564del19301)] among three patients. One of the three heterozygous p.R189W mutation patients also had homozygous HBB c.79G>A, p.E27K, causing mild anemia (Table 1 and Supplement Figure 1). For the seven patients with moyamoya disease, the WES study identified genetic variants of heterozygous RNF213 mutation (c.14429G>A, p.R4810K) in two patients (Table 1 and Supplement Figure 1).
Table 1.1 Table 1. Characteristics of the 14 patients with moyamoya. Patients 1-7 were classified as moyamoya syndrome, and Patients 8-14 as moyamoya disease.
Table 1.2
Neurological outcomes revealed neurologic deficits among seven patients. The modified Rankin scale (mRS) was higher in moyamoya syndrome (score 3 in craniopharyngioma post cranial radiation, score 2 in alpha thalassemia disease, and score 1 in Williams syndrome), indicating higher disability when compared with moyamoya disease (score 1 in two of seven patients).
Sickle cell anemia and β thalassemia had been reported in moyamoya syndrome.[1] The present study reported non-deletional α-thalassemia in moyamoya. The α-thalassemia disease usually involves moderate anemia and requires occasional red blood cell (RBC) transfusion. However, non-deletional α-thalassemia may exhibit more severe phenotypes, similar to patients with ꞵ-thalassemia.[3] The patients in this report, before the diagnosis of moyamoya, received occasional RBC transfusions. As a result, the high proportion of phosphatidylserine exposing RBC increased the hypercoagulable  state[4] and may contribute to the developing moyamoya. The additional prothrombotic risk factors were APAs in two α-thalassemia patients and PROC p.R189W in one patient. The p.R189W mutation, resulted in low or slightly low levels of PC activity. The related report suggested that p.R189W had a low binding affinity to endothelial PC receptors; however, some patients with p.R189W had normal PC activity.[5,6] WES demonstrated the prothrombotic genetic variants of the PROC p.R189W mutation in three patients. These findings indicated the overall prothrombotic risk factors in 35.7% of patients and 71.4% of moyamoya syndrome, including two patients with APS and three with PROC p.R189W mutation. A related study reported prothrombotic risk factors consisting of APAs and PS deficiencies in 40% of the investigated ten patients.[7]
In addition to prothrombotic risk factors, one patient in this study presented Williams syndrome associated with vascular abnormality, including peripheral pulmonary stenosis and moyamoya syndrome.[1] One patient developed moyamoya syndrome after 18 months of 54 Gy cranial radiation. Altogether, 14.2% of our reported patients and 28.6% of moyamoya disease demonstrated RNF213 p.R4810K, which was lower than that reported among Japanese (90.1%) and Korean (78.9%). Still, the same as in Chinese (23.2%) patients.[2] The incidence was higher than the prevalence in the general population (0-11.4%).[2] The RNF213 gene, located on chromosome 17q25.3, is related to angiogenesis and vascular inflammation with an unknown physiologic function.
Although the number of enrolled patients was small due to the rarity of the disease, our report demonstrated a non-deletional type α-thalassemia disease (--SEA/ α^csα), and APAs in moyamoya syndrome. In addition to the prothrombotic risk factor of genetic variants of PROC p.R189W. Moreover, the (RNF213 p.R4810K was identified in 28.6% of moyamoya disease. In addition, the RNF213 p.R4810K was identified in 28.6% of moyamoya disease.

Acknowledgements

We thank Dr. Chaiyos Khongkhatithum MD, Thipwimol Tim-Aroon MD, Duantida Songdej MD, and Pongpak Pongphitcha MD for their advice and contribution to the research. We thank the patients and their families, and all medical personnel involved in providing care to the patients. This research project is supported by the Faculty of Medicine Ramathibodi Hospital Mahidol University (Grant ID CF_63003).


References   

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Supplement Figure

Suppl Figure 1 Supplement Figure 1. Sequencing results of: (A), RNF213 c.14429G>A, p.R4810K in patients 11 and 13; and (B), PROC c.565C>T, p.R189W in patients 5, 7, and 9. Diagram (C) demonstrates the possible pathogeneses of moyamoya from this report, including genetic variants of RNF213 p.R4810K resulting in abnormal angiogenesis; PROC p.R189W resulting in increased thrombus formation, and contributing factors such as infection, inflammation, thalassemia disease, neurofibromatosis type I, cranial radiation and Williams syndrome.

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