RNF213 variant in a patient with Legius syndrome associated with moyamoya syndrome

Abstract

Neurofibromatosis type 1 (NF1) is a clinically heterogeneous neurocutaneous genetic disorder that may occasionally be associated with various cerebrovascular anomalies, including moyamoya syndrome (Dlamini et al., 2019; Guey et al., 2015; Sam et al., 2015). A similar vascular involvement has been very recently reported in a patient with Legius syndrome, a rare condition characterized by caféaulait spots and macrocephaly either with or without axillary and inguinal freckling, presenting with moyamoya syndrome (MMS) (Pabst et al., 2020). Although MMS has been largely reported in individuals with NF1, this recent association with Legius syndrome is relevant with regard to the phenotypic expansion of this rare condition. Legius syndrome has been long considered a milder clinical form of NF1, but it is a distinct autosomal dominant medical condition caused by pathogenic variants in SPRED1 (OMIM *609291). We report the second case of moyamoya syndrome in a patient with Legius syndrome caused by a novel frameshift variant in SPRED1. A 20yearold Caucasian woman was clinically diagnosed with NF1 during the first years of life, according to the presence of multiple caféaulait macules and axillary freckling. At the age of 4 years, she developed leftarm hemiparesis and motor weakness. Brain magnetic resonance imaging (MRI) showed ischemic infarction of the right cerebellar hemisphere and wholebody MRI revealed a mild aortic coarctation, without flow obstruction. At the age of 9 years, she presented with sudden generalized tonic– clonic seizures and brain MRI showed acute brainstem and left cerebellar hemisphere ischemia. One week later, an area of diffuse restriction in the left anterolateral portion of the ponsmidbrain junction was observed on brain magnetic resonance angiography (MRA) scans. No arterial narrowing could be noticed, but conventional angiography revealed tortuosity and irregularity of the distal branches of cerebellar arteries. One year later, MRA showed tortuosity and caliber irregularities of the superior cerebellar arteries, in combination with a moyamoya syndrome. Followup brain MRAs and perfusion studies were stable. However, the electroencephalogram (EEG) at the age of 19 years showed bilateral anterior and posterior irritative abnormalities induced by hyperventilation, suggestive of the typical “Rebuildup” phenomenon observed in moyamoya disease. A prophylactic therapy with acetylsalicylic acid (ASA) was therefore started. A targeted Next Generation Sequencing (NGS) panel including NF1 (OMIM *613113), SPRED1, and RNF213 (OMIM *613768) was performed within a retrospective study investigating the presence of RNF213 pathogenic variants in Caucasian subjects with NF1/Legius syndrome and moyamoya syndrome. After written informed consent was collected, DNA was extracted from blood samples of the proband and the parents using QIAamp DNA Mini Kit according to the manufacturer s instructions (Qiagen S.A.). DNA concentrations were quantified using the Nanodrop ND1000 UVVis spectrophotometer (Labtech France). We used the highthroughput NGS machine Ion Torrent PGM in combination with Ion AmpliSeq Designer Software v4.2 (Life Technologies). After the identification of the candidate variant, the proband s peripheral blood was collected in PAXgene Blood RNA tubes and subjected to total RNA extraction using the PAXgene Blood RNA kit (Qiagen) for cDNA analysis (further data available in the Supplementary Material). NGS analysis led to the identification of the novel paternally inherited null variant NM_001256071.3:c.1471+1dupG; p.(?) in RNF213 (NM_001256071.3, NP_001243000.2; https://www.ncbi.nlm.nih.gov/gene/57674) in our patient (Figure 1a) but, surprisingly, no pathogenic variant was observed in NF1. The patient carried instead the novel de novo NM_152594.3:c.330_331delGG; p.(Arg110Serfs*3) variant in SPRED1 (NM_152594.3, NP_689807.1, https://www.ncbi. nlm.nih.gov/gene/161742) (Figure 1b), leading to a diagnosis of Legius syndrome. Both the RNF213 and SPRED1 variants are rare (absent from GnomAD database) and predicted damage by in silico tools. The sequencing of RTPCR products generated by the RNF213 NM_001256071.3:c.1471+1dupG variant of the patient and wildtype allele showed the breakpoint of junction between the first 82 bps of exon 8 and the first bases of exon 9 causing by the intronic variant, supporting the pathogenic role of this variant (Figure 1c,d). To our knowledge, this is the second patient with moyamoya syndrome and Legius syndrome to be reported in the