Delineation of the KIAA2022 mutation phenotype: Two patients with X-linked intellectual disability and distinctive features

INTRODUCTION

X-linked ID (XLID) accounts for 5–10% of intellectual disability (ID) in males. Mutations causing XLID have been reported in over 100 genes [Tarpey et al., 2009; Lubs et al., 2012]. High-throughput sequencing has enabled the screening for a causative mutation in XLID using targeted panel, X-exome, and whole exome sequencing methods [de Ligt et al., 2012; Nava et al., 2012]. Cantagrel et al. [2004] previously identified an inactivating disruption of the X-linked KIAA2022 and P2RY8 in two male patients with severe ID [Cantagrel et al., 2004]. Van Maldergem et al. [2013] reported three additional families with moderate to severe ID with truncated KIAA2022 mutations and 70-kb microduplication encompassing KIAA2022 exon 1 [Van Maldergem et al., 2013]. Here, we report two unrelated male patients with nonsyndromic ID and novel KIAA2022 nonsense mutations detected by targeted sequencing.

CLINICAL REPORTS

We selected 13 Japanese male patients, including four sets of siblings (three male siblings and one female sibling) and nine sporadic patients with severe ID and slightly dysmorphic features inconsistent with established syndromes, after exclusion of chromosomal anomalies by array comparative genomic hybridization (CGH). Two of thirteen patients had KIAA2022 mutation.

Patient 1

Patient 1 was a 3-year-old boy. He was the first and only child of unrelated healthy parents with no family history of ID. He was delivered at 33 weeks gestation owing to premature rupture of membrane after an uneventful pregnancy. His birth weight was 1762 g [− 0.5 standard deviation (SD)], length was 40.5 cm (− 1.3 SD), and OFC was 30.0 cm (0.4 SD). He was admitted to the neonatal intensive care unit and was supplied oxygen because of respiratory distress. Hypotonia was noted at birth, associated with poor feeding and poor weight gain. After discharge without tube feeding, he had failure-ro-thrive due to gastroesophageal reflux with persistent vomiting more than twice daily. The patient started controlling his head at 6 months, rolling over at 9 months, and sitting with support at 21 months. He was referred at 1 year and 9 months for assessment of hypotonia and developmental delay. His height was 73.6 cm (− 3.3 SD), weight 9.8 kg (− 1.2 SD), and OFC45.6 cm (− 1.7 SD). He had postnatal growth retardation (Suppl. Fig. S1A,B). He was able to sit alone at age three years; however, he did not speak comprehensively. He had autistic behavior with repetitive hand flapping movement and tongue protrusion. The patient showed dysmorphic features, including upswept hair, narrow forehead, downslanting eyebrows, long nose, hypoplastic alae nasi, thin lips, open mouth, and large ears (Fig. 1A). Brain magnetic resonance imaging (MRI) revealed normal structure for his age.

MATERIALS AND METHODS

Written informed consent was obtained from the patients' parents in accordance with the Kanagawa Children's Medical Center Review Board and Ethics Committee.

Total genomic DNA was obtained from lymphocytes using standard techniques. Targeted panel was designed for targeted next-generation sequencing for 68 XLID genes and 101 autistic spectrum disorder (ASDs) genes, covering all coding regions and untranslated regions (in total, 1,164,259 bp). Candidate genes were selected from a literature review of XLID and ASDs genes associated with nonsyndromic ID [Vissers et al., 2010; Najmabadi et al., 2011; Lubs et al., 2012]. The list of genes is described in the supplemental data (Suppl. Table SI). DNA libraries were enriched for sequences using HaloPlex (Agilent Technologies Inc., Santa Clara, CA). Patient samples were sequenced by MiSeq (IlluminaInc., San Diego, CA) by 150-bp pair-end reads. Data were analyzed by Burrows–Wheeler alignment tool version 6 and Genome Analysis Toolkit pipeline (Broad Institute, Cambridge, MA) and visualized in the Integrative Genomics Viewer (IGV). Calling copy-number variation (CNV) was based on log–ratio analysis and the z-score of read depth on each exon. Mutations identified by targeted sequencing were confirmed by Sanger sequencing and demonstrated appropriate segregation with phenotype in unaffected mothers of each XLID case.

Array CGH was performed using the Agilent SurePrint G3 Human CGH Microarray Kit 8 × 60k (Agilent Technologies) according to manufacturer's instructions. The results were analyzed using Agilent Genomic Workbench software (Agilent Technologies).

RESULTS

The mean depth of coverage over all samples was 281× per base, and bases covered by at least five reads were 96.7% and those covered by at least 20 reads were 90.1% of coding regions. Targeted sequencing identified two novel mutations of KIAA2022 in two unrelated patients. Patient 1 had c.2113C>T/p.Q705X (NM_001008537) in the coding region of exon 3, and his mother did not have the mutation. Patient 2 had c.964C>T/p.R322X (NM_001008537) in the coding region of exon 3, and his mother did not have the mutation (Suppl. Fig. S2). Neither mutation was present in the human genetic variation database (Japanese genetic variation consortium, a reference database of genetic variations in Japanese population: 1,208 individuals http://www.genome.med.kyoto-u.ac.jp/SnpDB), the 1000 genomes project, and NHLBI grant opportunity exome sequencing project (ESP).

Array CGH revealed no significant copy number variants in either patient. In Patient 1, array CGH revealed an 886-kb duplication at 4q28.3 from position 133,225,227 to 134,111,432. The duplicated region included only a part of PCDH10. This CNV was derived from his healthy mother.

DISCUSSION

De novo KIAA2022 nonsense mutations were identified in two unrelated patients with severe ID. This report confirms that KIAA2022 mutations represent a clinically recognizable condition characterized by ID and distinctive dysmorphic features. KIAA2022 mutations and alterations have been reported in only four families with nonsyndromic ID and epilepsy [Cantagrel et al., 2004; Van Maldergem et al., 2013] (Table I). Moderate to severe ID was described in two families with frameshift mutations, one family with intron 1 disruption of KIAA2022, and the present patients with nonsense mutations. Mild ID was reported in one family with intragenic microduplication of KIAA2022, which is considered a hypomorphic phenotype. There is a correlation between the genotype and severity of ID. The consistent features among these patients were hypotonic face, narrow forehead, upturned nostrils, open mouth, and large ears. The patient with microduplication showed mild features. In this study, two of thirteen patients had KIAA2022 mutations. Significant high prevalence with these mutations is associated with selection of patients according to phenotypes and common features with KIAA2022 mutation or a chance occurrence. It is difficult to estimate the prevalence of KIAA2022 related ID.

KIAA2022 mutation may cause central hypothyroidism and nephrotic syndrome in addition to as ID. KIAA2022 is highly expressed in the fetal and adult brain and plays a role in neuronal development. KIAA2022 knockdown reduced the total neurite length in an in vitro study [Van Maldergem et al., 2013]. In the mouse brain, KIAA2022 expression occurred ubiquitously during the embryonic and perinatal stages and was maintained in the hippocampus, dentate gyrus, olfactory bulb, ventral thalamic nucleus, and ventromedial hypothalamic nucleus during the adult stage [Ishikawa et al., 2012]. KIAA2022 mutation is suspected to be associated with hypothalamic dysfunction, leading to central hypothyroidism in Patient 2. Xpn (mouse ortholog of KIAA2022) knockdown enhanced cell–cell and cell–matrix adhesion mediated by N-cadherin and beta 1 integrin [Magome et al., 2013]. The beta 1 integrin showed characteristic expression pattern during nephrogenesis and is crucial for normal development and function of the renal glomeruli [Kanasaki et al., 2008]. Overexpression of beta 1 integrin contributes to mesangial remodeling and is related to glomeluronephritis [Kuhara et al., 1997]. Alterations in beta 1 integrin expression pattern in location and intensity have been found in congenital nephrotic syndrome of the Finnish type [Ljungberg et al., 1996]. The occurrence of nephrotic syndrome in patient 2 suggested the possibility that dysfunction of cell adhesions could affect glomerular function due to KIAA2022 mutation. KIAA2022 is expressed in lower amounts in organ tissues other than brain, and its function in the kidney remains unclear. Other cases with KIAA2022 mutation revealed no disease or anomaly other than ID and epilepsy; there is a possibility of coincidental occurrence in Patient 2.

These additional patients support the evidence that KIAA2022 is a causative gene for X-linked nonsyndromic ID. The two patients showed consistent distinctive features and severe ID.