NSDHL-containing duplication at Xq28 in a male patient with autism spectrum disorder: a case report

Autism spectrum disorder (ASD) is a neurodevelopmental disorder that is defined in DSM-5 [1] as persistent deficits in social communication and social interaction across multiple contexts in conjunction with restricted, repetitive patterns, interests, or activities as manifested by at least two prototypically inflexible behaviours [2]. A high degree of heritability and modest environmental influences contribute to the aetiology of ASD [3, 4]. The genetic architecture of autism has proven to be complex and heterogeneous through linkage or association based on whole-genome or exome sequencing [5, 6]. In recent decades, 10–20% of ASD cases have been attributed to ASD susceptibility genes [6]. In the X chromosome, double expression of these related genes leads to functional disomy [7‐9]. Among the pathogenic X-linked duplications, the most prominent and well-studied is that of the Methyl-CpG-binding Protein 2 gene (MECP2; OMIM: 300005), located at Xq28, causing severe X-linked intellectual disability (XLID) [10], and its loss of function causes Rett syndrome (OMIM: 613454) [11, 12]. Duplication of chromosome 15q11–13, which includes a series of imprinting and non-imprinting genes, results in the recurrent cytogenetic abnormalities associated with ASD and represents one of the most frequently reported CNVs in ASD [13].

Duplication involving the int22h-1/int22h-2 LCR-flanked region in distal Xq28 was recently linked to intellectual and developmental disabilities [14, 15]. NAD(P)H steroid dehydrogenase-like protein (NSDHL), also located at Xq28, is expressed in developing cortical neurons and glia and encodes an enzyme in post-squalene cholesterol biosynthesis [16]. Mutations of NSDHL have been related to neurodevelopmental diseases including CHILD syndrome (OMIM: 308050) and CK syndrome (OMIM: 300831). In CHILD syndrome, NSDHL function is presumably eliminated or greatly reduced [17]; CK syndrome affects only males [16]. CK syndrome caused by NSDHL mutations is characterized by mild to severe cognitive impairment, seizures beginning in infancy, microcephaly, cerebral cortical malformations and a thin body habitus [18]. Most male patients with CK syndrome manifest behaviours of aggression, attention deficit hyperactivity disorder (ADHD), and irritability. Using Autism Diagnostic Interview-Revised (ADI-R) [19] or the Autism Diagnostic Observation Schedule (ADOS) [20], the affected males fulfil partial criteria for ASD [18]. However, little is known about the clinical consequences of NSDHL duplication, and no NSDHL whole-gene duplication with ASD has been reported in the literature.

Here, we report an 8-year-old boy diagnosed with ASD and possessing a 260-kb NSDHL-containing duplication at Xq28 inherited from his mother; this constitutes the first report of an ASD patient with a related NSDHL-containing duplication at Xq28. The patient underwent careful clinical observation, detailed examination, and comprehensive medical history recording and demonstrated symptoms of social deficiency, restricted interests, attention deficit and impulsive behaviour but normal cognition.

The patient in our study manifested core symptoms of ASD with accompanying ADHD and impulsiveness. CNV scanning revealed that the patient had a Xq28 duplication encompassing 8 genes, and that the most disease-associating gene NSDHL was among this duplication region; this is the first report of an ASD patient with a related NSDHL-containing duplication at Xq28.

We hypothesized that the NSDHL-containing duplication at Xq28 was responsible for the ASD. Four MAGE family genes [24] have been reported to be expressed at high levels in numerous tumours of various histological types, whereas in a large panel of healthy tissues, expression was observed only in the testis and placenta. CSAG family genes [25], which are expressed at quite low levels in brain, are mostly expressed in the testis, and CSAG1 is expressed in some higher-grade chondrosarcomas, chondrosarcoma-derived cell lines, and melanoma cell lines, as well as in some types of cartilage. CETN2 [26] is associated with the centrosome, whereas ZNF185 [27] is mostly expressed in the human adult prostate, testis, ovary, placenta, peripheral blood, and embryonic kidney; a partial copy was contained in this region (encompassing a minimum of 16 and a maximum of 18 exons). Since the levels of CETN2 and ZNF185 are also high in the brain, we searched more databases, including OMIM, Gene Cards and ClinVar, but still did not find any report indicating associations among CETN2, ZNF185 and neural development disorders. Diseases associated with CETN2 include Xeroderma pigmentosum [28], while diseases associated with NSDHL include CK Syndrome and CHILD Syndrome. Thus, most genes in the duplicated region are expressed beyond the brain or exhibit little relation to autism or neurodevelopmental disease (See Additional file 1: Table S3–1, S3–2). NSDHL, which is located at Xq28, is expressed in developing cortical neurons and glia and encodes an enzyme that functions in post-squalene cholesterol biosynthesis [16], thus affecting the steps of the cholesterol biosynthetic pathway. Research has shown that perturbations of cholesterol metabolism are associated with a variety of CNS disorders found in ASD [29] and other mental disorders [30]. Nevertheless, we still need more evidence to evaluate the effects of other genes in the duplicated region and more functional studies to show the importance of NSDHL to the development of ASD.

NSDHL mutations, which have been implicated in CK syndrome [18] in males, result in rare X-linked intellectual disabilities and cognitive impairment. Affected males also have behavioural problems, including aggression, ADHD, irritability, and seizures [18] and partially fulfil the criteria for ASD [18]. Our patient exhibited the characteristics of attention deficit and irritability but not seizure and satisfied the criteria for ASD diagnosis. The patient had an NSDHL duplication at Xq28, which was inherited from his mother. NSDHL is likely the most important disease-related gene within the duplicated region of this patient because the other genes in this duplicated region have little relevance to ASD. In contrast to patients with CK syndrome [16], our patient exhibited normal cognition, possibly because the NSDHL gene was duplicated and not mutated. His mother, who carried the NSDHL duplication on only one X chromosome, exhibited a few symptoms of ASD and had normal physical features. Her intellect and irritable disposition were consistent with previous reports of female carriers of NSDHL mutations [16]. Studies have demonstrated that female carriers of an NSDHL mutation generally do not manifest signs or symptoms of CK syndrome [31]. These carriers have normal physical features, intellect, and brain imaging findings but exhibit behavioural problems, such as irritability and aggression.

We also checked the common variation in DGV (http://​dgv.​tcag.​ca/​dgv/​app/​home) for the same CNV duplication region. We found that Jakobsson’s [32] study showed an observed gain including the entire NSDHL gene. Tracing the duplication region, we found out that a female had the duplication by checking the information of the raw data (https://​www.​ncbi.​nlm.​nih.​gov/​geo/​query/​acc.​cgi?​acc=​GSM264524; sample ID HGDP00959). We assumed that female carriers with NSDHL duplication exhibit few symptoms and tend to be normal. To our knowledge, the frequency of the CNV in the normal population remains uncertain.

Patients with similar duplications reported in the DECIPHER database share some phenotypes and exhibit intellectual disabilities, muscular hypotonia and obesity. Table 1 shows an overview of similar duplications in DECIPHER. We found one boy (No. 258546) who had inherited from his unaffected mother a similar duplication region as that in our patient also had Autism Spectrum Disorder (We contacted the clinician for more information on these patients via DECIPHER. However, some patients did not have any further follow-up. A negative finding for ASD diagnosis in DECIPHER does not necessarily indicate that the cases do not have ASD; therefore, the importance of NSDHL cannot be ignored). Furthermore, because NSDHL functions in the removal of two C-4 methyl groups in one of the later steps of cholesterol biosynthesis, it is more likely that patients with similar duplications will exhibit obesity, including our patient, who showed higher serum levels of free fatty acids. These data support the dysfunction of NSDHL in cholesterol biosynthesis and the development of CNS.

Here, we have identified an 8-year-old patient carrying a 260-kb NSDHL-containing duplication inherited from his mother. Since the data in DGV and DECIPHER are not considered part of an peer-reviewed publication, this is the first report of an ASD patient with a related NSDHL-containing duplication at Xq28. The patient fulfilled the criteria for ASD and shared some characteristics with CK syndrome patients [16, 18, 33] but exhibited normal cognition. Further studies and case reports are required to establish that both mutations and duplications of NSDHL cause neurodevelopmental diseases. The relationship between NSDHL and ASD also requires further study to obtain conclusive support.