NSD1 mutations in Sotos syndrome

Sotos syndrome is a genetic disorder characterized by a typical facial appearance, macrocephaly, accelerated growth, developmental delay, and a variable range of associated abnormalities. The NSD1 gene was recently found to be responsible for Sotos syndrome, and more than 150 patients with NSD1 alterations have been identified. A significant ethnic difference is found in the prevalence of different types of mutation, with a high percentage of microdeletions identified in Japanese Sotos syndrome patients and with intragenic mutations in most non-Japanese patients. NSD1 aberrations are rather specific for Sotos syndrome, but have also been detected in patients lacking one or more major criteria of the disorder, namely overgrowth, macrocephaly, and advanced bone age. Thus, new diagnostic criteria should be considered. Studies have reported different frequencies of mutations versus non-mutations in Sotos syndrome, thus indicating allelic or locus hetereogeneity. Although some authors have suggested genotype/phenotype correlations, further studies are needed.     

Missense and nonsense mutations in the alternatively-spliced exon 2 of COL2A1 cause the ocular variant of Stickler syndrome

Stickler syndrome type I (STL1) is a phenotypically heterogeneous disorder characterized by ocular and extraocular features. It is caused by null-allele mutations in the COL2A1 gene that codes for procollagen II. COL2A1 precursor mRNA undergoes alternative splicing, resulting in two isoforms, a long form including exon 2 (type IIA isoform) and a short form excluding exon 2 (type IIB isoform). The short form is predominantly expressed by differentiated chondrocytes in adult cartilage, and the long form in chondroprogenitor cells during early development and in the vitreous of the eye, which is the only adult tissue containing procollagen IIA. Recent evidence indicates that due to the tissue-specific expression of these two isoforms, premature termination codon mutations in exon 2 cause Stickler syndrome with minimal or no extraocular manifestations. We describe here two mutations in exon 2 of COL2A1 in three patients with predominantly ocular Stickler syndrome: Cys64Stop in two patients, and a novel structural mutation, Cys57Tyr, in one patient. RT-PCR of total lymphoblast RNA from one patient with the Cys64Stop mutation revealed that only the normal allele of the IIA form was present, indicating that the mutation resulted either in complete loss of the allele by nonsense-mediated mRNA decay or by skipping of exon 2 via nonsense-mediated altered splicing, resulting in production of the type IIB isoform. The results of COL2A1 minigene expression studies suggest that both Cys64Stop and Cys57Tyr alter positive cis regulatory elements for splicing, resulting in a lower IIA:IIB ratio.     

Heterogeneity of NSD1 alterations in 116 patients with Sotos syndrome

Sotos syndrome is an overgrowth syndrome characterized by distinctive facial features, learning difficulties, and macrocephaly with frequent pre- and postnatal overgrowth with advanced bone age. Here, we report on our experience in the molecular diagnostic of Sotos syndrome on 116 patients. Using direct sequencing and a quantitative multiplex PCR of short fluorescent fragments (QMPSF)-based assay allowing accurate detection of both total and partial NSD1 deletions, we identified NSD1 abnormalities in 104 patients corresponding to 102 Sotos families (90%). NSD1 point mutations were detected in 80% of the index cases, large deletions removing the NSD1 gene entirely in 14%, and intragenic NSD1 rearrangements in 6%. Among the 69 detected distinct point mutations, 48 were novel. The QMPSF assay detected an exonic duplication and a mosaic partial deletion. QMPSF mapping of the 15 large deletions revealed the heterogeneity of the deletions, which vary in size from 1 to 4.5 Mb. Clinical features of NSD1-positive Sotos patients revealed that the phenotype in patients with nontruncating mutations was less severe that in patients with truncating mutations. This study confirms the heterogeneity of NSD1 alterations in Sotos syndrome and therefore the need to complete sequencing analysis by screening for partial deletions and duplications to ensure an accurate molecular diagnosis of this syndrome.     

Ganglioglioma in a Sotos syndrome patient with an NSD1 deletion

Sotos syndrome, a disorder with macrocephaly, mental delay, and facial anomalies, has been noted to have an increased risk of neoplasia. Here, we report a patient with a microdeletion in nuclear receptor SET-domain-containing protein (NSD1) and a previously undescribed intracranial ganglioglioma.     

Analysis of the NSD1 promoter region in patients with a Sotos syndrome phenotype

Sotos syndrome (SoS, OMIM#117550) is an overgrowth disorder characterized by excessive growth—especially in the first years of childhood—distinctive craniofacial features, and various degrees of mental retardation. Haploinsufficiency of the nuclear receptor binding SET domain containing protein 1 (NSD1) gene, due to either intragenic mutations or whole-gene microdeletions, is found in the majority of patients with SoS. However, in approximately 10–40% of patients with a typical SoS phenotype, no abnormalities are detected. In this study, hemizygous hypermethylation or genomic sequence abnormalities of the promoter region of NSD1 were hypothesized to be the underlying cause in patients with a SoS phenotype, but without confirmed NSD1 alterations. In 18 patients, including one patient with a reported hepatocellular carcinoma, the promoter region of NSD1 was analyzed. However, no hypermethylation or sequence abnormalities in the promoter region could be detected. It therefore seems unlikely that such abnormalities of NSD1 are a major culprit in patients with phenotypical SoS. Additional methods are necessary for detection of other genetic or epigenetic causes of SoS.     

Partial NSD1 deletions cause 5% of Sotos syndrome and are readily identifiable by multiplex ligation dependent probe amplification

Background: Most cases of Sotos syndrome are caused by intragenic NSD1 mutations or 5q35 microdeletions. It is uncertain whether allelic or genetic heterogeneity underlies the residual cases and it has been proposed that other mechanisms, such as 11p15 defects, might be responsible for Sotos cases without NSD1 mutations or 5q35 microdeletions. Objective: To develop a multiplex ligation dependent probe amplification (MLPA) assay to screen NSD1 for exonic deletions/duplications. Methods: Analysis was undertaken of 18 classic Sotos syndrome cases in which NSD1 mutations and 5q35 microdeletions were excluded. Long range polymerase chain reaction (PCR) was used to characterise the mechanism of generation of the partial NSD1 deletions. Results: Eight unique partial NSD1 deletions were identified: exons 1–2 (n = 4), exons 3–5, exons 9–13, exons 19–21, and exon 22. Using long range PCR six of the deletions were confirmed and the precise breakpoints in five cases characterised. This showed that three had arisen through Alu-Alu recombination and two from non-homologous end joining. Conclusions: MLPA is a robust, inexpensive, simple technique that reliably detects both 5q35 microdeletions and partial NSD1 deletions that together account for ∼15% of Sotos syndrome.     

Mutations in NSD1 are responsible for Sotos syndrome, but are not a frequent finding in other overgrowth phenotypes

Recently, deletions encompassing the nuclear receptor binding SET-Domain 1 (NSD1) gene have been described as the major cause of Japanese patients with the Sotos syndrome, whereas point mutations have been identified in the majority of European Sotos syndrome patients. In order to investigate a possible phenotype-genotype correlation and to further define the predictive value of NSD1 mutations, we performed mutational analysis of the NSD1 gene in 20 patients and one familial case with Sotos syndrome, five patients with Weaver syndrome, six patients with unclassified overgrowth/mental retardation, and six patients with macrocephaly/mental retardation. We were able to identify mutations within the NSD1 gene in 18 patients and the familial case with Sotos syndrome (90%). The mutations (six nonsense, eight frame shifts, three splice site, one missense, one in-frame deletion) are expected to result in an impairment of NSD1 function. The best correlation between clinical assessment and molecular results was obtained for the Sotos facial gestalt in conjunction with overgrowth, macrocephaly, and developmental delay. In contrast to the high mutation detection rate in Sotos syndrome, none of the patients with Weaver syndrome, unclassified overgrowth/mental retardation and macrocephaly/mental retardation, harbored NSD1 mutations. We tested for large deletions by FISH analysis but were not able to identify any deletion cases. The results indicate that the great majority of patients with Sotos syndrome are caused by mutations in NSD1. Deletions covering the NSD1 locus were not found in the patients analyzed here.     

NSD1 analysis for Sotos syndrome: insights and perspectives from the clinical laboratory.

PURPOSE: Sotos syndrome is a genetic disorder characterized primarily by overgrowth, developmental delay, and a characteristic facial gestalt. Defects in the NSD1 gene are present in approximately 80% of patients with Sotos syndrome. The goal of this study was to determine the incidence of NSD1 abnormalities in patients referred to a clinical laboratory for testing and to identify clinical criteria that distinguish between patients with and without NSD1 abnormalities. METHODS: Deletion or mutation analysis of the NSD1 gene was performed on 435 patients referred to our clinical genetics laboratory. Detailed clinical information was obtained on 86 patients with and without NSD1 abnormalities, and a clinical checklist was developed to help distinguish between these two groups of patients. RESULTS: Abnormalities of the NSD1 gene were identified in 55 patients, including 9 deletions and 46 mutations. Thus, in the clinical laboratory setting, deletions were found in 2% and mutations in 21% of samples analyzed, because not all patients had both tests. Thirty-three previously unreported mutations in the NSD1 gene were identified. Clinical features typically associated with Sotos syndrome were not found to be significantly different between individuals with and without NSD1 abnormalities. The clinical checklist developed included poor feeding, increased body mass index, and enlarged cerebral ventricles, in addition to the typical clinical features of Sotos syndrome, and was able to distinguish between the two groups with 80% sensitivity and 70% specificity. CONCLUSIONS: The dramatic decrease in the frequency of finding NSD1 abnormalities in the clinical laboratory is likely because of the heterogeneity of the patient population. Our experience from a diagnostic laboratory can help guide clinicians in deciding for whom NSD1 genetic analysis is indicated.     

Spectrum of NSD1 mutations in Sotos and Weaver syndromes

Interestingly, mental retardation was consistently more severe in patients with NSD1 deletions. Macrocephaly and facial gestalt but not overgrowth and advanced bone age were consistently observed in Sotos syndrome patients. We suggest therefore considering macrocephaly and facial gestalt as mandatory criteria for the diagnosis of Sotos syndrome and overgrowth and advanced bone age as minor criteria.     

A novel homozygous nonsense NDNF variant in Kallmann syndrome

Kallmann syndrome (KS) is a rare genetic disease characterized by pubertal failure and olfactory defects. Although many genes associated with KS have been reported, most are rare. Recently, heterozygous inactivating mutations in the neuron-derived neurotrophic factor gene (NDNF) were reported to cause KS. Here, we present a 14-year-old Kurdish boy with KS who has a novel homozygous nonsense c.1251C>A (p.Tyr417Ter) variant in NDNF. The variant was not observed in reference population databases and was predicted to be deleterious. Segregation analysis performed with Sanger sequencing indicated the autosomal recessive inheritance of the clinical phenotype. His heterozygous parents have experienced timely pubertal development and normal reproductive features. This study reported the first homozygous truncating NDNF variant, enabling the direct observation of the clinical consequences of predictively absent NDNF function. These results support the contention that the inactivating mutations in NDNF cause KS, and provide additional evidence for the complex inheritance of KS.     

Craniofacial and oral features of Sotos syndrome: differences in patients with submicroscopic deletion and mutation of NSD1 gene

Sotos syndrome is a well-known overgrowth syndrome caused by haploinsufficiency of NSD1 gene located at 5q35. There are two types of mutations that cause NSD1 haploinsufficiency: mutations within the NSD1 gene (mutation type) and a 5q35 submicroscopic deletion encompassing the entire NSD1 gene (deletion type). We investigated detailed craniofacial, dental, and oral findings in five patients with deletion type, and three patients with mutation type Sotos syndrome. All eight patients had a high palate, excessive tooth wear, crowding, and all but one patient had hypodontia and deep bite. Hypodontia was exclusively observed in the second premolars, and there were no differences between the deletion and mutation types in the number of missing teeth. Another feature frequently seen in common with both types was maxillary recession. Findings seen more frequently and more pronounced in deletion-type than in mutation-type included mandibular recession, scissors or posterior cross bite, and small dental arch with labioclination of the maxillary central incisors. It is noteworthy that although either scissors bite or cross bite was present in all of the deletion-type patients, neither of these was observed in mutation-type patients. Other features seen in a few patients include enamel hypoplasia (two deletion patients), and ectopic tooth eruption (one deletion and one mutation patients). Our study suggests that Sotos syndrome patients should be observed closely for possible dental and oral complications especially for malocculusion in the deletion-type patients.     

Adults with Sotos syndrome: review of 21 adults with molecularly confirmed NSD1 alterations, including a detailed case report of the oldest person

Sotos syndrome is a well‐described multiple anomaly syndrome characterized by overgrowth, distinctive craniofacial appearance, and variable learning disabilities. The diagnosis of Sotos syndrome relied solely on these clinical criteria until haploinsufficiency of the NSD1 gene was identified as causative. We describe a 63‐year‐old woman with classic features and a pathogenic NSD1 mutation, who we believe to be the oldest reported person with Sotos syndrome. She is notable for the diagnosis of Sotos syndrome late in life, mild cognitive limitation, and chronic kidney disease attributed to fibromuscular dysplasia for which she recently received a transplant. She has basal cell and squamous cell carcinoma for which her lifetime of sun exposure and fair cutaneous phototype are viewed as risk factors. We also reviewed previous literature reports (n = 11) for adults with Sotos syndrome, and studied patients ascertained in the Spanish Overgrowth Syndrome Registry (n = 15). Analysis was limited to 21/27 (78%) total patients who had molecular confirmation of Sotos syndrome (15 with a mutation, 6 with a microdeletion). With a mean age of 26 years, the most common features were learning disabilities (90%), scoliosis (52%), eye problems (43%), psychiatric issues (30%), and brain imaging anomalies (28%). Learning disabilities were more severe in patients with a microdeletion than in those with a point mutation. From this small study with heterogeneous ascertainment we suggest modest adjustments to the general healthcare monitoring of individuals with Sotos syndrome. Although this series includes neoplasia in four cases, this should not be interpreted as incidence. Age‐appropriate cancer surveillance should be maintained. © 2011 Wiley‐Liss, Inc.