Delineation of the adult phenotype of Coffin-Siris syndrome in 35 individuals.

Coffin-Siris syndrome (CSS) is a rare multisystemic autosomal dominant disorder. Since 2012, alterations in genes of the SWI/SNF complex were identified as the molecular basis of CSS, studying largely pediatric cohorts. Therefore, there is a lack of information on the phenotype in adulthood, particularly on the clinical outcome in adulthood and associated risks. In an international collaborative effort, data from 35 individuals ≥ 18 years with a molecularly ascertained CSS diagnosis (variants in ARID1B, ARID2, SMARCA4, SMARCB1, SMARCC2, SMARCE1, SOX11, BICRA) using a comprehensive questionnaire was collected. Our results indicate that overweight and obesity are frequent in adults with CSS. Visual impairment, scoliosis, and behavioral anomalies are more prevalent than in published pediatric or mixed cohorts. Cognitive outcomes range from profound intellectual disability (ID) to low normal IQ, with most individuals having moderate ID. The present study describes the first exclusively adult cohort of CSS individuals. We were able to delineate some features of CSS that develop over time and have therefore been underrepresented in previously reported largely pediatric cohorts, and provide recommendations for follow-up.

A PAK1 Mutational Hotspot Within the Regulatory CRIPaK Domain is Associated With Severe Neurodevelopmental Disorders in Children.

BACKGROUND: P-21-activated kinases (PAKs) are protein serine/threonine kinases, part of the RAS/mitogen-activated protein kinase pathway. PAK1 is highly expressed in the central nervous system and crucially involved in neuronal migration and brain developmental processes. Recently, de novo heterozygous missense variants in PAK1 have been identified as an ultrarare cause of pediatric neurodevelopmental disorders. METHODS: We report a series of children affected with postnatal macrocephaly, neurodevelopmental impairment, and drug-resistant epilepsy. Repeated electroencephalographic (EEG) and video-EEG evaluations were performed over a two- to 10-year period during follow-up to delineate electroclinical histories. Genetic sequencing studies and computational evaluation of the identified variants were performed in our patient cohort. RESULTS: We identified by whole-exome sequencing three novel de novo variants in PAK1 (NM_001128620: c.427A>G, p.Met143Val; c.428T>C, p.Met143Thr; c.428T>A, p.Met143Lys) as the underlying cause of the disease in our families. The three variants affected the same highly conserved Met143 residue within the cysteine-rich inhibitor of PAK1 (CRIPaK) domain, which was identified before as a PAK1 inhibitor target. Computational studies suggested a defective autoinhibition presumably due to impaired PAK1 autoregulation as a result of the recurrent substitution. CONCLUSIONS: We delineated the electroclinical phenotypes of PAK1-related neurological disorders and highlight a novel mutational hotspot that may involve defective autoinhibition of the PAK1 protein. The three novel variants affecting the same hotspot residue within the CRIPaK domain highlight potentially impaired PAK1-CRIPaK interaction as a novel disease mechanism. These findings shed light on possible future treatments targeted at the CRIPaK domain, to modulate PAK1 activity and function.

Clustered variants in the 5' coding region of TRA2B cause a distinctive neurodevelopmental syndrome.

PURPOSE: Transformer2 proteins (Tra2α and Tra2ß) control splicing patterns in human cells, and no human phenotypes have been associated with germline variants in these genes. The aim of this work was to associate germline variants in the TRA2B gene to a novel neurodevelopmental disorder. METHODS: A total of 12 individuals from 11 unrelated families who harbored predicted loss-of-function monoallelic variants, mostly de novo, were recruited. RNA sequencing and western blot analyses of Tra2ß-1 and Tra2ß-3 isoforms from patient-derived cells were performed. Tra2ß1-GFP, Tra2ß3-GFP and CHEK1 exon 3 plasmids were transfected into HEK-293 cells. RESULTS: All variants clustered in the 5' part of TRA2B, upstream of an alternative translation start site responsible for the expression of the noncanonical Tra2ß-3 isoform. All affected individuals presented intellectual disability and/or developmental delay, frequently associated with infantile spasms, microcephaly, brain anomalies, autism spectrum disorder, feeding difficulties, and short stature. Experimental studies showed that these variants decreased the expression of the canonical Tra2ß-1 isoform, whereas they increased the expression of the Tra2ß-3 isoform, which is shorter and lacks the N-terminal RS1 domain. Increased expression of Tra2ß-3-GFP were shown to interfere with the incorporation of CHEK1 exon 3 into its mature transcript, normally incorporated by Tra2ß-1. CONCLUSION: Predicted loss-of-function variants clustered in the 5' portion of TRA2B cause a new neurodevelopmental syndrome through an apparently dominant negative disease mechanism involving the use of an alternative translation start site and the overexpression of a shorter, repressive Tra2ß protein.

Behavioral and psychological features in girls and women with triple-X syndrome.

Triple-X syndrome is a common sex chromosome aneuploidy, which appears in 1 out of 1,000 females. The aim of our study was to describe the behavioral features of a large group of girls and women with triple-X in comparison to a control group. A total of 72 subjects with triple-X and 69 subjects of an age-matched control group were included. Psychological and behavioral questionnaires were allocated to three age groups, representing a range of ages from young childhood to adulthood. Regarding the females between 4 and 7 years of age, we found significant differences for social problems, attention problems, and school performance. For the age group 8-17 years, we found larger significant differences for the majority of the scales listed in the child behavior checklist. The most significant differences (p < .001) were from total behavior problems, internalizing problems, and four other scales. Young females with triple-X have significantly lower general self-esteem, especially concerning school and family. In the adults, there were significant differences concerning psychological symptoms and distress, with higher scores in the triple-X subjects. Regardless, their mean scores were still in the normal range. We did not find clinical evidence for more than 50% of the triple-X females in any age group, indicating that approximately half of them do not have behavioral problems, and that more than 60% do not differ in their competence from the control group. However, our findings suggest that triple-X influences mental health and the overall well-being of the individuals across their whole life spans.

Maternal variants in NLRP and other maternal effect proteins are associated with multilocus imprinting disturbance in offspring.

BACKGROUND: Genomic imprinting results from the resistance of germline epigenetic marks to reprogramming in the early embryo for a small number of mammalian genes. Genetic, epigenetic or environmental insults that prevent imprints from evading reprogramming may result in imprinting disorders, which impact growth, development, behaviour and metabolism. We aimed to identify genetic defects causing imprinting disorders by whole-exome sequencing in families with one or more members affected by multilocus imprinting disturbance. METHODS: Whole-exome sequencing was performed in 38 pedigrees where probands had multilocus imprinting disturbance, in five of whom maternal variants in NLRP5 have previously been found. RESULTS: We now report 15 further pedigrees in which offspring had disturbance of imprinting, while their mothers had rare, predicted-deleterious variants in maternal effect genes, including NLRP2, NLRP7 and PADI6. As well as clinical features of well-recognised imprinting disorders, some offspring had additional features including developmental delay, behavioural problems and discordant monozygotic twinning, while some mothers had reproductive problems including pregnancy loss. CONCLUSION: The identification of 20 putative maternal effect variants in 38 families affected by multilocus imprinting disorders adds to the evidence that maternal genetic factors affect oocyte fitness and thus offspring development. Testing for maternal-effect genetic variants should be considered in families affected by atypical imprinting disorders.

Noncoding copy-number variations are associated with congenital limb malformation.

PurposeCopy-number variants (CNVs) are generally interpreted by linking the effects of gene dosage with phenotypes. The clinical interpretation of noncoding CNVs remains challenging. We investigated the percentage of disease-associated CNVs in patients with congenital limb malformations that affect noncoding cis-regulatory sequences versus genes sensitive to gene dosage effects.MethodsWe applied high-resolution copy-number analysis to 340 unrelated individuals with isolated limb malformation. To investigate novel candidate CNVs, we re-engineered human CNVs in mice using clustered regularly interspaced short palindromic repeats (CRISPR)-based genome editing.ResultsOf the individuals studied, 10% harbored CNVs segregating with the phenotype in the affected families. We identified 31 CNVs previously associated with congenital limb malformations and four novel candidate CNVs. Most of the disease-associated CNVs (57%) affected the noncoding cis-regulatory genome, while only 43% included a known disease gene and were likely to result from gene dosage effects. In transgenic mice harboring four novel candidate CNVs, we observed altered gene expression in all cases, indicating that the CNVs had a regulatory effect either by changing the enhancer dosage or altering the topological associating domain architecture of the genome.ConclusionOur findings suggest that CNVs affecting noncoding regulatory elements are a major cause of congenital limb malformations.

The maternal uniparental disomy of chromosome 6 (upd(6)mat) "phenotype": result of placental trisomy 6 mosaicism?

BACKGROUND: Maternal uniparental disomy of chromosome 6 (upd(6)mat) is a rare finding and its clinical relevance is currently unclear. Based on clinical data from two new cases and patients from the literature, the pathogenetic significance of upd(6)mat is delineated. METHODS: Own cases were molecularly characterized for isodisomic uniparental regions on chromosome 6. For further cases with upd(6)mat, a literature search was conducted and genetic and clinical data were ascertained. RESULTS: Comparison of isodisomic regions between the new upd(6)mat cases and those from four reports did not reveal any common isodisomic region. Among the patients with available cytogenetic data, five had a normal karyotype in lymphocytes, whereas a trisomy 6 (mosaicism) was detected prenatally in four cases. A common clinical picture was not obvious in upd(6)mat, but intrauterine growth restriction (IUGR) and preterm delivery were frequent. CONCLUSION: A common upd(6)mat phenotype is not obvious, but placental dysfunction due to trisomy 6 mosaicism probably contributes to IUGR and preterm delivery. In fact, other clinical features observed in upd(6)mat patients might be caused by homozygosity of recessive mutations or by an undetected trisomy 6 cell line. Upd(6)mat itself is not associated with clinical features, and can rather be regarded as a biomarker. In case upd(6)mat is detected, the cause for the phenotype is identified indirectly, but the UPD is not the basic cause.

Tubulin-related cerebellar dysplasia: definition of a distinct pattern of cerebellar malformation.

OBJECTIVE: To determine the neuroimaging pattern of cerebellar dysplasia (CD) and other posterior fossa morphological anomalies associated with mutations in tubulin genes and to perform clinical and genetic correlations. METHODS: Twenty-eight patients harbouring 23 heterozygous pathogenic variants (ten novel) in tubulin genes TUBA1A (n = 10), TUBB2B (n = 8) or TUBB3 (n = 5) were studied by a brain MRI scan performed either on a 1.5 T (n = 10) or 3 T (n = 18) MR scanner with focus on the posterior fossa. RESULTS: Cerebellar anomalies were detected in 24/28 patients (86%). CD was recognised in 19/28 (68%) including cortical cerebellar dysplasia (CCD) in 18/28, either involving only the cerebellar hemispheres (12/28) or associated with vermis dysplasia (6/28). CCD was located only in the right hemisphere in 13/18 (72%), including four TUBB2B-, four TUBB3- and five TUBA1A-mutated patients, while in the other five TUBA1A cases it was located only in the left hemisphere or in both hemispheres. The postero-superior region of the cerebellar hemispheres was most frequently affected. CONCLUSIONS: The cerebellar involvement in tubulinopathies shows specific features that may be labelled as 'tubulin-related CD'. This pattern is unique and differs from other genetic causes of cerebellar dysplasia. KEY POINTS: • Cortical cerebellar dysplasia without cysts is suggestive of tubulin-related disorder. • Cerebellar dysplasia in tubulinopathies shows specific features labelled as 'tubulin-related CD'. • Focal and unilateral involvement of cerebellar hemispheres has important implications for counselling.

Dravet syndrome: a new causative SCN1A mutation?

Dravet syndrome is often caused by SCN1A mutations and has a wide variation in clinical appearance. Indication for genetic analysis should be an epileptic encephalopathy or severe clinical course of seizures in infants with episodes of fever before the first year of life.

47 patients with FLNA associated periventricular nodular heterotopia.

BACKGROUND: Heterozygous loss of function mutations within the Filamin A gene in Xq28 are the most frequent cause of bilateral neuronal periventricular nodular heterotopia (PVNH). Most affected females are reported to initially present with difficult to treat seizures at variable age of onset. Psychomotor development and cognition may be normal or mildly to moderately impaired. Distinct associated extracerebral findings have been observed and may help to establish the diagnosis including patent ductus arteriosus Botalli, progressive dystrophic cardiac valve disease and aortic dissection, chronic obstructive lung disease or chronic constipation. Genotype-phenotype correlations could not yet be established. METHODS: Sanger sequencing and MLPA was performed for a large cohort of 47 patients with Filamin A associated PVNH (age range 1 to 65 years). For 34 patients more detailed clinical information was available from a structured questionnaire and medical charts on family history, development, epileptologic findings, neurological examination, cognition and associated clinical findings. Available detailed cerebral MR imaging was assessed for 20 patients. RESULTS: Thirty-nine different FLNA mutations were observed, they are mainly truncating (37/39) and distributed throughout the entire coding region. No obvious correlation between the number and extend of PVNH and the severity of the individual clinical manifestation was observed. 10 of the mutation carriers so far are without seizures at a median age of 19.7 years. 22 of 24 patients with available educational data were able to attend regular school and obtain professional education according to age. CONCLUSIONS: We report the clinical and mutation spectrum as well as MR imaging for a large cohort of 47 patients with Filamin A associated PVNH including two adult males. Our data are reassuring in regard to psychomotor and cognitive development, which is within normal range for the majority of patients. However, a concerning median diagnostic latency of 17 to 20 years was noted between seizure onset and the genetic diagnosis, intensely delaying appropriate medical surveillance for potentially life threatening cardiovascular complications as well as genetic risk assessment and counseling prior to family planning for this X-linked dominant inherited disorder with high perinatal lethality in hemizygous males.

Congenital CLN disease in two siblings.

BACKGROUND: Neuronal ceroid lipofuscinoses (NCL) is characterized by a combination of retinopathy, dementia, and epilepsy. As a group, they encompass ten distinct biological and clinical entities and are the most common type of childhood neurodegenerative disease. PATIENTS AND METHODS: Case reports. RESULTS: We demonstrate the clinical course of two neonates (brother and sister) with infantile neuronal ceroid lipofuscinoses (NCL) (CLN 10 disease) presenting with intractable seizures and respiratory insufficiency immediately after birth. Characteristic clinical, radiological and pathological findings of this form of NCL are presented. CONCLUSIONS: We conclude that the diagnosis of CLN10 should be kept in mind as a differential diagnosis in newborns presenting with respiratory insufficiency and severe epilepsy that is largely refractory to anti-epileptic drugs (AED) treatment. Because of the severity of CLN10 disease and futility of treatment, important ethical issues arise when caring for children with this clinical entity.

Next-generation sequencing in X-linked intellectual disability.

X-linked intellectual disability (XLID) is a genetically heterogeneous disorder with more than 100 genes known to date. Most genes are responsible for a small proportion of patients only, which has hitherto hampered the systematic screening of large patient cohorts. We performed targeted enrichment and next-generation sequencing of 107 XLID genes in a cohort of 150 male patients. Hundred patients had sporadic intellectual disability, and 50 patients had a family history suggestive of XLID. We also analysed a sporadic female patient with severe ID and epilepsy because she had strongly skewed X-inactivation. Target enrichment and high parallel sequencing allowed a diagnostic coverage of >10 reads for ~96% of all coding bases of the XLID genes at a mean coverage of 124 reads. We found 18 pathogenic variants in 13 XLID genes (AP1S2, ATRX, CUL4B, DLG3, IQSEC2, KDM5C, MED12, OPHN1, SLC9A6, SMC1A, UBE2A, UPF3B and ZDHHC9) among the 150 male patients. Thirteen pathogenic variants were present in the group of 50 familial patients (26%), and 5 pathogenic variants among the 100 sporadic patients (5%). Systematic gene dosage analysis for low coverage exons detected one pathogenic hemizygous deletion. An IQSEC2 nonsense variant was detected in the female ID patient, providing further evidence for a role of this gene in encephalopathy in females. Skewed X-inactivation was more frequently observed in mothers with pathogenic variants compared with those without known X-linked defects. The mutation rate in the cohort of sporadic patients corroborates previous estimates of 5-10% for X-chromosomal defects in male ID patients.

Deletions of exons with regulatory activity at the DYNC1I1 locus are associated with split-hand/split-foot malformation: array CGH screening of 134 unrelated families.

BACKGROUND: A growing number of non-coding regulatory mutations are being identified in congenital disease. Very recently also some exons of protein coding genes have been identified to act as tissue specific enhancer elements and were therefore termed exonic enhancers or "eExons". METHODS: We screened a cohort of 134 unrelated families with split-hand/split-foot malformation (SHFM) with high resolution array CGH for CNVs with regulatory potential. RESULTS: In three families with an autosomal dominant non-syndromic SHFM phenotype we detected microdeletions encompassing the exonic enhancer (eExons) 15 and 17 of DYNC1I1. In a fourth family, who had hearing loss in addition to SHFM, we found a larger deletion of 510 kb including the eExons of DYNC1I1 and, in addition, the human brain enhancer hs1642. Exons 15 and 17 of DYNC1I1 are known to act as tissue specific limb enhancers of DLX5/6, two genes that have been shown to be associated with SHFM in mice. In our cohort of 134 unrelated families with SHFM, deletions of the eExons of DYNC1I1 account for approximately 3% of the cases, while 17p13.3 duplications were identified in 13% of the families, 10q24 duplications in 12%, and TP63 mutations were detected in 4%. CONCLUSIONS: We reduce the minimal critical region for SHFM1 to 78 kb. Hearing loss, however, appears to be associated with deletions of a more telomeric region encompassing the brain enhancer element hs1642. Thus, SHFM1 as well as hearing loss at the same locus are caused by deletion of regulatory elements. Deletions of the exons with regulatory potential of DYNC1I1 are an example of the emerging role of exonic enhancer elements and their implications in congenital malformation syndromes.

Deletions in 14q24.1q24.3 are associated with congenital heart defects, brachydactyly, and mild intellectual disability.

Interstitial deletions of chromosome band 14q24.1q24.3 are apparently very rare. We report on three unrelated patients with overlapping de novo deletions of sizes 5.4, 2.8, and 2.3 Mb in this region. While some clinical problems such as intestinal malrotation, cryptorchidism, and ectopic kidney were only observed in single patients, all three patients had mild intellectual disability, congenital heart defects (truncus arteriosus, pulmonary atresia, atrial septal defect, and/or ventricular septal defect), brachydactyly, hypertelorism, broad nasal bridge, and thin upper lips. Likely haploinsufficiency of one or several of the 19 genes in the common deleted interval (ACTN1, DCAF5, EXD2, GALNTL1, ERH, SLC39A9, PLEKHD1, CCDC177, KIAA0247, LOC100289511, SRSF5, SLC10A1, SMOC1, SLC8A3, ADAM21P1, COX16, SYNJ2BP, SYNJ2BP-COX16, ADAM21) was responsible for these manifestations, but apart from SMOC1, mutations in which cause autosomal recessive Waardenburg anophthalmia syndrome, and ACTN1, mutations in which are associated with congenital macrothrombocytopenia, no disease associations have so far been reported for the other genes. Functional studies and a systematic search for mutations or chromosome aberrations in this region will elucidate the role of individual genes in the clinical manifestations and will provide insight into the underlying biological mechanisms.

Molecular karyotyping as a relevant diagnostic tool in children with growth retardation with Silver-Russell features.

OBJECTIVE: To determine the contribution of submicroscopic chromosomal imbalances to the etiology of Silver-Russell syndrome (SRS) and SRS-like phenotypes. STUDY DESIGN: We performed molecular karyotyping in 41 patients with SRS or SRS-like features without known chromosome 7 and 11 defects using the Affymetrix SNP Array 6.0 system (Affymetrix, High Wycombe, United Kingdom). RESULTS: In 8 patients, pathogenic copy number variations with sizes ranging from 672 kb to 9.158 Mb were identified. The deletions in 1q21, 15q26, 17p13, and 22q11 were associated with known microdeletion syndromes with overlapping features with SRS. The duplications in 22q13 and Xq25q27 represent unique novel copy number variations but have an obvious influence on the phenotype. In 5 additional patients, the pathogenetic relevance of the detected variants remained unclear. CONCLUSION: Pathogenic submicroscopic imbalances were detectable in a significant proportion of patients with short stature and features reminiscent of SRS. Therefore, molecular karyotyping should be implemented in routine diagnostics for growth-retarded patients with even slight dysmorphisms suggestive for SRS.

Phenotypic spectrum associated with CASK loss-of-function mutations.

BACKGROUND: Heterozygous mutations in the CASK gene in Xp11.4 have been shown to be associated with a distinct brain malformation phenotype in females, including disproportionate pontine and cerebellar hypoplasia. METHODS: The study characterised the CASK alteration in 20 new female patients by molecular karyotyping, fluorescence in situ hybridisation, sequencing, reverse transcriptase (RT) and/or quantitative real-time PCR. Clinical and brain imaging data of a total of 25 patients were reviewed. RESULTS: 11 submicroscopic copy number alterations, including nine deletions of ~11 kb to 4.5 Mb and two duplications, all covering (part of) CASK, four splice, four nonsense, and one 1 bp deletion are reported. These heterozygous CASK mutations most likely lead to a null allele. Brain imaging consistently showed diffuse brainstem and cerebellar hypoplasia with a dilated fourth ventricle, but of remarkably varying degrees. Analysis of 20 patients in this study, and five previously reported patients, revealed a core clinical phenotype comprising severe developmental delay/intellectual disability, severe postnatal microcephaly, often associated with growth retardation, (axial) hypotonia with or without hypertonia of extremities, optic nerve hypoplasia, and/or other eye abnormalities. A recognisable facial phenotype emerged, including prominent and broad nasal bridge and tip, small or short nose, long philtrum, small chin, and/or large ears. CONCLUSIONS: These findings define the phenotypic spectrum associated with CASK loss-of-function mutations. The combination of developmental and brain imaging features together with mild facial dysmorphism is highly suggestive of this disorder and should prompt subsequent testing of the CASK gene.

Christianson syndrome in a patient with an interstitial Xq26.3 deletion.

Interstitial deletions of chromosome band Xq26.3 are rare. We report on a 2-year-old boy in whom array comparative genomic hybridization analysis revealed an interstitial 314 kb deletion in Xq26.3 affecting SLC9A6 and FHL1. Mutations in SLC9A6 are associated with Christianson syndrome (OMIM 300243), a syndromic form of X-linked mental retardation (XLMR) characterized by microcephaly, severe global developmental delay, ataxia and seizures. FHL1 mutations cause Emery-Dreifuss muscular dystrophy (OMIM 310300), X-linked myopathy with postural muscle atrophy (XMPMA, OMIM 300696), scapuloperoneal myopathy (OMIM 300695), or reducing body myopathy (OMIM 300717, 300718). The clinical problems of the patient reported here comprised severe intellectual disability, absent speech, ataxia, epilepsy, and gastroesophageal reflux, and could mostly be attributed to SLC9A6 insufficiency. In contrast to the majority of reported Christianson syndrome patients who were microcephalic, this patient was normocephalic, but his head circumference had decelerated from the 50th centile at birth to the 25th centile at the age of 2 ²/¹² years. Muscle problems due to the FHL1 deletion are not to be expected before late childhood, which is the earliest age of onset for FHL1 associated Emery-Dreifuss muscular dystrophy. This patient broadens the spectrum of SLC9A6 mutations and contributes to the clinical delineation of Christianson syndrome. This is also the first patient with a deletion affecting both SLC9A6 and the complete FHL1 gene.