Introduction: Williams-Beuren syndrome (WBS) is a rare genetic disorder characterized by congenital heart defects, dysmorphic features, intellectual delay, and a distinctive social behavioral profile. This highly recurrent and homogeneous phenotype has been curiously reported to be associated with autism spectrum disorders (ASD). Both genetic and environmental origins have been implicated. This study aimed to describe Tunisian patients associated with WBS and ASD and explore the underlying etiologies. Methods: Thirty-one clinically suspected WBS were referred for genetic exploration. A comprehensive evaluation using karyotyping, fluorescence in situ hybridization (FISH), and array comparative genomic hybridization (array-CGH) was performed. Results: All patients were clinically diagnosed and confirmed to have WBS through karyotyping and FISH analysis. Notably, six patients with complex or atypical clinical presentations underwent array-CGH. Two of these patients presented with ASD. Array CGH showed microdeletions ranging from 1.4 to 1.7 Mb in the 7q11.2 region. Further analysis of the extended region deletion identified a gene closely located in the deleted region, the HIP1 gene, involved in the central nervous system trafficking protein. Discussion: The recurrent deletion in WBS, as well as the mirror duplication, may contribute to ASD development in some cases, suggesting a potential involvement of the ASD genes pathway in this region. However, recessive genetic origins should also be considered, particularly in consanguineous families. Furthermore, our findings highlight the potential role of genetic factors and regulatory elements within the deleted region in modulating gene expression, notably the HIP1 gene. This underscores the implications of gene dosage and environmental factors in the broader WBS region, notably with language and social development. Conclusion: The presence of ASD in WBS patients emphasizes the need to investigate all WBS patients for autistic traits to establish a better genotype-phenotype correlation. We underline the utility of array-CGH as a valuable genetic diagnostic tool for characterizing WBS cases, and we shed light on the complex interplay of behavioral disorders in the 7q11.2 region rearrangements.

Background Forty-six ,XY Differences/Disorders of Sex Development (DSD) are characterized by a broad phenotypic spectrum ranging from typical female to male with undervirilized external genitalia, or more rarely testicular regres-sion with a typical male phenotype. Despite progress in the genetic diagnosis of DSD, most 46,XY DSD cases remain idiopathic.Methods To determine the genetic causes of 46,XY DSD, we studied 165 patients of Tunisian ancestry, who pre-sented a wide range of DSD phenotypes. Karyotyping, candidate gene sequencing, and whole-exome sequencing (WES) were performed.Results Cytogenetic abnormalities, including a high frequency of sex chromosomal anomalies (85.4%), explained the phenotype in 30.9% (51/165) of the cohort. Sanger sequencing of candidate genes identified a novel pathogenic vari-ant in the SRYgene in a patient with 46,XY gonadal dysgenesis. An exome screen of a sub-group of 44 patients with 46,XY DSD revealed pathogenic or likely pathogenic variants in 38.6% (17/44) of patients. Conclusion Rare or novel pathogenic variants were identified in the AR, SRD5A2, ZNRF3, SOX8, SOX9 and HHATgenes. Overall our data indicate a genetic diagnosis rate of 41.2% (68/165) in the group of 46,XY DSD.

Abstract Background Corpus callosum malformations (CCM) represent one of the most common congenital cerebral malformations with a prevalence of around one for 4000 births. There have been at least 230 reports in the literature concerning 1q43q44 deletions of varying sizes discovered using chromosomal microarrays. This disorder is distinguished by global developmental delay, seizures, hypotonia, corpus callosum defects, and significant craniofacial dysmorphism. In this study, we present a molecular cytogenetic analysis of 2 Tunisian patients with corpus callosum malformations. Patient 1 was a boy of 3 years old who presented psychomotor retardation, microcephaly, behavioral problems, interventricular septal defect, moderate pulmonary stenosis, hypospadias, and total CCA associated with delayed encephalic myelination. Patient 2 was a boy of 9 months. He presented a facial dysmorphia, a psychomotor retardation, an axial hypotonia, a quadri pyramidal syndrome, a micropenis, and HCC associated with decreased volume of the periventricular white matter. Both the array comparative genomic hybridization and fluorescence in situ hybridization techniques were used. Results Array CGH analysis reveals that patient 1 had the greater deletion size (11,7 Mb) at 1q43. The same region harbors a 2,7 Mb deletion in patient 2. Here, we notice that the larger the deletion, the more genes are likely to be involved, and the more severe the phenotype is likely to be. In both patients, the commonly deleted region includes six genes: PLD5, AKT3, ZNF238, HNRNPU, SDCCAG8 and CEP170. Based on the role of the ZNF238 gene in neuronal proliferation, migration, and cortex development, we hypothesized that the common deletion of ZNF238 in both patients seems to be the most responsible for corpus callosum malformations. Its absence may directly cause CCM. In addition, due to their high expression in the brain, PLD5 and FMN2 could modulate in the CCM phenotype. Conclusion Our findings support and improve the complex genotype–phenotype correlations previously reported in the 1qter microdeletion syndrome and define more precisely the neurodevelopmental phenotypes associated with genetic alterations of several genes related to this pathology.

Abstract Background Williams Beuren Syndrome is a multisystemic disorder manifested by congenital heart defects associated with dysmorphic features, intellectual delay, and a particular behavioural profile due to a microdeletion in 7q11.2. Methods To establish a genotype-phenotype correlation; we carried out a molecular cytogenetic analysis on 31 Tunisian WBS patients using the CGH-array and FISH techniques. Results 6 patients were investigated by CGH-array. All of them had a typical WBS deletion ranging from 1.4Mb to 1.7Mb. Curiously in 2 patients autistic spectrum disorders were noted in contrast to the behavioural profile generally observed in the other patients which are characterized by good contact. If we analyse the distal region of the generally deleted region, we found that the HIP1 gene is included. HIP1 encodes a central nervous system expressed protein and is considered the candidate gene for autism in this region. Conclusion Considering the presence of autism, a CGH analysis is essential to determine the exact etiology of this disorder, which seems strange for this syndrome, but is becoming progressively frequent. We suggest that alteration of the HIP1 gene could be indirectly responsible for autism, but specific environmental factors might act as risk factors triggering the development of this trait...

Introduction X-linked recessive ichthyosis (XLI) is a genodermatosis, caused by a deficiency of the steroid sulphatase enzyme encoded by the STS gene (OMIM # 300,747). Adopted XLI molecular diagnosis approaches differ from one laboratory to another depending on available technical facilities. Our work aims to figure out a sound diagnostic strategy for XLI. Patients and methods We collected 8 patients with XLI, all males, from 3 unrelated Tunisian families from central Tunisia. Genetic diagnosis was conducted through a large panel of genetic techniques including: Sanger Sequencing, haplotype analysis of STR markers, MLPA analysis, FISH and array CGH. Results Direct Sanger sequencing of the STS gene showed the same deletion of 13 base pairs within the exon 4 in all patients resulting in a premature stop codon. However, all patients' mothers were not carriers of this variant and no common haplotype flanking STS gene was shared between affected patients. Sequence alignment with reference human genome revealed an unprocessed pseudogene of the STS gene located on the Y chromosome, on which the 13 bp deletion was actually located. STS MLPA analysis identified a deletion of the entire STS gene on X chromosome for all affected patients. This deletion was confirmed by FISH and delineated by array CGH. Conclusion All our patients shared a deletion of the entire STS gene revealed by MLPA, confirmed by FISH and improved by array CGH. Geneticists must be aware of the presence of pseudogenes that can lead to XLI genetic misdiagnosis.

Background: Lissencephaly represents a rare subgroup of genetically distinct neurological disorders of neuronal migration characterized by a paucity or absence of cerebral gyration. The most common form of lissencephaly has been isolated and referred to as classic or type 1 lissencephaly. It is frequently related to abnormalities within LIS1 or DCX genes, with abnormalities ranging from single base pair substitutions to contiguous gene deletions. Methods: In this study, we report, for the first time, a clinical and genetic characterization of eight unrelated Tunisian children presenting type 1 lissencephaly. We screened LIS1 and DCX abnormalities thanks to a combination of molecular cytogenetic methods and next generation sequencing. Results: One deletion of DCX and three deletions of LIS1 were observed. One of these deletions was inherited from a maternal reciprocal translocation and estimated to approximately 2,9 Mb length. In addition, a 26 Kb LIS1 deletion was detected and refined between exon 3 up to exon 11 by target capture and sequencing. The last LIS1 rearrangement was a mutation (c.779T>A, p.V260E). Finally, two novel DCX mutations were found out (c.910G>C, p.G304R/c.436T>C, p.F146L). Conclusions: Our data confirm the individuality and originality of type 1 lissencephaly on both the phenotypic and the genetic levels. Furthermore, our data confirm again that LIS1 and DCX are the most genes associated with type 1 lissencephaly and spotlight the usefulness of developing approaches and methods for detecting a large number of known causative gene mutations.

Background: While Miller-Dieker syndrome critical region deletions are well known delineated anomalies, submicroscopic duplications in this region have recently emerged as a new distinctive syndrome. So far, only few cases have been described overlapping 17p13.3 duplications.Methods: In this study, we report on clinical and cytogenetic characterization of two new cases involving 17p13.3 and 3p26 chromosomal regions in two sisters with familial history of lissencephaly. Fluorescent In Situ Hybridization and array Comparative Genomic Hybridization were performed.Results: A deletion including the critical region of the Miller-Dieker syndrome of at least 2,9 Mb and a duplication of at least 3,6 Mb on the short arm of chromosome 3 were highlighted in one case. The opposite rearrangements, 17p13.3 duplication and 3p deletion were observed in the second case. This double chromosomal aberration is the result of an adjacent 1:1 meiotic segregation of a maternal reciprocal translocation t(3;17)(p26.2;p13.3).Conclusions: 17p13.3 and 3p26 deletions have a clear range of phenotypic features while duplications still have an uncertain clinical significance. However, we could suggest that regardless of the type of the rearrangement, the gene dosage and interactions of CNTN4, CNTN6 and CHL1 in the 3p26 and PAFAH1B1, YWHAE in 17p13.3 could result in different clinical spectrums.

Searchable abstracts of presentations at key conferences in endocrinology ISSN 1470-3947 (print) | ISSN 1479-6848 (online)

BackgroundSmall Supernumerary Marker Chromosomes (sSMC) are rare chromosomal abnormalities, which have abnormal banding arrangement and take many shapes. Several disorders have been correlated with sSMC presence. The aim of this study is to characterize the sSMC derived from chromosome 18 by Fluorescence in situ hybridization (FISH) and Array Comparative Genomic Hybridization (aCGH).ResultsNine children with dysmorphic features have been investigated. They have these features in common: a triangular face, low-set ears, a large mouth with a thin upper lip, and a horizontal palpebral fissure. Epicanthus and strabismus were present in two patients. In addition, we have noticed microcephaly and mental and/or developmental delay with low birth weight. However, two patients had standard birth weight; one patient had hypospadias; two had skin problems; and three showed different congenital heart defects. One patient had corpus callosum hypoplasia. Systematic karyotype analysis revealed a de novo supernumerary chromosome. Array CGH showed a gain in copy number on the short arm of chromosome 18 in the nine cases. In one case, the sSMC seemed to be in mosaic. The breakpoints of the marker were identified using aCGH and FISH. Thus, the sSMC led to 18p tetrasomy with approximately 14Mb lengths, between 364344 and 14763575 based on the human genome version 18.ConclusionsThese results have been completed by FISH in order to ascertain the shape of the sSMC. Our results confirm the uniqueness and particularity of the iso18p syndrome on the phenotypic as well as on the genetic level.

Objective: To determine the genetic cause of 46, XY primary amenorrhea in three 46, XY girls.Design: Whole exome sequencing. Setting: University cytogenetics center.Patient(s): Three patients with unexplained 46, XY primary amenorrhea were included in the study.Intervention(s): Potentially pathogenic variants were confirmed by Sanger sequencing, and familial segregation was determined where parents' DNA was available.Main Outcome Measure(s): Exome sequencing was performed in the three patients, and the data were analyzed for potentially pathogenic mutations. The functional consequences of mutations were predicted.Result(s): Three novel homozygous nonsense mutations in the luteinizing hormone receptor (LHCGR) gene were identified: c.1573 C -> T, p. Gln525Ter, c. 1435 C -> T p. Arg479Ter, and c. 508 C -> T, p. Gln170Ter.Conclusion(s): Inactivating mutations of the LHCGR gene may be a more common cause of 46, XY primary amenorrhea than previously considered. (C) 2016 by American Society for Reproductive Medicine.

In recent years, molecular genetics has been playing an increasing role in the diagnostic process of monogenic epilepsies. Knowing the genetic basis of one patient's epilepsy provides accurate genetic counseling and may guide therapeutic options. Genetic diagnosis of epilepsy syndromes has long been based on Sanger sequencing and search for large rearrangements using MLPA or DNA arrays (array-CGH or SNP-array). Recently, next-generation sequencing (NGS) was demonstrated to be a powerful approach to overcome the wide clinical and genetic heterogeneity of epileptic disorders. Coverage is critical for assessing the quality and accuracy of results from NGS. However, it is often a difficult parameter to display in practice. The aim of the study was to compare two library-building methods (Haloplex, Agilent and SeqCap EZ, Roche) for a targeted panel of 41 genes causing monogenic epileptic disorders. We included 24 patients, 20 of whom had known disease-causing mutations. For each patient both libraries were built in parallel and sequenced on an Ion Torrent Personal Genome Machine (PGM). To compare coverage and depth, we developed a simple homemade tool, named DeCovA (Depth and Coverage Analysis). DeCovA displays the sequencing depth of each base and the coverage of target genes for each genomic position. The fraction of each gene covered at different thresholds could be easily estimated. None of the two methods used, namely NextGene and Ion Reporter, were able to identify all the known mutations/CNVs displayed by the 20 patients. Variant detection rate was globally similar for the two techniques and DeCovA showed that failure to detect a mutation was mainly related to insufficient coverage.

Infantile Spasms syndrome (ISs) is a characterized by epileptic spasms occurring in clusters with an onset in the first year of life. West syndrome represents a subset of ISs that associates spasms in clusters, a hypsarrhythmia EEG pattern and a developmental arrest or regression. Aetiology of ISs is widely heterogeneous including many genetic causes. Many patients, however, remain without etiological diagnosis, which is critical for prognostic purpose and genetic counselling. In the present study, we performed genetic screening of 73 patients with different types of ISs by array-CGH and molecular analysis of 5 genes: CDKL5, STXBP1, KCNQ2, and GRIN2A, whose mutations cause different types of epileptic encephalopathies, including ISs, as well as MAGI2, which was suggested to be related to a subset of ISs. In total, we found a disease-causing mutation or CNV (Copy Number Variation) in 15% of the patients. These included 6 point mutations found in CDKL5 (n = 3) and STXBP1 (n = 3), 3 microdeletions (10 Mb in 2q24.3, 3.2 Mb in 5q14.3 including the region upstream to MEF2C, and 256 kb in 9q34 disrupting EHMT1), and 2 microduplications (671 kb in 2q24.3 encompassing SCN2A, and 11.93 Mb in Xq28). In addition, we discuss 3 CNVs as potential risk factors, including one 16p12.1 deletion, one intronic deletion of the NEDD4 gene, and one intronic deletion of CALN1 gene. The present findings highlight the efficacy of combined cytogenetic and targeted mutation screening to improve the diagnostic yield in patient with ISs.

We report on three patients presenting moderate intellectual disability, delayed language acquisition, and mild facial dysmorphia. Array-CGH studies revealed overlapping interstitial 12p13.1 microdeletions encompassing all or part of GRIN2B. GRIN2B encodes the NR2B subunit of the N-methyl-D-aspartate (NMDA) receptor. This receptor is a heteromeric glutamate-activated ion channel, present throughout the central nervous system. It plays a critical role in corticogenesis, neuronal migration, and synaptogenesis during brain development. GRIN2B alterations, including mutation and gene disruption by apparently balanced chromosomal rearrangements, have been described in patients with intellectual disability and autism spectrum disorder. We report here on the first cases of GRIN2B deletion, enlarging the spectrum of GRIN2B abnormalities. Our findings confirm the involvement of this gene in neurodevelopmental disorders. © 2013 Wiley Periodicals, Inc.

OBJECTIVES:Rolandic epilepsies (REs) represent the most frequent epilepsy in childhood. Patients may experience cognitive, speech, language, reading, and behavioral issues. The genetic origin of REs has long been debated. The participation of rare copy number variations (CNVs) in the pathophysiology of various human epilepsies has been increasingly recognized. However, no systematic search for microdeletions or microduplications has been reported in RE so far. METHODS:Array comparative genomic hybridization (aCGH) and quantitative polymerase chain reaction (qPCR) were used to analyze the genomic status of a series of 47 unrelated RE patients who displayed various types of electroclinical manifestations. RESULTS:Thirty rare CNVs were detected in 21 RE patients. Two CNVs were de novo, 12 were inherited, and 16 were of unknown inheritance. Each CNV was unique to one given patient, except for a 16p11.2 duplication found in two patients. The CNVs of highest interest comprised or disrupted strong candidate or confirmed genes for epileptic and other neurodevelopmental disorders, including BRWD3, GRIN2A, KCNC3, PRKCE, PRRT2, SHANK1, and TSPAN7. SIGNIFICANCE:Patients with REs showed rare microdeletions and microduplications with high frequency and heterogeneity. Whereas only a subset of all genomic alterations found here may actually participate in the phenotype, the novel de novo events as well as several inherited CNVs contain or disrupt genes, some of which are likely to influence the emergence, the presentation, or the comorbidity of RE. The future screening of cohorts of larger size will help in detecting more de novo or recurrent events and in appreciating the possible enrichment of specific CNVs in patients with RE.