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In this report we describe a male patient with a rare de novo interstitial deletion of chromosome 2q14.1-q22.1. His karyotype was reported as 46,XY,del(2)(q13q21) but subsequent array comparative genomic hybridization (array CGH) analysis redefined the deletion breakpoints as 2q14.1 and 2q22.1. Eight patients have been reported with deletions either within or spanning the region 2q13 or 2q14 to 2q22.1. In five patients the diagnosis was made by karyotype analysis alone and in three reported patients and the proband array CGH analysis was also performed. When the proband was compared with the eight previously reported patients it was apparent that they shared many clinical findings suggesting that patients with a de novo interstitial deletion involving 2q13 or 2q14 to 2q21 or 2q22 may have a recognizable phenotype. There are 14 known disease-associated genes in the deleted region of 2q14.1-q22.1 and their possible phenotypic effects on the proband and the eight previously reported patients are discussed., (© 2014 Wiley Periodicals, Inc.)

Our aim was to construct a streamlined technical workflow to facilitate a prospective, multi-centre evaluation of array comparative genomic hybridisation (array-CGH) in the prenatal diagnostic context. A collection of commercially available DNA extraction and quantification techniques were evaluated and compared using minimal quantities of amniotic fluid, chorionic villi and cultured cells. When prenatal DNA of suitable quality and quantity was obtained, array-CGH was performed using Oxford Gene Technology's (OGT, Oxford, UK) CytoSure™ ISCA 8 × 60 K oligo array platform. With starting quantities of 2-4 ml amniotic fluid, 2-5 mg chorionic villi or under 150,000 cultured cells the following optimised technical workflow was identified: DNA extraction using the iGENatal™ kit (igenbiotech, Madrid, Spain) and quantification by the Qubit® 2.0 Fluorometer with the Qubit® dsDNA BR assay kit (Invitrogen™, Eugene, OR, USA). In addition, it was elucidated that array-CGH can be successfully performed with as little as 125 ng DNA in the experiment using the OGT CytoSure™ ISCA 8 × 60 K oligo array platform. Amidst an on-going debate on whether array-CGH should be applied in the prenatal diagnostic setting, by following the technical recommendations described here genetics laboratories can now gain exposure to prenatal array-CGH testing without compromising the conventional karyotype result.

The clinical utility of microarray technologies when used in the context of prenatal diagnosis lies in the technology's ability to detect submicroscopic copy number changes that are associated with clinically significant outcomes. We have carried out a systematic review of the literature to calculate the utility of prenatal microarrays in the presence of a normal conventional karyotype. Amongst 12,362 cases in studies that recruited cases from all prenatal ascertainment groups, 295/12,362 (2.4%) overall were reported to have copy number changes with associated clinical significance (pCNC), 201/3090 (6.5%) when ascertained with an abnormal ultrasound, 50/5108 (1.0%) when ascertained because of increased maternal age and 44/4164 (1.1%) for all other ascertainment groups (e.g. parental anxiety and abnormal serum screening result). When additional prenatal microarray studies are included in which ascertainment was restricted to fetuses with abnormal ultrasound scans, 262/3730 (7.0%) were reported to have pCNCs., (© 2013 John Wiley & Sons, Ltd.)

Background: In cultured, dividing transformed T lymphocytes and in dividing bone marrow cells from normal men and those with a haematological malignancy, sex chromosome aneuploidy has been found to increase in prevalence and degree with age. This has rarely been investigated in non-dividing uncultured blood samples. The loss and gain of the X chromosome in dividing transformed lymphocytes in women with age is much more frequent than that of the Y chromosome in males. However, paradoxically X chromosome aneuploidy is rarely seen in the dividing cells of bone marrow of females., Methods: In blood samples from 565 men with breast cancer and 54 control men from the England and Wales general population, 80 cell nuclei per sample were scored for presence of X and Y chromosomes using fluorescent centromeric probes., Results: Sex chromosome aneuploidy, largely Y chromosome loss, was present in 63% of cases and 57% of controls, with the prevalence and degree of aneuploidy increasingly sharply and highly significantly with age. At ages 65-80 years, 71% of cases and 85% of controls showed aneuploidy and 15% and 25%, respectively, had ≥ 10% of cells aneuploid. Allowing for age, aneuploidy was less prevalent (P=0.03) in cases than controls., Conclusion: Sex chromosome aneuploidy in non-dividing nuclei of peripheral blood cells is frequent in adult men, the prevalence and degree increasing sharply with age. The possible relation of sex chromosome aneuploidy to breast cancer risk in men, and to cancer risk generally, needs further investigation, ideally in cohort studies.

The 8p23.1 duplication syndrome is a relatively rare genomic condition that has been confirmed with molecular cytogenetic methods in only 11 probands and five family members. Here, we describe another prenatal and five postnatal patients with de novo 8p23.1 duplications analyzed with oligonucleotide array comparative genomic hybridization (oaCGH). Of the common features, mild or moderate developmental delays and/or learning difficulties have been found in 11/12 postnatal probands, a variable degree of mild dysmorphism in 8/12 and congenital heart disease (CHD) in 4/5 prenatal and 3/12 postnatal probands. Behavioral problems, cleft lip and/or palate, macrocephaly, and seizures were confirmed as additional features among the new patients, and novel features included neonatal respiratory distress, attention deficit hyperactivity disorder (ADHD), ocular anomalies, balance problems, hypotonia, and hydrocele. The core duplication of 3.68 Mb contains 31 genes and microRNAs of which only GATA4, TNKS, SOX7, and XKR6 are likely to be dosage sensitive genes and MIR124-1 and MIR598 have been implicated in neurocognitive phenotypes. A combination of the duplication of GATA4, SOX7, and related genes may account for the variable penetrance of CHD. Two of the duplications were maternal and intrachromosomal in origin with maternal heterozygosity for the common inversion between the repeats in 8p23.1. These additional patients and the absence of the 8p23.1 duplications in published controls, indicate that the 8p23.1 duplication syndrome may now be considered a pathogenic copy number variation (pCNV) with an estimated population prevalence of 1 in 58,000., (Copyright © 2013 Wiley Periodicals, Inc.)

We report a case of Albright hereditary osteodystrophy (AHO) in a three-year-old girl with a microduplication at 17q11.2. The child developed obesity within the first 6 months of life. A diagnosis of Albright was made at age 2 years when biochemical evidence of parathyroid resistance was found. No mutations were identified in guanine nucleotide-binding protein G (s) subunit alpha (GNAS1). Subsequent investigations revealed methylation disturbance at GNAS1A, neuroendocrine secretory protein antisense (NESPAS) and neuroendocrine secretory protein 55 (NESP55) confirming a diagnosis of pseudohypothyroidism type 1B. A deletion of NESP55 and uniparental disomy chromosome 20 were excluded which suggested that the features of AHO arose through a purely epigenetic mechanism. Further investigation revealed a de novo microduplication at 17q11.2 encompassing the neurofibromatosis type 1 (NF1) gene. The combination of two rare de novo events in the same child raises the possibility that duplication of a gene within the 17q11.2 region may have triggered abnormal methylation in the GNAS cluster region on chromosome 20.

Duplications leading to functional disomy of chromosome Xq28, including MECP2 as the critical dosage-sensitive gene, are associated with a distinct clinical phenotype in males, characterized by severe mental retardation, infantile hypotonia, progressive neurologic impairment, recurrent infections, bladder dysfunction, and absent speech. Female patients with Xq duplications including MECP2 are rare. Only recently submicroscopic duplications of this region on Xq28 have been recognized in four females, and a triplication in a fifth, all in combination with random X-chromosome inactivation (XCI). Based on this small series, it was concluded that in females with MECP2 duplication and random XCI, the typical symptoms of affected boys are not present. We present clinical and molecular data on a series of five females with an Xq28 duplication including the MECP2 gene, both isolated and as the result of a translocation, and compare them with the previously reported cases of small duplications in females. The collected data indicate that the associated phenotype in females is distinct from males with similar duplications, but the clinical effects may be as severe as seen in males., (Copyright © 2012 Elsevier Masson SAS. All rights reserved.)

We report a large series of 173 patients with physical and/or neurological abnormalities and a de novo imbalance identified by array CGH. Breakpoint intervals were screened for the presence of low copy repeats (LCRs) to distinguish between rearrangements formed by non-allelic homologous recombination (NAHR) and rearrangements formed by other mechanisms. We identified significant differences in size and parental origin between the LCR-mediated and non-LCR groups. Non-LCR imbalances were evenly distributed among the four size intervals we defined, whereas LCR-mediated rearrangements had a narrow size distribution, predominantly between 1 and 5 Mb (P = 0.001). Among the LCR-mediated rearrangements there were equal numbers of maternally and paternally derived cases. In contrast, for the non-LCR rearrangements there was a significant excess of paternal cases (P = 0.024) over a wide size range including below 1 Mb. Our results provide novel evidence that unbalanced chromosome rearrangements are not only more frequent in males, but may also arise through different mechanisms than those seen in females. Although the paternal imbalances identified in our study are evenly distributed throughout the four size groups, there are very few maternal imbalances either <1 Mb or >10 Mb. Furthermore, a lower proportion of paternal imbalances are LCR mediated (13/71) compared with the maternal imbalances (12/30). We hypothesise that imbalances of maternal origin arise predominantly through NAHR during meiosis, while the majority of imbalances of paternal origin arise through male-specific mechanisms other than NAHR. Our data suggest that mitotic mechanisms could be important for the formation of chromosome imbalances; however, we found no association with increased paternal age.

In several laboratories, genome-wide array analysis has been implemented as the first tier diagnostic test for the identification of copy number changes in patients with mental retardation and/or congenital anomalies. The identification of a pathogenic copy number variant (CNV) is not only important to make a proper diagnosis but also to enable the accurate estimation of the recurrence risk to family members. Upon the identification of a de novo interstitial loss or gain, the risk recurrence is considered very low. However, this risk is 50% if one of the parents is carrier of a balanced insertional translocation (IT). The apparently de novo imbalance in a patient is then the consequence of the unbalanced transmission of a derivative chromosome involved in an IT. To determine the frequency with which insertional balanced translocations would be the origin of submicroscopic imbalances, we investigated the potential presence of an IT in a consecutive series of 477 interstitial CNVs, in which the parental origin has been tested by FISH, among 14,293 patients with developmental abnormalities referred for array. We demonstrate that ITs underlie ~2.1% of the apparently de novo, interstitial CNVs, indicating that submicroscopic ITs are at least sixfold more frequent than cytogenetically visible ITs. This risk estimate should be taken into account during counseling, and warrant parental and proband FISH testing wherever possible in patients with an apparently de novo, interstitial aberration.

Background: Analysis of cell free fetal (cff) DNA in maternal plasma is used routinely for non invasive prenatal diagnosis (NIPD) of fetal sex determination, fetal rhesus D status and some single gene disorders. True positive results rely on detection of the fetal target being analysed. No amplification of the target may be interpreted either as a true negative result or a false negative result due to the absence or very low levels of cffDNA. The hypermethylated RASSF1A promoter has been reported as a universal fetal marker to confirm the presence of cffDNA. Using methylation-sensitive restriction enzymes hypomethylated maternal sequences are digested leaving hypermethylated fetal sequences detectable. Complete digestion of maternal sequences is required to eliminate false positive results., Methods: cfDNA was extracted from maternal plasma (n = 90) and digested with methylation-sensitive and insensitive restriction enzymes. Analysis of RASSF1A, SRY and DYS14 was performed by real-time PCR., Results: Hypermethylated RASSF1A was amplified for 79 samples (88%) indicating the presence of cffDNA. SRY real time PCR results and fetal sex at delivery were 100% accurate. Eleven samples (12%) had no detectable hypermethylated RASSF1A and 10 of these (91%) had gestational ages less than 7 weeks 2 days. Six of these samples were male at delivery, five had inconclusive results for SRY analysis and one sample had no amplifiable SRY., Conclusion: Use of this assay for the detection of hypermethylated RASSF1A as a universal fetal marker has the potential to improve the diagnostic reliability of NIPD for fetal sex determination and single gene disorders.

Purpose: Copy number variants have emerged as a major cause of human disease such as autism and intellectual disabilities. Because copy number variants are common in normal individuals, determining the functional and clinical significance of rare copy number variants in patients remains challenging. The adoption of whole-genome chromosomal microarray analysis as a first-tier diagnostic test for individuals with unexplained developmental disabilities provides a unique opportunity to obtain large copy number variant datasets generated through routine patient care., Methods: A consortium of diagnostic laboratories was established (the International Standards for Cytogenomic Arrays consortium) to share copy number variant and phenotypic data in a central, public database. We present the largest copy number variant case-control study to date comprising 15,749 International Standards for Cytogenomic Arrays cases and 10,118 published controls, focusing our initial analysis on recurrent deletions and duplications involving 14 copy number variant regions., Results: Compared with controls, 14 deletions and seven duplications were significantly overrepresented in cases, providing a clinical diagnosis as pathogenic., Conclusion: Given the rapid expansion of clinical chromosomal microarray analysis testing, very large datasets will be available to determine the functional significance of increasingly rare copy number variants. This data will provide an evidence-based guide to clinicians across many disciplines involved in the diagnosis, management, and care of these patients and their families.

The availability of microarray technology has led to the recent recognition of copy number abnormalities of distal chromosome 22q11.2 that are distinct from the better-characterized deletions and duplications of the proximal region. This report describes five unrelated individuals with copy number abnormalities affecting distal chromosome 22q11.2. We report on novel phenotypic features including diaphragmatic hernia and uterine didelphys associated with the distal microdeletion syndrome; and frontomedial polymicrogyria and callosal agenesis associated with the distal microduplication syndrome. We describe the third distal chromosome 22q11.2 microdeletion patient with Goldenhar syndrome. Patients with distal chromosome 22q11.2 copy number abnormalities exhibit inter- and intra-familial phenotypic variability, and challenge our ability to draw meaningful genotype-phenotype correlations., (Copyright © 2011 Wiley-Liss, Inc.)

We report six patients with array deletions encompassing 12q14. Out of a total of 2538 array investigations carried out on children with developmental delay and dysmorphism in three diagnostic testing centres, six positive cases yielded a frequency of 1 in 423 for this deletion syndrome. The deleted region in each of the six cases overlaps significantly with previously reported cases with microdeletions of this region. The chromosomal range of the deletions extends from 12q13.3q15. In the current study, we report overlapping deletions of variable extent and size but primarily comprising chromosomal bands 12q13.3q14.1. Four of the six deletions were confirmed as de novo events. Two cases had deletions that included HMGA2, and both children had significant short stature. Neither case had osteopoikilosis despite both being deleted for LEMD3. Four cases had deletions that ended proximal to HMGA2 and all of these had much better growth. Five cases had congenital heart defects, including two with atrial septal defects, one each with pulmonary stenosis, sub-aortic stenosis and a patent ductus. Four cases had moderate delay, two had severe developmental delay and a further two had a diagnosis of autism. All six cases had significant speech delay with subtle facial dysmorphism.

We describe a patient with striking generalized symmetrical enchondromatosis of the tubular bones and a de novo duplication of chromosome 12p11.23 to 12p11.22. The PTHLH gene within this region encodes a ligand for PTHR1: mutations in the gene encoding this receptor are associated with some cases of Ollier disease, several skeletal dysplasias including Blomstrand, Eiken, and Jansen and down-regulation of PTHLH expression in brachydactyly type E. Our findings suggest that abnormal PTHLH-PTHR1 signaling may underly this unusual form of enchondromatosis and indicate that unlike most cases of Ollier disease it is dominantly inherited., (© 2010 Wiley-Liss, Inc.)

Autism spectrum disorders (ASD) and schizophrenia are neurodevelopmental disorders for which recent evidence indicates an important etiologic role for rare copy number variants (CNVs) and suggests common genetic mechanisms. We performed cytogenomic array analysis in a discovery sample of patients with neurodevelopmental disorders referred for clinical testing. We detected a recurrent 1.4 Mb deletion at 17q12, which harbors HNF1B, the gene responsible for renal cysts and diabetes syndrome (RCAD), in 18/15,749 patients, including several with ASD, but 0/4,519 controls. We identified additional shared phenotypic features among nine patients available for clinical assessment, including macrocephaly, characteristic facial features, renal anomalies, and neurocognitive impairments. In a large follow-up sample, the same deletion was identified in 2/1,182 ASD/neurocognitive impairment and in 4/6,340 schizophrenia patients, but in 0/47,929 controls (corrected p = 7.37 × 10⁻⁵). These data demonstrate that deletion 17q12 is a recurrent, pathogenic CNV that confers a very high risk for ASD and schizophrenia and show that one or more of the 15 genes in the deleted interval is dosage sensitive and essential for normal brain development and function. In addition, the phenotypic features of patients with this CNV are consistent with a contiguous gene syndrome that extends beyond RCAD, which is caused by HNF1B mutations only., (Copyright © 2010 The American Society of Human Genetics. Published by Elsevier Inc. All rights reserved.)

This study was an investigation of 90 patients referred to the Wessex Regional Genetics Laboratory for and negative by molecular cytogenetic analysis using array comparative genomic hybridization. This patient cohort represents typical referrals to a regional genetic centre. Methylation analysis was performed at 13 imprinted loci [PLAGL1, IGF2R, MEST, GRB10, H19, IGF2 DMR2 (IGF2P0), KCNQ1OT1 (KvDMR), MEG3, SNRPN, PEG3, GNAS (GNAS exon 1a and NESP55) and GNASAS]. In total 6/90 (6.67%) were shown to have a methylation defect, 2 of which were associated with known imprinting disorders: 1 patient had isolated hypomethylation at IGF2P0, an atypical epigenotype associated with Russell-Silver syndrome, and 1 showed hypomethylation at KvDMR consistent with a diagnosis of Beckwith-Wiedemann syndrome. A further 4 patients, 3 exhibiting complete hypermethylation, and 1 partial hypomethylation, had aberrations at IGF2R, the clinical significance of which remains unclear. This study demonstrates the potential utility of epigenetic investigation in routine diagnostic testing.

Monosomy 1p36 is the most common terminal deletion syndrome seen in humans, occurring in approximately 1 in 5,000 live births. Common features include mental retardation, characteristic dysmorphic features, hypotonia, seizures, hearing loss, heart defects, cardiomyopathy, and behavior abnormalities. Similar phenotypes are seen among patients with a variety of deletion sizes, including terminal and interstitial deletions, complex rearrangements, and unbalanced translocations. Consequently, critical regions harboring causative genes for each of these features have been difficult to identify. Here we report on five individuals with 200-823 kb overlapping deletions of proximal 1p36.33, four of which are apparently de novo. They present with features of monosomy 1p36, including developmental delay and mental retardation, dysmorphic features, hypotonia, behavioral abnormalities including hyperphagia, and seizures. The smallest region of deletion overlap is 174 kb and contains five genes; these genes are likely candidates for some of the phenotypic features in monosomy 1p36. Other genes deleted in a subset of the patients likely play a contributory role in the phenotypes, including GABRD and seizures, PRKCZ and neurologic features, and SKI and dysmorphic and neurologic features. Characterization of small deletions is important for narrowing critical intervals and for the identification of causative or candidate genes for features of monosomy 1p36 syndrome.

Chromosomal microarray (CMA) is increasingly utilized for genetic testing of individuals with unexplained developmental delay/intellectual disability (DD/ID), autism spectrum disorders (ASD), or multiple congenital anomalies (MCA). Performing CMA and G-banded karyotyping on every patient substantially increases the total cost of genetic testing. The International Standard Cytogenomic Array (ISCA) Consortium held two international workshops and conducted a literature review of 33 studies, including 21,698 patients tested by CMA. We provide an evidence-based summary of clinical cytogenetic testing comparing CMA to G-banded karyotyping with respect to technical advantages and limitations, diagnostic yield for various types of chromosomal aberrations, and issues that affect test interpretation. CMA offers a much higher diagnostic yield (15%-20%) for genetic testing of individuals with unexplained DD/ID, ASD, or MCA than a G-banded karyotype ( approximately 3%, excluding Down syndrome and other recognizable chromosomal syndromes), primarily because of its higher sensitivity for submicroscopic deletions and duplications. Truly balanced rearrangements and low-level mosaicism are generally not detectable by arrays, but these are relatively infrequent causes of abnormal phenotypes in this population (<1%). Available evidence strongly supports the use of CMA in place of G-banded karyotyping as the first-tier cytogenetic diagnostic test for patients with DD/ID, ASD, or MCA. G-banded karyotype analysis should be reserved for patients with obvious chromosomal syndromes (e.g., Down syndrome), a family history of chromosomal rearrangement, or a history of multiple miscarriages., (Copyright (c) 2010 The American Society of Human Genetics. Published by Elsevier Inc. All rights reserved.)

Congenital pancreatic hypoplasia is a rare cause of neonatal diabetes. We report on a series of three patients with pancreatic agenesis and congenital heart defects. All had abdominal scan evidence of pancreatic agenesis. In addition, Patient 1 had a ventricular septal defect, patent ductus arteriosus and pulmonary artery stenosis; Patient 2 had a truncus arteriosus and Patient 3 had tetralogy of Fallot. Two of the three patients have developmental delay. All three patients were isolated cases within the family. Investigations included sequencing of GCK, ABCC8, IPF1, NEUROD1, PTF1A, HNF1B, INS, ISL1, NGN3, HHEX, G6PC2, TCF7L2, SOX4, FOXP3 (Patients 1 and 2), GATA4 and KCNJ11 genes (all three patients), but no mutations were found. Genetic investigation to exclude paternal UPD 6, methylation aberrations and duplications of 6q24 was also negative in all three. 22q11 deletion was excluded in all three patients. Array CGH in Patient (1) showed a approximately 250 kb, paternally inherited duplication of chromosome 12q [arr cgh 12q24.33 (B35:CHR12:131808577-132057649++) pat], not found in the other two patients. Permanent neonatal diabetes mellitus due to pancreatic hypoplasia with congenital heart defects has been reported before and may represent a distinct condition. We discuss this rare association and review previously reported literature., (Copyright 2010 Wiley-Liss, Inc.)

Background: Congenital chromosome abnormalities are relatively common in our species and among structural abnormalities the most common class is balanced reciprocal translocations. Determining the parental origin of de novo balanced translocations may provide insights into how and when they arise. While there is a general paternal bias in the origin of non-recurrent unbalanced rearrangements, there are few data on parental origin of non-recurrent balanced rearrangements., Methods: The parental origin of a series of de novo balanced reciprocal translocations was determined using DNA from flow sorted derivative chromosomes and linkage analysis., Results: Of 27 translocations, we found 26 to be of paternal origin and only one of maternal origin. We also found the paternally derived translocations to be associated with a significantly increased paternal age (p<0.008)., Conclusion: Our results suggest there is a very pronounced paternal bias in the origin of all non-recurrent reciprocal translocations and that they may arise during one of the numerous mitotic divisions that occur in the spermatogonial germ cells prior to meiosis.

Background: Recurrent 15q13.3 microdeletions were recently identified with identical proximal (BP4) and distal (BP5) breakpoints and associated with mild to moderate mental retardation and epilepsy., Methods: To assess further the clinical implications of this novel 15q13.3 microdeletion syndrome, 18 new probands with a deletion were molecularly and clinically characterised. In addition, we evaluated the characteristics of a family with a more proximal deletion between BP3 and BP4. Finally, four patients with a duplication in the BP3-BP4-BP5 region were included in this study to ascertain the clinical significance of duplications in this region., Results: The 15q13.3 microdeletion in our series was associated with a highly variable intra- and inter-familial phenotype. At least 11 of the 18 deletions identified were inherited. Moreover, 7 of 10 siblings from four different families also had this deletion: one had a mild developmental delay, four had only learning problems during childhood, but functioned well in daily life as adults, whereas the other two had no learning problems at all. In contrast to previous findings, seizures were not a common feature in our series (only 2 of 17 living probands). Three patients with deletions had cardiac defects and deletion of the KLF13 gene, located in the critical region, may contribute to these abnormalities. The limited data from the single family with the more proximal BP3-BP4 deletion suggest this deletion may have little clinical significance. Patients with duplications of the BP3-BP4-BP5 region did not share a recognisable phenotype, but psychiatric disease was noted in 2 of 4 patients., Conclusions: Overall, our findings broaden the phenotypic spectrum associated with 15q13.3 deletions and suggest that, in some individuals, deletion of 15q13.3 is not sufficient to cause disease. The existence of microdeletion syndromes, associated with an unpredictable and variable phenotypic outcome, will pose the clinician with diagnostic difficulties and challenge the commonly used paradigm in the diagnostic setting that aberrations inherited from a phenotypically normal parent are usually without clinical consequences.

Trisomy and tetrasomy of distal chromosome 15q have rarely been reported. Although most of the described patients have some learning difficulties and are overgrown, the phenotype associated with distal trisomy/tetrasomy 15q is uncertain due to the small numbers of reported cases and the common co-occurrence of additional chromosome deletions in many patients with trisomy 15q. We present five individuals with overgrowth, learning difficulties and increased dosage of distal 15q. Partial trisomy 15q was identified in four of these cases. Two were generated through recombination of a parental pericentric inversion and two were generated through malsegregation of a maternal balanced 14;15 reciprocal translocation. In all four cases the trisomy can be considered "pure" as the 14p and 15p monosomies will exert no phenotypic effect. Partial tetrasomy 15q, as the result of an analphoid supernumerary chromosome derived from an inverted duplication of distal 15q, was identified in the fifth patient. In addition to the overgrowth and learning difficulties, all five had a characteristic facial appearance and three had renal anomalies. The gestalt consists of a long, thin face with a prominent chin and nose. Renal anomalies included renal agenesis, horseshoe kidney, and hydronephrosis. We provide further support for a distinct "15q overgrowth syndrome" caused by either trisomy or tetrasomy resulting in increased dosage of distal 15q. In addition we propose that renal anomalies and a distinctive facial appearance be considered major features of this condition., ((c) 2009 Wiley-Liss, Inc.)

Chromosome microdeletions or duplications are detected in 10-20% of patients with mental impairment and normal karyotypes. A few cases have been reported of mental impairment with microdeletions comprising tumor suppressor genes. By array-CGH we detected 4 mentally impaired individuals carrying de novo microdeletions sharing an overlapping segment of approximately 180 kb in 17p13.1. This segment encompasses 18 genes, including 3 involved in cancer, namely KCTD11/REN, DLG4/PSD95, and GPS2. Furthermore, in 2 of the patients, the deletions also included TP53, the most frequently inactivated gene in human cancers. The 3 tumor suppressor genes KCTD11, DLG4, and GPS2, in addition to the GABARAP gene, have a known or suspected function in neuronal development and are candidates for causing mental impairment in our patients. Among our 4 patients with deletions in 17p13.1, 3 were part of a Brazilian cohort of 300 mentally retarded individuals, suggesting that this segment may be particularly prone to rearrangements and appears to be an important cause (approximately 1%) of mental retardation. Further, the constitutive deletion of tumor suppressor genes in these patients, particularly TP53, probably confers a significantly increased lifetime risk for cancer and warrants careful oncological surveillance of these patients. Constitutional chromosome deletions containing tumor suppressor genes in patients with mental impairment or congenital abnormalities may represent an important mechanism linking abnormal phenotypes with increased risks of cancer., (Copyright 2009 S. Karger AG, Basel.)

Severe ocular malformations, including anophthalmia-microphthalmia (AM), are responsible for around 25% of severe visual impairment in childhood. Recurrent interstitial deletions of 14q22-23 are associated with AM and a wide range of extra-ocular phenotypes including brain anomalies. The homeobox gene OTX2 is located at 14q22.3 and has recently been identified as mutated in AM patients. Eight human OTX2 mutations have been reported in subjects with severe eye malformations, including AM, and variable developmental delay. We screened a novel AM cohort for mutations and deletions in OTX2, and identified four new mutations in six individuals and two cases of whole gene deletions. Our data suggest that OTX2 mutations and deletions account for 2-3% of AM cases., ((c) 2008 Wiley-Liss, Inc.)

The severe mental retardation and speech deficits associated with 22q13 terminal deletions have been attributed in large part to haploinsufficiency of SHANK3, which maps to all 22q13 terminal deletions, although more proximal genes are assumed to have minor effects. We report two children with interstitial deletions of 22q13 and two copies of SHANK3, but clinical features similar to the terminal 22q13 deletion syndrome, including mental retardation and severe speech delay. Both these interstitial deletions are completely contained within the largest terminal deletion, but do not overlap with the nine smallest terminal deletions. These interstitial deletions indicate that haploinsufficiency for 22q13 genes other than SHANK3 can have major effects on cognitive and language development. However, the relatively mild speech problems and normal cognitive abilities of a parent who transmitted her identical interstitial deletion to her more severely affected son suggests that the phenotype associated with this region may be more variable than terminal deletions and therefore contribute to the relative lack of correlation between clinical severity and size of terminal deletions. The phenotypic similarity between the interstitial deletions and non-overlapping small terminal 22q13 deletions emphasizes the general nonspecificity of the clinical picture of the 22q13 deletion syndrome and the importance of molecular analysis for diagnosis.

We present a 12-year-old with a de novo interstitial deletion of approximately 2.3Mb in chromosome band 17q24.2-q24.3, which was identified by array CGH. The most characteristic features in this case are posterior laryngeal cleft and the presence of numerous freckles and lentigines in childhood. Growth restriction, microcephaly and moderate mental retardation are also prominent features but are frequently seen with other chromosomal anomalies. The microdeletion causes haploinsufficiency of PRKAR1A (protein kinase, cAMP-dependent, regulatory 1alpha), which is known to cause Carney Complex but this diagnosis alone does not account for all of her problems and she therefore has 'Carney Complex plus'. This report illustrates the practical benefits associated with a clear cytogenetic diagnosis, as regular endocrinological and cardiac screening is required.

Background: Duplications and deletions in the human genome can cause disease or predispose persons to disease. Advances in technologies to detect these changes allow for the routine identification of submicroscopic imbalances in large numbers of patients., Methods: We tested for the presence of microdeletions and microduplications at a specific region of chromosome 1q21.1 in two groups of patients with unexplained mental retardation, autism, or congenital anomalies and in unaffected persons., Results: We identified 25 persons with a recurrent 1.35-Mb deletion within 1q21.1 from screening 5218 patients. The microdeletions had arisen de novo in eight patients, were inherited from a mildly affected parent in three patients, were inherited from an apparently unaffected parent in six patients, and were of unknown inheritance in eight patients. The deletion was absent in a series of 4737 control persons (P=1.1x10(-7)). We found considerable variability in the level of phenotypic expression of the microdeletion; phenotypes included mild-to-moderate mental retardation, microcephaly, cardiac abnormalities, and cataracts. The reciprocal duplication was enriched in nine children with mental retardation or autism spectrum disorder and other variable features (P=0.02). We identified three deletions and three duplications of the 1q21.1 region in an independent sample of 788 patients with mental retardation and congenital anomalies., Conclusions: We have identified recurrent molecular lesions that elude syndromic classification and whose disease manifestations must be considered in a broader context of development as opposed to being assigned to a specific disease. Clinical diagnosis in patients with these lesions may be most readily achieved on the basis of genotype rather than phenotype., (2008 Massachusetts Medical Society)

Langer-Giedion syndrome results from a microdeletion at 8q24.1 encompassing the EXT1 and the adjacent TRPS1 gene. We report on a boy with an oligo array-cgh characterized small microdeletion involving EXT1 alone but with some features of Langer-Giedion syndrome suggesting a functional disturbance of TRPS1. This boy, in addition to a mild Langer-Giedion like phenotype, also had some unusual features including prominent toe pads and fat pads on the soles of his feet similar to those described in Pierpont syndrome., (2008 Wiley-Liss, Inc.)

Context: Heterozygous, de novo mutations in the transcription factor SOX2 are associated with bilateral anophthalmia or severe microphthalmia and hypopituitarism. Variable additional abnormalities include defects of the corpus callosum and hippocampus., Objective: We have ascertained a further three patients with severe eye defects and pituitary abnormalities who were screened for mutations in SOX2. To provide further evidence of a direct role for SOX2 in hypothalamo-pituitary development, we have studied the expression of the gene in human embryonic tissues., Results: All three patients harbored heterozygous SOX2 mutations: a deletion encompassing the entire gene, an intragenic deletion (c.70&#95;89del), and a novel nonsense mutation (p.Q61X) within the DNA binding domain that results in impaired transactivation. We also show that human SOX2 can inhibit beta-catenin-driven reporter gene expression in vitro, whereas mutant SOX2 proteins are unable to repress efficiently this activity. Furthermore, we show that SOX2 is expressed throughout the human brain, including the developing hypothalamus, as well as Rathke's pouch, the developing anterior pituitary, and the eye., Conclusions: Patients with SOX2 mutations often manifest the unusual phenotype of hypogonadotropic hypogonadism, with sparing of other pituitary hormones despite anterior pituitary hypoplasia. SOX2 expression patterns in human embryonic development support a direct involvement of the protein during development of tissues affected in these individuals. Given the critical role of Wnt-signaling in the development of most of these tissues, our data suggest that a failure to repress the Wnt-beta-catenin pathway could be one of the underlying pathogenic mechanisms associated with loss-of-function mutations in SOX2.

A two-year-and-eight-month-old girl presented with clitoromegaly and short stature. Two cell lines, 45,X and 46,X,idic(Y)(q11.2), were observed. Cytogenetic and fluorescence in situ hybridisation investigations were carried out on her peripheral lymphocytes and gonadal cells, to determine the genotype-phenotype effect with respect to differential tissue distribution, effects of the sex determining region of the Y chromosome, and the break-points in the azoospermia factor region.

We report the analyses of breakpoints in 31 phenotypically normal and 14 abnormal carriers of balanced translocations. Our study assesses the differences between balanced translocations in normal carriers and those in abnormal carriers, focusing on the presence of genomic imbalances at the breakpoints or elsewhere in the genome, presence of cryptic chromosome rearrangements, and gene disruption. Our hypothesis is that all four features will be associated with phenotypic abnormalities and absent or much less frequent in a normal population. In the normal cohort, we identified neither genomic imbalances at the breakpoints or elsewhere in the genome nor cryptic chromosome rearrangements. In contrast, we identified candidate disease-causing imbalances in 4/14 abnormal patients. These were three breakpoint associated deletions and three deletions unrelated to the breakpoints. All six de novo deletions originated on the paternally inherited chromosome. Additional complexity was also present in one of these cases. Gene disruption by the breakpoints was present in 16/31 phenotypically normal individuals and in 5/14 phenotypically abnormal patients. Our results show that translocations in phenotypically abnormal patients are molecularly distinct from those in normal individuals: the former are more likely to be associated with genomic imbalances at the breakpoints or elsewhere and with chromosomal complexity, whereas the frequency of gene disruption is similar in both normal and abnormal translocation carriers.

A series of 125 patients referred primarily with aniridia classified as either sporadic (74), familial (24), or in association with WAGR syndrome (14) or other malformations (13) was analysed for mutations, initially by karyotyping and targeted FISH analysis of chromosome 11p13. These methods identified mutations in a significant proportion of patients, 34/125 (27%). Two cases had chromosome rearrangements involving 11p13, 16 cases had visible deletions, and 16 cases had cryptic deletions identified by FISH. The frequency of cryptic deletions in familial aniridia was 27% and in sporadic isolated aniridia was 22%. Of the 14 cases referred with WAGR syndrome, 10 (71%) had chromosomal deletions, 2 cryptic and 8 visible. Of the 13 cases with aniridia and other malformations, 5 (38%) had a chromosomal rearrangement or deletion. In 37 cases with no karyotypic or cryptic chromosome abnormality, sequence analysis of the PAX6 gene was performed. Mutations were identified in 33 cases; 22 with sporadic aniridia, 10 with familial aniridia and 1 with aniridia and other non-WAGR syndrome associated anomalies. Overall, 67 of 71 cases (94%) undergoing full mutation analysis had a mutation in the PAX6 genomic region., ((c) 2008 Wiley-Liss, Inc.)

Background: Developmental eye anomalies, which include anophthalmia (absent eye) or microphthalmia (small eye) are an important cause of severe visual impairment in infants and young children. Heterozygous mutations in SOX2, a SOX1B-HMG box transcription factor, have been found in up to 10% of individuals with severe microphthalmia or anophthalmia and such mutations could also be associated with a range of non-ocular abnormalities., Methods: We performed mutation analysis on a new cohort of 120 patients with congenital eye abnormalities, mainly anophthalmia, microphthalmia and coloboma. Multiplex ligation-dependent probe amplification (MLPA) and fluorescence in situ hybridisation (FISH) were used to detect whole gene deletion., Results: We identified four novel intragenic SOX2 mutations (one single base deletion, one single base duplication and two point mutations generating premature translational termination codons) and two further cases with the previously reported c.70del20 mutation. Of 52 patients with severe microphthalmia or anophthalmia analysed by MLPA, 5 were found to be deleted for the whole SOX2 gene and 1 had a partial deletion. In two of these, FISH studies identified sub-microscopic deletions involving a minimum of 328 Kb and 550 Kb. The SOX2 phenotypes include a patient with anophthalmia, oesophageal abnormalities and horseshoe kidney, and a patient with a retinal dystrophy implicating SOX2 in retinal development., Conclusion: Our results provide further evidence that SOX2 haploinsufficiency is a common cause of severe developmental ocular malformations and that background genetic variation determines the varying phenotypes. Given the high incidence of whole gene deletion we recommend that all patients with severe microphthalmia or anophthalmia, including unilateral cases be screened by MLPA and FISH for SOX2 deletions.

Using fluorescent in situ hybridization (FISH) with the probe p15 (D15Z1), we investigated the distribution of the polymorphic 15p signal which has been reported to occur on acrocentric chromosomes in addition to chromosome 15. The short arm of chromosome 15 has a characteristic signal pattern when hybridized with the FISH probe D15Z1. However, the D15Z1 signal can occasionally be seen on the short arm of other acrocentric chromosomes. We studied the distribution of the D15Z1 probe in 1657 patients consisting both of individuals with a normal karyotype and those with a variety of chromosome abnormalities involving the acrocentric chromosomes. Our results show that one in six individuals, regardless of their patient ascertainment category or karyotypic status, had one or more additional D15Z1 signals, and that there were no significant differences in the distribution of extra signals among the patient groups.

A de novo complex chromosome rearrangement (CCR) found in a phenotypically abnormal boy was characterized by G-bands, FISH with subtelomere probes, and M-FISH. The G-banding analysis revealed involvement of chromosomes 1, 2, 3, 15, and 18 with (at least) eight breakpoints, five nonreciprocal translocations (1q --> 2q --> 8q --> 15q --> 2p --> 1q), and a 3p insertion into the der(2); there was also a presumptive deletion of 1q41. The 5 derivatives were described as follows: der(1)(1pter --> 1q32.3?::2p21--> 2pter),der(2)(1qter --> 1q42?::2q24.2 --> 2p21::3p13 --> 3p26::15q15 --> 15qter),der(3)(3qter --> 3p13:),der(15)(15pter --> 15q15::18q11 --> 18qter),der(18)(18pter --> 18q11::2q24.2 --> 2qter). The molecular assays confirmed the segmental composition of each derivative and documented the localization of most relevant telomeres. In addition to the novelty of the 1, 2, 3, 15 and 18 combination, this CCR may also be unique in the sense that it represents a cluster of 6 nonreciprocal transpositions regardless of the occurrence (or lack thereof) of secondary unbalances. Finally, there appears to be an excess of CCRs in fetuses conceived by intracytoplasmic sperm injection.