Directly Transmitted 12.3-Mb Deletion with a Consistent Phenotype in the Variable 11q21q22.3 Region.

A phenotype is emerging for the proximal pair of G-dark bands in 11q (11q14.1 and q14.3) but not yet for the distal pair (11q22.1 and q22.3). A mother and daughter with the same directly transmitted 12.3-Mb interstitial deletion of 11q21q22.3 (GRCh37: 93,551,765-105,817,723) both had initial feeding difficulties and failure to thrive, speech delay, learning difficulties, and mild dysmorphism. Among 17 patients with overlapping deletions, developmental or speech delay, dysmorphism, hypotonia, intellectual disability or learning difficulties, short stature, and coloboma were each found in 2 or more. These results may provide the basis for a consistent phenotype for this region. Among the 53 deleted and additional breakpoint genes, CNTN5, YAP1, and GRI4 were the most likely candidates. Non-penetrance of haploinsufficient genes and dosage compensation among related genes may account for the normal cognition in the mother and variable phenotypes that can extend into the normal range.

Incomplete penetrance, variable expressivity, or dosage insensitivity in four families with directly transmitted unbalanced chromosome abnormalities.

The direct transmission of microscopically visible unbalanced chromosome abnormalities (UBCAs) is rare and usually has phenotypic consequences. Here we report four families in which a normal phenotype was initially found in one or more family members. Each UBCA was interpreted with regard to overlapping examples and factors previously associated with transmitted imbalances including incidental ascertainment, low gene density, benign copy number variation (CNV) content, and gene relatedness. A 4.56 Mb deletion of 8p23.1-p23.2 was thought to be causal in the affected proband but showed incomplete penetrance in her mother and sibling (Family 1). Incomplete penetrance was also associated with a 10.88 Mb duplication of 13q21.31-q22.1 (Family 3) and dosage insensitivity with a 17.6 Mb deletion of 22pter-q11.21 (Family 4) that were both ascertained at prenatal diagnosis and each found in 4 unaffected family members. The 22pter-q11.21 deletion is part of a region with high benign CNV content and supports the mapping of cat eye syndrome to a 600 kb interval of 22q11.1-q11.21. Low gene densities of less than 2.0 genes/Mb were found in each of these three families but only after segmentally duplicated genes were excluded from the deletions of 8p and 22q. In contrast, gene density was average and variable expressivity associated with a 3.59 Mb duplication of 8p23.1 incidentally ascertained for paternal infertility (Family 2). Our results indicate that a greater degree of direct parental transmission, incomplete penetrance, and variable expression are features of both sub-microscopic CNVs and UBCAs with relatively low gene and high benign CNV content.

Inside the 8p23.1 duplication syndrome; eight microduplications of likely or uncertain clinical significance.

The 8p23.1 duplication syndrome (8p23.1 DS) is a recurrent genomic condition with an estimated prevalence of 1 in 58,000. The core 3.68 Mb duplication contains 32 genes of which five are currently candidates for the phenotypic features. Here we describe four patients and five families with eight microduplications of 8p23.1 ranging from 187 to 1082 kb in size and one atypical duplication of 4 Mb. These indicate that a minimal region of overlap (MRO) in medial 8p23.1 can give rise to features of 8p23.1 DS including developmental delay, dysmorphism, macrocephaly and otitis media, but not congenital heart disease (CHD). This MRO spans 776 kb (chr8:10,167,881-10,943,836 hg19) and contains SOX7 and seven of the other 32 core 8p23.1 DS genes. In centromeric 8p23.1, microduplications including GATA4 can give rise to non-syndromic CHD but the clinical significance of two smaller centromeric microduplications without GATA4 was uncertain due to severe neurological profiles not usually found in 8p23.1 DS. The clinical significance of three further 8p23.1 microduplications was uncertain due to additional genetic factors without which the probands might not have come to medical attention. Variable expressivity was indicated by the almost entirely unaffected parents in all five families and the mildly affected sibling in one. Intronic interruptions of six genes by microduplication breakpoint intervals had no apparent additional clinical consequences. Our results suggest that 8p23.1 DS is an oligogenetic condition largely caused by the duplication and interactions of the SOX7 and GATA4 transcription factors.

8p23.1 duplication syndrome; common, confirmed, and novel features in six further patients.

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.

16p11.2-p12.2 duplication syndrome; a genomic condition differentiated from euchromatic variation of 16p11.2.

Chromosome 16 contains multiple copy number variations (CNVs) that predispose to genomic disorders. Here, we differentiate pathogenic duplications of 16p11.2-p12.2 from microscopically similar euchromatic variants of 16p11.2. Patient 1 was a girl of 18 with autism, moderate intellectual disability, behavioural difficulties, dysmorphic features and a 7.71-Mb (megabase pair) duplication (16:21 521 005-29 233 146). Patient 2 had a 7.81-Mb duplication (16:21 382 561-29 191 527), speech delay and obsessional behaviour as a boy and, as an adult, short stature, macrocephaly and mild dysmorphism. The duplications contain 65 coding genes of which Polo-like kinase 1 (PLK1) has the highest likelihood of being haploinsufficient and, by implication, a triplosensitive gene. An additional 1.11-Mb CNV of 10q11.21 in Patient 1 was a possible modifier containing the G-protein-regulated inducer of neurite growth 2 (GPRIN2) gene. In contrast, the euchromatic variants in Patients 3 and 4 were amplifications from a 945-kb region containing non-functional immunoglobulin heavy chain (IGHV), hect domain pseudogene (HERC2P4) and TP53-inducible target gene 3 (TP53TG3) loci in proximal 16p11.2 (16:31 953 353-32 898 635). Paralogous pyrosequencing gave a total copy number of 3-8 in controls and 8 to >10 in Patients 3 and 4. The 16p11.2-p12.2 duplication syndrome is a recurrent genomic disorder with a variable phenotype including developmental delay, dysmorphic features, mild to severe intellectual disability, autism, obsessive or stereotyped behaviour, short stature and anomalies of the hands and fingers. It is important to differentiate pathogenic 16p11.2-p12.2 duplications from harmless, microscopically similar euchromatic variants of proximal 16p11.2, especially at prenatal diagnosis.

Transmitted deletions of medial 5p and learning difficulties; does the cadherin cluster only become penetrant when flanking genes are deleted?

The central portion of the short arm of chromosome 5 is unusual in that large, cytogenetically visible interstitial deletions segregate in families with and without phenotypic consequences. Here we present a family in which a transmitted interstitial deletion of 5p13.3 to 5p14.3 co-segregated with learning and/or behavioral difficulties in six family members. Facial dysmorphism was not striking but a father and daughter both had lacrimal fistulae. The deletion was 12.23 Mb in size (chr5:20,352,535-32,825,775) and contained fifteen known protein coding genes. Five of these (GOLPH3; MTMR12; ZFR; SUB1; and NPR3) and an ultra-conserved microRNA (hsa-miR-579) were present in an 883 kb candidate gene region in 5p13.3 that was deleted in the present family but not in previously reported overlapping benign deletions. Members of the cadherin precursor gene cluster, with brain specific expression, were deleted in both affected and benign deletion families. The candidate genes in 5p13.3 may be sufficient to account for the consistent presence or absence of phenotype in medial 5p deletions. However, we consider the possibility of position effects in which CDH6, and/or other cadherin genes, become penetrant when adjacent genes, or modifiers of gene expression, are also deleted. This could account for the absence of intellectual disability in benign deletions of the cadherin cluster, the cognitive phenotype in medial 5p deletion syndrome and the greater severity of intellectual disability in patients with cri-du-chat syndrome and deletions of 5p15 that extend into the region deleted in the present family.

Further molecular and clinical delineation of the Wisconsin syndrome phenotype associated with interstitial 3q24q25 deletions.

Deletions of the distal 3q22.3 region encompassing the gene forkhead transcription factor FOXL2 (FOXL2) usually result in intellectual disability (ID) and the highly recognizable blepharophimosis-ptosis-epicanthus inversus syndrome (BPES). We encountered three patients with molecularly defined interstitial deletions distal to the FOXL2 gene. They present with remarkably similar manifestations comprising variable ID, a coarse facial appearance, including prominent nose and eyebrows, hypogonadism and skin pigmentation abnormalities, and they share an approximately 8.8 Mb overlapping 3q24q25 deletion. Interestingly, one of the present patients was described previously in a clinical report with emphasis on her clinical similarity to the Wisconsin syndrome, suggesting that Wisconsin syndrome might be caused by a (micro) deletion within the 3q24q25 region.

A de novo 4q34 interstitial deletion of at least 9.3 Mb with no discernible phenotypic effect.

Cytogenetically visible imbalances without phenotypic effect are still rare despite the extent of large-scale copy number variation in the normal population revealed by array CGH. Here we report on a phenotypically normal 30-year-old female with a de novo, cytogenetically visible, interstitial deletion of band 4q34. She was referred following three successive miscarriages, one of which was an intra-uterine death with subendocardial fibroelastosis and dilated cardiomyopathy. There was no other notable medical or family history, she was of normal intelligence and had no dysmorphic features. FISH and Array CGH with a customized 1 Mb BAC array showed that the deletion is a minimum of 9.3 and a maximum of 10.7 Mb in size, between approximately 173 Mb in 4q34.1 and approximately 182 Mb in 4q34.3. The deletion contains only 23 known coding genes giving a low average gene density of approximately 2 genes/Mb. This case further illustrates that (1) sizeable imbalances can be associated with apparent phenotypic normality, (2) gene density is a better guide to possible phenotypic consequences than aberration size, and (3) it is not always safe to assume that de novo imbalances will be causal.

15q overgrowth syndrome: a newly recognized phenotype associated with overgrowth, learning difficulties, characteristic facial appearance, renal anomalies and increased dosage of distal chromosome 15q.

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.

Transmitted duplication of 8p23.1-8p23.2 associated with speech delay, autism and learning difficulties.

Duplications of distal 8p with and without significant clinical phenotypes have been reported and are often associated with an unusual degree of structural complexity. Here, we present a duplication of 8p23.1-8p23.2 ascertained in a child with speech delay and a diagnosis of ICD-10 autism. The same duplication was found in his mother who had epilepsy and learning problems. A combination of cytogenetic, FISH, microsatellite, MLPA and oaCGH analysis was used to show that the duplication extended over a minimum of 6.8 Mb between 3 539 893 and 10 323 426 bp. This interval contains 32 novel and 41 known genes, of which only microcephalin (MCPH1) is a plausible candidate gene for autism at present. The distal breakpoint of the duplicated region interrupts the CSMD1 gene in 8p23.2 and the medial breakpoint lies between the MSRA and RP1L1 genes in 8p23.1.An interchromosomal insertion between a normal and polymorphically inverted chromosome 8 is proposed to explain the origin of this duplication. Further mapped imbalances of distal 8p are needed to determine whether the autistic component of the phenotype in this family results from the cumulative imbalance of many genes or dosage imbalance of an individual susceptibility gene.

Defensins and the dynamic genome: what we can learn from structural variation at human chromosome band 8p23.1.

Over the past four years, genome-wide studies have uncovered numerous examples of structural variation in the human genome. This includes structural variation that changes copy number, such as deletion and duplication, and structural variation that does not change copy number, such as orientation and positional polymorphism. One region that contains all these types of variation spans the chromosome band 8p23.1. This region has been studied in some depth, and the focus of this review is to examine our current understanding of the variation of this region. We also consider whether this region is a good model for other structurally variable regions in the genome and what the implications of this variation are for clinical studies. Finally, we discuss the bioinformatics challenges raised, discuss the evolution of the region, and suggest some future priorities for structural variation research.

8p23.1 duplication syndrome; a novel genomic condition with unexpected complexity revealed by array CGH.

The 8p23.1 deletion syndrome is established but not an equivalent duplication syndrome. Here, we report five patients; a de novo prenatal case and two families in which 8p23.1 duplications have been directly transmitted from mothers to children. Dual-colour fluorescent in situ hybridisation, multiplex ligation-dependent probe amplification analysis and customised oligonucleotide array comparative genomic hybridisation (oaCGH) indicated an approximately 3.75 Mb duplication of most of band 8p23.1 between the olfactory receptor/defensin repeats (ORDRs) in all cases. However, oaCGH revealed an additional duplication of 500 kb adjacent to the proximal ORDR in Family 1 and an additional deletion of 3.14 Mb within the Nablus Mask-Like Facial Syndrome region of 8q22.1 in Family 2. Copy number variation at introns 4-5 of the GATA4 gene was also identified. This 8p23.1 duplication syndrome is associated with a characteristic facial phenotype including a prominent forehead and arched eyebrows. Adrenal insufficiency, Tetralogy of Fallot, partial 2/3 syndactyly of the toes and cleft palate in some individuals may be explained by ascertainment bias, incomplete penetrance and/or the presence of the microdeletion in Family 2. The duplication is compatible with normal early childhood development but, although our adult cases live independent lives with varying degrees of support, learning difficulties have been experienced by some family members. We conclude that the 8p23.1 duplication syndrome is a genomic condition with an emerging but variable phenotype that may be under-diagnosed. Our results demonstrate that direct transmission does not distinguish genuine duplications from euchromatic variants and illustrate the power of array CGH to reveal unexpected additional imbalances in affected patients.

The variant inv(2)(p11.2q13) is a genuinely recurrent rearrangement but displays some breakpoint heterogeneity.

Human chromosome 2 contains large blocks of segmental duplications (SDs), both within and between proximal 2p and proximal 2q, and these may contribute to the frequency of the common variant inversion inv(2)(p11.2q13). Despite their being cytogenetically homogeneous, we have identified four different breakpoint combinations by fluorescence in situ hybridization mapping of 40 cases of inv(2)(p11.2q13) of European origin. For the vast majority of inversions (35/40), the breakpoints fell within the same spanning BACs, which hybridized to both 2p11.2 and 2q13 on the normal and inverted homologues. Sequence analysis revealed that these BACs contain a significant proportion of intrachromosomal SDs with sequence homology to the reciprocal breakpoint region. In contrast, BACs spanning the rare breakpoint combinations contain fewer SDs and with sequence homology only to the same chromosome arm. Using haplotype analysis, we identified a number of related family subgroups with identical or very closely related haplotypes. However, the majority of cases were not related, demonstrating for the first time that the inv(2)(p11.2q13) is a truly recurrent rearrangement. Therefore, there are three explanations to account for the frequent observation of the inv(2)(p11.2q13): the majority have arisen independently in different ancestors, while a minority either have been transmitted from a common founder or have different breakpoints at the molecular cytogenetic level.

Parental mosaic trisomy 21 detected following maternal cell contamination of an amniotic fluid specimen from a normal male pregnancy.

We report a case of maternal mosaic trisomy 21 ascertained at prenatal diagnosis as a result of maternal cell contamination of an amniotic fluid sample. A 34 year old female was referred for karyotyping because of a previous trisomy 21 pregnancy. Chromosome analysis of primary in situ cultures showed a karyotype of 47,XX, + 21[6]/46,XY[32]/46,XX[2]. Molecular testing demonstrated maternal cell contamination of the amniotic fluid sample and G-banded karyotyping of maternal blood showed that 3/200 cells had trisomy 21, consistent with the mother being a Down syndrome mosaic. A normal male baby with a 46,XY chromosome complement was delivered at 30 weeks. This case emphasises the need for close collaboration between cytogenetic and molecular genetics laboratories in resolving unusual cases of mosaicism.

Novel deletion variants of 9q13-q21.12 and classical euchromatic variants of 9q12/qh involve deletion, duplication and triplication of large tracts of segmentally duplicated pericentromeric euchromatin.

Large-scale copy number variation that is cytogenetically visible in normal individuals has been described as euchromatic variation but needs to be distinguished from pathogenic euchromatic deletion or duplication. Here, we report eight patients (three families and two individuals) with interstitial deletions of 9q13-q21.12. Fluorescence in situ hybridisation with a large panel of BACs showed that all the deleted clones were from extensive tracts of segmentally duplicated euchromatin, copies of which map to both the long and short arms of chromosome 9. The variety of reasons for which these patients were ascertained, and the phenotypically normal parents, indicates that this is a novel euchromatic variant with no phenotypic effect. Further, four patients with classical euchromatic variants of 9q12/qh or 9p12 were also shown to have duplications or triplications of this segmentally duplicated material common to both 9p and 9q. The cytogenetic boundaries between the segmentally duplicated regions and flanking unique sequences were mapped to 9p13.1 in the short arm (BAC RP11-402N8 at 38.7 Mb) and to 9q21.12 in the long arm (BAC RP11-88I18 at 70.3 Mb). The BACs identified in this study should in future make it possible to differentiate between clinically significant deletions or duplications and euchromatic variants with no established phenotypic consequences.