Nablus mask-like facial syndrome: Report of an atypical case with 8q21.3-q22.1 deletion.

Nablus mask-like facial syndrome (NMLFS) is a rare condition characterized by unique facial features, initially described in a 4-year-old boy from Nablus, Palestine. These features include expressionless facial appearance, tight facial skin, blepharophimosis, sparse eyebrows, and a flat nose. Genetic studies have identified a deletion of 8q22.1 as the cause of the syndrome, however while 26 patients have been reported with the deletion, only 13 displayed the characteristic facial features. Here we report on a 35-year-old male with 8q21.3-q22.1 deletion identified by whole exome sequencing and Chromosomal microarray analysis (CMA) that presents with typical and atypical features, including neurodevelopmental disorder, mild facial features, and myopathy, which has not been described in a patient with NMLFS to date. Further research will be required to understand the underlying pathogenetic mechanism of this rare genetic disorder.

Prenatal Array-CGH Detection of 3q26.32q26.33 Interstitial Deletion Encompassing the SOX2 Gene: Ultrasound, Pathological, and Cytogenetic Findings.

Background: SOX2 disorders are associated with anophthalmia-esophageal-genital syndrome or microphthalmia, syndromic 3 (MCOPS3- # 206900). Case Report: We describe a third fetal case with a de novo 3q26.32q26.33 deletion extending for 4.31 Mb, detected in a 15-week fetus. After legal interruption of pregnancy, at autopsy, the fetus presented bilateral microphthalmia, right cleft lip and palate, bilateral cerebral ventriculomegaly and dilated third ventricle, microcystic left lung, and intestinal malrotation. Histologically, the left lung showed congenital pulmonary airway malformation (CPAM) type 2. Retinal dysplasia was found in both eyes. Discussion/Conclusion: The human SOX2 gene (OMIM #184429) is located on chromosome 3 at position q26.3-27 and encodes a transcription factor involved in the development of the central and peripheral nervous systems, retina, and lung. In our case, the combination of cerebral, retinal, and pulmonary anomalies, not previously described, are consistent with SOX2 haploinsufficiency due to chromosomal deletion.

Detecting regions of homozygosity improves the diagnosis of pathogenic variants and uniparental disomy in pediatric patients.

Chromosomal microarray analysis using single nucleotide polymorphism probes can detect regions of homozygosity (ROH). This confers a potential utility in revealing autosomal recessive (AR) diseases and uniparental disomy (UPD). Results of genetic testing among pediatric patients from 2015 to 2019 were evaluated. Diagnostic findings with detected ROH from large consecutive case series in the literature were reviewed. Of 2050 pediatric patients, 65 (3%) had one or more ROH and 31 (53%) had follow-up whole exome sequencing (WES) and methylation studies. Seven homozygous variants were detected and four of them from three patients (9.6%) were within the detected ROH and classified as pathogenic or likely pathogenic variants for AR diseases. One patient (3%) had segmental UPD15q for a diagnosis of Prader-Willi syndrome. Additive diagnostic yield from ROH reporting was at least 0.2% (4/2050) of pediatric patients. These results were consistent with findings from several large case series reported in the literature. Detecting ROH had an estimated baseline predictive value of 10% for AR diseases and 3% for UPD. Consanguinity revealed by multiple ROH was a strong predictor for AR diseases. These results provide evidence for genetic counseling and recommendation of follow-up WES and methylation studies for pediatric patients reported with ROH.

Genetic Analysis of Circulating Tumour Cells.

The classification of human cancers has traditionally relied on the tissue of origin, the histologic appearance and anatomical extent of disease, otherwise referred to as grade and stage. However, this system fails to explain the highly variable clinical behaviour seen for any one cancer. Molecular characterization through techniques such as next-generation sequencing (NGS) has led to an appreciation of the extreme genetic heterogeneity that underlies most human cancers. Because of the difficulties associated with fresh tissue biopsy, interest has increased in using circulating tumour material, such as circulating tumour cells (CTCs), as a non-invasive way to access tumour tissue. CTC enumeration has been demonstrated to have prognostic value in metastatic breast, colon and prostate cancers. Recent studies have also shown that CTCs are suitable material for molecular characterization, using techniques such as reverse transcription-polymerase chain reaction (RT-PCR), fluorescence in situ hybridization (FISH), array comparative genomic hybridization (aCGH) and NGS. Furthermore, genetic analysis of CTCs may be more suitable to study tumour heterogeneity and clonal evolution than fresh tissue biopsy. Whether blood-based biopsy techniques will be accepted as a replacement to fresh tissue biopsies remains to be seen, but there is reason for optimism. While significant barriers to this acceptance exist, blood-based biopsy techniques appear to be reliable and representative alternatives to fresh tissue biopsy.

Molecular Cytogenomic Characterization of the Murine Breast Cancer Cell Lines C-127I, EMT6/P and TA3 Hauschka.

BACKGROUND: To test and introduce effective and less toxic breast cancer (BC) treatment strategies, animal models, including murine BC cell lines, are considered as perfect platforms. Strikingly, the knowledge on the genetic background of applied BC cell lines is often sparse though urgently necessary for their targeted and really justified application. METHODS: In this study, we performed the first molecular cytogenetic characterization for three murine BC cell lines C-127I, EMT6/P and TA3 Hauschka. Besides fluorescence in situ hybridization-banding, array comparative genomic hybridization was also applied. Thus, overall, an in silico translation for the detected imbalances and chromosomal break events in the murine cell lines to the corresponding homologous imbalances in humans could be provided. The latter enabled a comparison of the murine cell line with human BC cytogenomics. RESULTS: All three BC cell lines showed a rearranged karyotype at different stages of complexity, which can be interpreted carefully as reflectance of more or less advanced tumor stages. CONCLUSIONS: Accordingly, the C-127I cell line would represent the late stage BC while the cell lines EMT6/P and TA3 Hauschka would be models for the premalignant or early BC stage and an early or benign BC, respectively. With this cytogenomic information provided, these cell lines now can be applied really adequately in future research studies.

Fetal Cystic Hygroma Associated with Terminal 2p25.1 Duplication and Terminal 3p25.3 Deletion: Cytogenetic, Fluorescent in Situ Hybridization and Microarray Familial Characterization of Two Different Chromosomal Structural Rearrangements.

We report a prenatally diagnosed case of partial trisomy 2p and partial monosomy 3p, resulting from unbalanced translocation (2;3)(p25.1;p25.3) of paternal origin. Parents were non consanguineous Caucasians, with familial history of recurrent miscarriages on the father's side. Detailed sonographic examination of the fetus showed a septated cystic hygroma measuring 6 mm at 13 weeks' gestation. Karyotyping and fluorescent in situ hybridization (FISH) analysis of cultured amniotic fluid cells revealed an unbalanced translocation der(3)t(2;3)(p25.1; p25.3) and apparently balanced inv(3)(p13p25.3) in a fetus. Parental cytogenetic evaluation using karyotyping and FISH analysis showed the presence of both a balanced translocation and a paracentric inversion in father t(2;3) (p25.1;p25.3) inv(3)(p13p25.3). Microarray analysis showed a 11.6 Mb deletion at 3p26.3-p25.3 and duplication of 10.5 Mb at the 2p25.3-p25 region. The duplicated region at 2p25.1p25.3 contains 45 different genes, where 12 are reported as OMIM morbid genes with different phenotypical implications. The deleted region at 3p26.3-p25.3 contains 65 genes, out of which 27 are OMIM genes. Three of these (CNTN4, SETD5 and VHL) were curated by Clingene Dosage Gene Map and were given a high haplo-insufficiency score. Genes affected by the unbalanced translocation could have contributed to some specific phenotypic changes of the fetus in late pregnancy. The application of different cytogenetic methods was essential in our case, allowing the detection of different types of structural chromosomal aberrations and more thorough genetic counseling for future pregnancies.

A Very Rare Partial Trisomy Syndrome: De Novo Duplication of 16q12.1q23.3 in a Turkish Girl with Developmental Delay and Facial Dysmorphic Features.

Trisomy 16 is the most common type of autosomal trisomy associated with spontaneous abortion and is incompatible with life. Upon examining previously reported cases of partial chromosome 16q duplication, it was noted that the majority of cases had complex chromosomal abnormalities due to parental balanced chromosomal translocation carriage. The clinical presentation of very rare pure partial trisomy 16q cases was associated with congenital anomalies, facial dysmorphic findings and intellectual disability. In this study, we evaluated the physical characteristics and genetic data of an 8-month-old girl with developmental delay and facial dysmorphic features. Dysmorphic features including prominent metopic suture, synophrys, asymmetric head shape, triangular and asymmetric face, telecanthus, epicanthal folds, down-slanting palpebral fissures, microphthalmia of the left eye, anteverted nares, smooth and tented philtrum, microretrognathia, low-set posteriorly rotated ears, auricular pits, high-arched palate, thin upper lip and hypotonia were recorded. Her karyotype was 46,XX,add(16)(q24). To identify the extension of the duplicated section, array comparative genomic hybridization (aCGH) analysis was performed, which showed a de novo 29.8 Mb duplication [arr[hgl9] 16q12.1q23.3(52459169-82285105) x 3], interpreted to be pathogenic. We present this case report to clarify the clinical findings of a rare chromosomal anomaly, discuss the genes that may be related to the phenotype and advance the literature in terms of knowledge regarding genotypephenotype correlation.

Array-comparative Genomic Hybridization Results in Clinically Affected Cases with Apparently Balanced Chromosomal Rearrangements.

Carriers of apparently balanced chromosomal rearrangements (ABCRs) have a 2-3-fold higher risk of carrying an abnormal phenotype, when compared to the average population. Apparently balanced chromosomal rearrangements can be imbalanced at the submicroscopic level, and changes in the gene structure, formation of a new chimeric gene, gain or loss of function of the genes and altered imprinting pattern may also affect the phenotype. Chromosomal microarray (CMA) is an efficient tool to detect submicroscopic imbalances at the breakpoints as well as in the whole genome. We aimed to determine the effectiveness of array-comparative genomic hybridization (aCGH) application in phenotypically affected cases with ABCRs at a single center from Turkey. Thirty-four affected cases (13 prenatal, 21 postnatal) carrying ABCRs were investigated with CMA. In postnatal series, ABCRs were familial in 7 and de novo in 14 cases. Seven de novo cases were imbalanced (in postnatal series 33.3% and in de novo cases 50.0%). Out of 13 prenatal cases, five were familial and eight were de novo in origin and two de novo cases were imbalanced (in 15.4% prenatal series and in 25.0% de novo cases). No cryptic imbalance was observed in familial cases. The anomaly rates with array studies ranged between 14.3-25.0% in familial and between 20.0-57.5% in de novo cases of postnatal series in the literature. Studies focused on prenatal ABCR cases with abnormal ultrasound findings are limited and no submicroscopic imbalance was reported in the cohorts. When de novo postnatal or prenatal results were combined, the percentage of abnormalities detected by CMA was 40.9%. Taking this contribution into consideration, all ABCRs should be investigated by CMA even if the fetal ultrasound findings are normal.

A Systematic Clinical Review of Prenatally Diagnosed Tetrasomy 9p.

Tetrasomy 9p was first described in 1973 and approximately 68 cases with a variable phenotype have been reported to date with 22 of them being detected prenatally. The objective of this study was to review prenatally-reported cases of tetrasomy 9p thus far and to identify ultrasound phenotypes that may be suggestive of this specific syndrome. A PubMed database search was done in February 2018 without any restriction of publication date orjournals, with the use of the following keywords: tetrasomy 9p, tetrasomy 9p prenatal, mosaic tetrasomy 9p, mosaic tetrasomy 9p prenatal, isochromosome 9p, duplication 9p prenatal, trisomy 9p prenatal. Reported cases were included if the clinical presentation and diagnostic approach of each case was clearly described. The most common characteristics of prenatally-detected tetrasomy 9p are intrauterine growth retardation (IUGR, 57.0%), central nervous system (CNS) abnormalities (59.0%), skeletal anomalies (29.0%), genitourinary and renal anomalies (29.0%) and cardiac defects (29.0%). The phenotypic spectrum of tetrasomy 9p is rather unspecific as these findings are commonly associated with other chromosome anomalies, as well as microdeletion/microduplication or monogenic syndromes. The combination of early fetal morphology and diagnostic genetic testing enables a definite tetrasomy 9p diagnosis and effective further pregnancy management.

A Novel de Novo Paracentric Inversion [inv(20)(q13.1q13.3)] Accompanied by an 11q14.3-q21 Microdeletion in a Pediatric Patient with an Intellectual Disability.

A novel de novo paracentric inversion of the long arm of chromosome 20 [inv(20)(q13.1q13.3)], detected by conventional karyotyping in a 14-year-old boy with mental retardation is described. Further investigation by array comparative genomic hybridization (aCGH) revealed that the 20q inversion was not accompanied by microdeletions/microduplications containing disease-associated genes near or at the breakpoints. Two deletions at chromosomal regions 11q14.3q21 and 20q12 of 4.5 and 1.97 Mb size, respectively, containing important online Mendelian inheritance in man (OMIM) genes, were detected. The 4.5Mb 11q14.3q21 microdeletion was contained within a region that is involved, in most of the reported cases, with the interstitial 11q deletion and may be related to the mental retardation and developmental delay present in the patient. On the other hand, the published data about the 20q12 microdeletion are very few and it is not possible to correlate this finding with our patient's phenotype. This case report contributes to the description of a new chromosomal entity, not previously reported, and is therefore important, especially in prenatal diagnosis and management of patients. Array comparative genomic hybridization has proven a useful technique for detecting submicroscopic rearrangements and should be offered prenatally, especially in cases of de novo karyotypically balanced chromosomal inversions or translocations in order to unveil other unbalanced chromosomal abnormalities such as deletions and amplifications.

Characterization of a Large Novel α-Globin Gene Cluster Deletion Causing α0-Thalassemia in a Chinese Family.

We report a large novel α-globin cluster deletion that we named - -PG (NG_000006.1: g.93628_542759del450131), in a Chinese family. This large deletion is approximately 450 kb long, spanning from upstream of the PolR3k gene at the 5' end to the RAB11FIP3 gene at the 3' end of chromosome 16p13.3. This deletion removes all the globin distal regulatory elements as well as the whole α-globin gene cluster. Patients with heterozygous - -PG/αα had red blood cell (RBC) indices consistent with α-thalassemia (α-thal) trait, but no apparent increase in a cancer tendency or mental disability, microcephaly, relative hypertelorism, unusual facies or genital anomalies.

[Comparative results of preimplantation genetic screening by array comparative genomic hybridization and new-generation sequencing].

Aneuploidies as quantitative chromosome abnormalities are a main cause of failed development of morphologically normal embryos, implantation failures, and early reproductive losses. Preimplantation genetic screening (PGS) allows a preselection of embryos with a normal karyotype, thus increasing the implantation rate and reducing the frequency of early pregnancy loss after IVF. Modern PGS technologies are based on a genome-wide analysis of the embryo. The first pilot study in Russia was performed to assess the possibility of using semiconductor new-generation sequencing (NGS) as a PGS method. NGS data were collected for 38 biopsied embryos and compared with the data from array comparative genomic hybridization (array-CGH). The concordance between the NGS and array-CGH data was 94.8%. Two samples showed the karyotype 47,XXY by array-CGH and a normal karyotype by NGS. The discrepancies may be explained by loss of efficiency of array-CGH amplicon labeling.

Discrepant diagnosis rate of array comparative genomic hybridization in thawed euploid blastocysts.

PURPOSE: Preimplantation genetic screening (PGS) and diagnosis (PGD) with euploid embryo transfer is associated with improved implantation and live birth rates as compared to routine in vitro fertilization. However, misdiagnosis of the embryo is a potential risk. The purpose of this study was to investigate the clinical discrepant diagnosis rate associated with transfer of trophectoderm-biopsied blastocysts deemed to be euploid via array comparative genomic hybridization (aCGH). METHODS: This is a retrospective cohort study including cycles utilizing PGS or PGD with trophectoderm biopsy, aCGH, and euploid embryo transfer at a large university-based fertility center with known birth outcomes from November 2010 through July 2014 (n = 520). RESULTS: There were 520 embryo transfers of 579 euploid embryos as designated by aCGH. Five discrepant diagnoses were identified. Error rate per embryo transfer cycle was 1.0 %, 0.9 % per embryo transferred, and 1.5 % per pregnancy with a sac. The live birth (LB) error rate was 0.7 % (both sex chromosome errors), and the spontaneous abortion (SAB) error rate was 17.6 % (3/17 products of conception tested, but could range from 3/42 to 7/42). No single gene disorders were mistakenly selected for in any known cases.  CONCLUSIONS: Although aCGH has been shown to be a highly sensitive method of comprehensive chromosome screening, several possible sources of error still exist. While the overall error rate is low, these findings have implications for counseling couples that are contemplating PGS and PGD with aCGH.

Craniofacial abnormalities and developmental delay in two families with overlapping 22q12.1 microdeletions involving the MN1 gene.

Deletions spanning the MN1 gene (22q12.1) have recently been proposed as playing a role in craniofacial abnormalities that include cleft palate, as mouse studies have demonstrated that Mn1 haploinsufficiency results in skull abnormalities and secondary cleft palate. We report on four patients (two families) with craniofacial abnormalities and intellectual disability with overlapping microdeletions that span the MN1 gene. Comparative genomic hybridization microarray analysis revealed a 2.76 Mb deletion in the 22q12.1 region, in three family members (Family 1), that contains the MN1 gene. In addition, a complex 22q12 rearrangement, including a 1.61 Mb deletion containing the MN1 gene and a 2.28 Mb deletion encompassing the NF2 gene, has been identified in another unrelated patient (Family 2). Based upon genotype-phenotype correlation among our patients and those previously reported with overlapping 22q12 deletions, we identified a 560 kb critical region containing the MN1 gene that is implicated in human cleft palate formation. Importantly, NF2 was also found within the 22q12 deletion region in several patients which enabled specific clinical management for neurofibromatosis 2.

An economic analysis of prenatal cytogenetic technologies for sonographically detected fetal anomalies.

When congenital anomalies are diagnosed on prenatal ultrasound, the current standard of care is to perform G-banded karyotyping on cultured amniotic cells. Chromosomal microarray (CMA) can detect smaller genomic deletions and duplications than traditional karyotype analysis. CMA is the first-tier test in the postnatal evaluation of children with multiple congenital anomalies. Recent studies have demonstrated the utility of CMA in the prenatal setting and have advocated for widespread implementation of this technology as the preferred test in prenatal diagnosis. However, CMA remains significantly more expensive than karyotype. In this study, we performed an economic analysis of cytogenetic technologies in the prenatal diagnosis of sonographically detected fetal anomalies comparing four strategies: (i) karyotype alone, (ii) CMA alone, (iii) karyotype and CMA, and (iv) karyotype followed by CMA if the karyotype was normal. In a theoretical cohort of 1,000 patients, CMA alone and karyotype followed by CMA if the karyotype was normal identified a similar number of chromosomal abnormalities. In this model, CMA alone was the most cost-effective strategy, although karyotype alone and CMA following a normal karyotype are both acceptable alternatives. This study supports the clinical utility of CMA in the prenatal diagnosis of sonographically detected fetal anomalies.

Absence of skeletal anomalies in siblings with a maternally inherited 12q13.13-q13.2 microdeletion partially involving the HOXC gene cluster.

Microdeletions (12q13.13-q13.2) involving the HOXC gene cluster are rare. Only three patients with this contiguous deletion have been reported, all resulting in phenotypic features that include skeletal anomalies, facial dysmorphism, and intellectual disability. The deletion of the HOXC gene cluster is thought to result in skeletal anomalies in these patients. We report on siblings with a 969 kb deletion in the 12q13.13-q13.2 region detected by array comparative genomic hybridization (aCGH). This deletion spans seven of nine HOXC cluster genes. FISH analysis confirmed the siblings and mother were carriers of the 12q13.13-q13.2 deletion. Although minor facial dysmorphic features were present in both siblings, no skeletal anomalies were present in the siblings or the mother. The proband had autistic-like features and mild developmental delay, while the sibling and mother are of normal intelligence. The absence of skeletal anomalies in our family suggests that deletion of the entire HOXC gene cluster may be required to result in an abnormal skeletal phenotype, or those skeletal anomalies in previously reported patients may be attributed to other genes within the deletion interval.

Cancer Sample Analysis Utilizing Single-Nucleotide Polymorphism Array and Array Comparative Genomic Hybridization.

Chromosomal microarray, including single-nucleotide polymorphism (SNP) array and array comparative genomic hybridization (aCGH), enables the detection of DNA copy-number loss and/or gain associated with unbalanced chromosomal aberrations. In addition, SNP array and aCGH with SNP component also detect copy-neutral loss of heterozygosity (CN-LOH). Here we describe the chromosomal microarray procedure from the sample preparation using extracted DNA to the scanning of the array chip.

A feasible strategy of preimplantation genetic diagnosis for carriers with chromosomal translocation: Using blastocyst biopsy and array comparative genomic hybridization.

BACKGROUND/PURPOSE: Patients with chromosomal translocation are highly vulnerable to produce unbalanced gametes that result in recurrent miscarriages, affected offspring, or infertility. Preimplantation genetic diagnosis (PGD) with blastomere biopsy and fluorescent in-situ hybridization (FISH) has been used to select normal/balanced embryos for transfer. However, FISH is inherent with some technical difficulties such as cell fixation and signal reading. Here we introduce a strategy of PGD using blastocyst biopsy and array comparative genomic hybridization (aCGH) for reproductive problems of patients with chromosomal translocation. METHODS: Twelve patients diagnosed as having chromosomal translocation who underwent PGD cycles were included in this single-center observational study. Blastocyst biopsy was performed and biopsied blastocysts were cryopreserved individually. Testing was performed with aCGH, and the euploid embryos were transferred in the following thawing cycles. RESULTS: The overall diagnostic efficiency was 90.2% (55/61) and the euploidy rate was 32.7% (18/55). Ten cycles of thawed embryo transfer (ET) were carried out, resulting in three live births and another three ongoing pregnancies with an ongoing pregnancy rate of 60%/transfer cycle. The prenatal diagnosis with chorionic villi sampling confirmed the results of PGD/aCGH in all six pregnant women. No miscarriage happened in our case series. CONCLUSION: Our study demonstrates an effective PGD strategy with promising outcomes. Blastocyst biopsy can retrieve more genetic material and may provide more reliable results, and aCGH offers not only detection of chromosomal translocation but also more comprehensive analysis of 24 chromosomes than traditional FISH. More cases are needed to verify our results and this strategy might be considered in general clinical practice.

Cytogenomic characteristics of murine breast cancer cell line JC.

BACKGROUND: Breast cancer (BC), one of the most frequent human tumors, is genetically and histologically heterogeneous. Treatment options can be adapted according to BC subtype. Still, research is necessary to characterize BC biology better and to study potential new treatment options. Murine BC-cell lines can be used as model systems in this respect. RESULTS: Here for the first time murine BC-cell line JC was cytogenomically characterized as being complex rearranged and near-tetraploid. Multicolor banding and array comparative genomic hybridization were applied and the result was in silico translated to the human genome. CONCLUSIONS: Even though being commercially available, cell line JC was yet not much included in BC-research, most likely due to a lack of cytogenomic data. Thus, here comprehensive data is provided on chromosomal aberrations, genomic imbalances and involved breakpoints of JC cell line. Also JC could be characterized as a model for BC of luminal B type, basal-like tumor rather than for luminal A type.