Array Comparative Genomic Hybridization Analysis Research Articles

A Novel 4q32.3 Deletion in a Child: Additional Signs and the Role of MARCH1.

The 4q deletion syndrome is an uncommon condition manifesting with broad clinical expression and phenotypic variability. We report a 5-year-old boy affected by 4q deletion syndrome who showed minor craniofacial features, growth failure, mild developmental delay, severe speech delay, and marked irascibility and aggressivity. Moreover, he showed precocious and crowded primary dentition, digital hyperlaxity, and congenital bilateral adducted thumbs, signs which were previously unreported in the syndrome. The array comparative genomic hybridization analysis revealed a 4q partial terminal deletion of ∼329.6 kb extending from 164.703.186 to 165.032.803 nt, which includes part of MARCH1 (membrane associated ring-CH-type finger 1) gene (OMIM#613331). Same rearrangement was found in his healthy mother. Clinical phenotype of the child and its relationship to the deleted region is presented with a revision of the cases having the same copy number losses from the literature and genomic variant databases.

Application of array comparative genomic hybridization analysis for rapid differential diagnosis of a chromosome euchromatic variant of dup 22q13.3 at amniocentesis

Application of array comparative genomic hybridization analysis for rapid differential diagnosis of a chromosome euchromatic variant of dup 22q13.3 at amniocentesis

Molecular features and vulnerabilities of recurrent chordomas

BackgroundTumor recurrence is one of the major challenges in clinical management of chordoma. Despite R0-resection, approximately 50% of chordomas recur within ten years after initial surgery. The underlying molecular processes are poorly understood resulting in the lack of associated therapeutic options. This is not least due to the absence of appropriate cell culture models of this orphan disease.MethodsThe intra-personal progression model cell lines U-CH11 and U-CH11R were compared using array comparative genomic hybridization, expression arrays, RNA-seq, and immunocytochemistry. Cell line origin was confirmed by short tandem repeat analysis. Inter-personal cell culture models (n = 6) were examined to validate whether the new model is representative. Cell viability after HOX/PBX complex inhibition with small peptides was determined by MTS assays.ResultsUsing whole genome microarray analyses, striking differences in gene expression between primary and recurrent chordomas were identified. These expression differences were confirmed in the world’s first intra-personal model of chordoma relapse consisting of cell lines established from a primary (U-CH11) and the corresponding recurrent tumor (U-CH11R). Array comparative genomic hybridization and RNA-sequencing analyses revealed profound genetic similarities between both cell lines pointing to transcriptomic reprogramming as a key mechanism of chordoma progression. Network analysis of the recurrence specific genes highlighted HOX/PBX signaling as a common dysregulated event. Hence, HOX/PBX complexes were used as so far unknown therapeutic targets in recurrent chordomas. Treating chordoma cell lines with the complex formation inhibiting peptide HXR9 induced cFOS mediated apoptosis in all chordoma cell lines tested. This effect was significantly stronger in cell lines established from chordoma relapses.ConclusionClearly differing gene expression patterns and vulnerabilities to HOX/PBX complex inhibition in highly therapy resistant chordoma relapses were identified using the first intra-personal loco-regional and further inter-personal chordoma progression models. For the first time, HOX/PBX interference was used to induce cell death in chordoma and might serve as the basic concept of an upcoming targeted therapy for chordomas of all progression stages.

The Fanconi Pathway in Pediatric T-Cell Acute Lymphoblastic Leukemia

BRCA2 (also known as FANCD1), a core component of the Fanconi pathway, suppresses transformation of immature T-cell progenitors in mice. However, whether the Fanconi pathway contributes to human T-cell acute lymphoblastic leukemia (T-ALL) pathogenesis is unknown. Using targeted exon sequencing of childhood T-ALL diagnostic specimens, we found heterozygous mutations of Fanconi pathway genes in 23% of 40 cases analyzed. Array comparative genomic hybridization (CGH) analysis revealed that an additional 15% of these cases harbored large heterozygous deletions involving Fanconi pathway genes. The majority of these mutations appear to be somatic although in 2 of 8 available remission samples, the mutations were present, suggestive that these mutations are likely to be germline. Fanconi genes are typically considered two-hit tumor suppressors, where breast or ovarian cancers arising in heterozygous carriers often lose activity of the wild-type allele, leading to genomic instability. However, T-ALL cases with heterozygous Fanconi mutations lacked evidence of increased genomic instability based on array CGH analysis, were not hypersensitive to the PARP inhibitor olaparib, and did not demonstrate increased radial chromosome formation in response to mitomycin C (MMC), indicating that these cases retained activity of the wild-type allele. Complementation studies of Fanconi mutant cells revealed that most of the identified Fanconi mutations encoded pathogenic alleles, as evidenced by their impaired ability to rescue Fanconi-deficient cells from mitomycin C-induced radial chromosome formation and cytotoxicity. Treatment of a patient-derived xenograft sample harboring a heterozygous BRCA2 mutation with mitomycin C revealed that these cells had an intermediate increase in sensitivity to mitomycin C-induced cytotoxicity compared to Fanconi wild-type T-ALLs, suggesting that Fanconi haploinsufficiency is pathogenic in T-ALL. Together, these findings implicate a tumor suppressor role for Fanconi pathway haploinsufficiency in childhood T-ALL. DisclosuresHunger:Jazz Pharmaceuticals: Honoraria; Novartis: Consultancy; Amgen: Consultancy, Equity Ownership; Erytech Pharmaceuticals: Consultancy. Neuberg:Synta Pharmaceuticals: Other: Stock shares.

The Effect of 5-Azacitidine Treatment on the Biologic Characteristics of Bone Marrow Mesenchymal Stem Cells in Patients with Myelodysplastic Syndromes

Myelodysplastic syndromes (MDS) are clonal disorders of the hematopoietic stem cell characterized by ineffective bone marrow (BM) hematopoiesis and increased risk for leukemic evolution. An abnormal BM microenvironment has been reported to contribute to BM failure in MDS through defective support of hematopoiesis. The cellular elements of the BM microenvironment derive from a common progenitor cell, namely the mesenchymal stem cell (MSC). We have previously shown that MSCs from MDS patients display impaired proliferative and clonogenic potential and develop chromosomal alterations during passages.It has been shown that DNA methylation occurs at a high frequency in MDS and that gene reactivation by hypo-methylating agents may have a positive impact on patients' outcome. Recent evidence also suggests altered global DNA methylation status in BM-MSCs of MDS patients. The methyl transferase inhibitor 5-Azacitidine (ΑΖΑ) is widely used for the treatment of MDS; however the effect of AZA on patients' BM microenvironment and specifically on the MSC compartment has not been extensively studied. In the present study we investigate the functional, molecular and cytogenetic characteristics of ex vivo expanded BM-MSCs from MDS patients receiving AZA, before and after 6-cycle therapy.MSCs were isolated and expanded from 14 patients with high risk MDS and 10 age- and sex-matched hematologically healthy subjects undergoing surgery for hip replacement, after informed consent. MSCs were expanded and re-seeded for a total of 5 passages (P) and phenotypically characterized by flow cytometry. MSCs were induced to differentiate to adipocytes and osteoblasts. Differentiation was assessed by cytochemical stains and by the expression of adipocyte- and osteocyte-specific genes. MSC growth characteristics were assessed by the population doubling time (PDT) and the methyl-triazolyl-tetrazolium (MTT) assay throughout passages. The clonogenic potential of MSCs was evaluated by the colony forming unit fibroblast assay. Conventional cytogenetic and array comparative genomic hybridization (aCGH) analyses were performed in cultured expanded MSCs at P2.MSCs from MDS patients before and after therapy, displayed normal spindle-shape morphology and immunophenotypic characteristics as shown by the expression of CD90, CD105, CD44, CD29, CD73, and the lack of expression of CD45, CD14 and CD34. MSCs were also able to differentiate into adipocytes and osteoblasts before and after therapy, as evidenced by the Oil Red O and Alizarin Red staining, respectively, and the expression of differentiation specific genes. Cumulative analysis showed that the proliferation potential of patient MSCs, evaluated by the PDT and MTT assays, was statistically significant decreased compared to healthy individuals (P<0.01) but increased significantly following AZA treatment compared to baseline (P<0.05), although not at the level of the healthy controls. A paired analysis of patient samples before and after therapy, also showed that AZA treatment improves significantly the proliferation rate of MDS-MSCs (P<0.05). Similarly, a cumulative and paired analysis showed that the defective (compared to healthy individuals) clonogenic potential of patient MSCs at P2, increased significantly following AZA treatment (P<0.05 and P<0.05, respectively), although not at the level of the healthy individuals. Karyotypic analysis did not reveal any chromosomal abnormalities in MSC samples, before or after AZA treatment. An aCGH analysis of P2 MSCs has - thus far - been performed in three MDS patients before and after AZA therapy. Results have indicated genetic abnormalities (duplications/deletions) in all three samples before treatment that remained unchanged after AZA therapy whereas some new abnormalities also appeared.The data of this ongoing study on the biological properties of MSCs from MDS patients receiving AZA, show an improvement of the clonogenic and proliferative potential of MSCs following treatment. According to aCGH analysis, the genetic abnormalities of MSCs remain unaffected. The findings of the functional and cytogenetic characteristics of patient MSCs before and after therapy will be individually correlated with the MSC methylation status and with patients' outcome and the results are anticipated to provide novel insights in the role of AZA therapy in the BM microenvironment of MDS patients. DisclosuresNo relevant conflicts of interest to declare.

Retinal Features of Family Members With Familial Exudative Vitreoretinopathy Caused By Mutations in KIF11 Gene.

PurposeTo determine the clinical characteristics of patients and family members with familial exudative vitreoretinopathy (FEVR) caused by mutations in the KIF11 gene.MethodsTwenty-one patients from 10 FEVR families with mutations in the KIF11 gene were studied. The retinal and systemic features were examined. The genetic analyses performed included Sanger sequencing of the KIF11 gene, whole exome sequencing, as well as array comparative genomic hybridization (CGH) analysis and multiple ligation probe assay (MLPA).ResultsSequence analysis revealed seven different KIF11 mutations. Array CGH with MLPA revealed two different exon deletions. All probands had advanced FEVR with retinal detachments (RDs) and microcephaly with or without developmental disabilities. Patients with bilateral RDs were more frequently associated with developmental disabilities (P = 0.023). Multimodal imaging of the family members revealed that six of nine patients without RDs (66%) had varying degrees of chorioretinopathy. The retinal folds in FEVR patients were associated with severe retinal avascularization. However, funduscopic changes in the peripheral retina were unremarkable in family members without RDs. A score representing the peripheral vascular anomalies determined from the fluorescein angiograms was lower than that of control eyes of patients with mutations of the Wnt signaling genes (P = 0.0029).ConclusionsThe probands with KIF11 mutations were associated with severe ocular and systemic pathologies, whereas affected family members showed highly variable clinical manifestations. Peripheral vascular anomalies can often be unremarkable in eyes without RDs.Translational RelevanceThese findings highlight more diverse mechanisms that underlie the pathological changes in patients with FEVR.

Correlating genomic copy number alterations with clinicopathologic findings in 75 cases of hepatocellular carcinoma

BackgroundOligonucleotide array comparative genomic hybridization (aCGH) analysis has been used for detecting somatic copy number alterations (CNAs) in various types of tumors. This study aimed to assess the clinical utility of aCGH for cases of hepatocellular carcinoma (HCC) and to evaluate the correlation between CNAs and clinicopathologic findings.MethodsaCGH was performed on 75 HCC cases with paired DNA samples from tumor and adjacent nontumor tissues. Survival outcomes from these cases were analyzed based on Barcelona-Clinic Liver Cancer Stage (BCLC), Edmondson-Steiner grade (E-S), and recurrence status. Correlation of CNAs with clinicopathologic findings was analyzed by Wilcoxon rank test and clustering vs. K means.ResultsThe survival outcomes indicated that BCLC stages and recurrence status could be predictors and E-S grades could be a modifier for HCC. The most common CNAs involved gains of 1q and 8q and a loss of 16q (50%), losses of 4q and 17p and a gain of 5p (40%), and losses of 8p and 13q (30%). Analyses of genomic profiles and clusters identified that losses of 4q13.2q35.2 and 10q22.3q26.13 seen in cases of stage A, grade III and nonrecurrence were likely correlated with good survival, while loss of 1p36.31p22.1 and gains of 2q11.2q21.2 and 20p13p11.1 seen in cases of stage C, grade III and recurrence were possibly correlated with worst prognosis.ConclusionsThese results indicated that aCGH analysis could be used to detect recurrent CNAs and involved key genes and pathways in patients with HCC. Further analysis on a large case series to validate the correlation of CNAs with clinicopathologic findings of HCC could provide information to interpret CNAs and predict prognosis.

CDK4 overexpression is a predictive biomarker for resistance to conventional chemotherapy in patients with osteosarcoma.

Osteosarcoma (OS) is the most common malignant bone tumor, and its sensitivity to preoperative chemotherapy is a significant prognostic factor. The present study aimed to identify potential genomic markers for the prediction of chemosensitivity in patients with OS using a genomic approach. A total of 50 pediatric and adolescent patients diagnosed with high‑grade OS were selected. Each pre‑therapeutic biopsy sample was subjected to comparative genomic hybridization array analysis and targeted exome sequencing. Although no recurrent gene mutation was observed in chemoresistant tumors, copy number analysis detected recurrent gain of chromosome 12q14.1, which was significantly more frequent (5/21; 24%) in the poor responder cohort than in the good responder cohort (0/29; 0%; P<0.01). Subsequent expression analysis revealed that CDK4 was the only gene in the 12q14.1 gained region with an expression level that was positively associated with copy number gains. In order to elucidate the effect of CDK4 on drug sensitivity, CDK4‑overexpressing OS cell lines were treated with cisplatin (CDDP); significant attenuation of CDDP sensitivity, demonstrated by increased cell viability and decreased expression of cleaved caspase‑9, was induced by enforced expression of CDK4. In addition, treatment with CDK4/6 inhibitor palbociclib in CDK4‑overexpressing U2OS cells facilitated apoptosis and a significant decrease in cell viability in a dose‑dependent manner. In conclusion, the results of the present study showed that higher expression and amplification of CDK4 in tumors is a predictive biomarker for resistance to conventional chemotherapy in patients with OS and that palbociclib is a promising drug for this therapeutically challenging cohort.

Evaluation of extracellular vesicles and gDNA from culture medium as a possible indicator of developmental competence in human embryos.

Human embryos generated in vitro have a high incidence of chromosomal abnormalities that negatively affect pregnancy rate. Embryos generated in vitro secrete extracellular vesicles (EVs) into the culture medium that could be used potentially as indicators of embryo competence. This research aimed to evaluate the concentration and size of EVs and their gDNA content as an indicator of developmental competence in human embryos. Human embryos generated by intracytoplasmic sperm injection (ICSI) were classified morphologically as of either TOP, FAIR or POOR quality. Culture medium and developmentally arrested embryos (which were not able to be used for embryo transfer) were collected. Microvesicles, exosomes (MV/Exo) and apoptotic bodies (ABs) were isolated from culture medium. Nanoparticle tracking analysis (NTA) and array comparative genomic hybridization (aCGH) analysis were performed to evaluate EVs and their gDNA content. From NTA, the diameter (mean) of MVs/Exo from TOP quality embryos was higher (112.17 nm) compared with that of FAIR (108.02) and POOR quality embryos (102.78 nm) (P < 0.05). aCGH analysis indicated that MVs/Exo and ABs carried gDNA with the presence of 23 chromosome pairs. However, when arrested embryos were compared with their respective MVs/Exo and ABs, the latter had an increased rate of chromosomal abnormalities (24.9%) compared with embryos (8.7%) (P < 0.05). In conclusion, the size of EVs from culture medium might be an alternative for evaluating competence of human embryos, however more studies are needed to validate the use of gDNA from EVs as an indicator of embryo competence.

A new childhood ALL case with an extremely complex karyotype and acute spontaneous tumor lysis syndrome

BackgroundB cell precursor acute lymphoblastic leukemia (B-ALL) is the most common malignancy of childhood, with, after corresponding treatment, an overall complete remission rate of 90%. Approximately 75% of B-ALL cases harbor recurrent abnormalities, including so-called complex karyotypes (CK). Tumor lysis syndrome (TLS) is a metabolic abnormality which may arise during cancer therapy and also, extremely rarely, as spontaneous TLS before initiation of chemotherapy in patients with ALL.Case presentationHere we report a 9-year-old male, diagnosed with a de novo pre-B-ALL according to the WHO classification. Cytogenetic, molecular cytogenetic approaches and array comparative genomic hybridization analyses revealed a unique CK involving five chromosomes. It included four yet unreported chromosomal aberrations: a der(11)t(7;11)(p22.1;q24.2), a der(18)t(7;18)(q21.3;p11.22), del(11)(q24.2q25) and dup(18)(q11.1q23). Unfortunately, the patient died 3 months after the initial diagnosis.ConclusionsTo the best of our knowledge, a comparable childhood ALL case was not previously reported. Thus, the combination of the here seen chromosomal aberrations in childhood primary ALL seems to indicate for an extremely adverse prognosis.

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.

Characterization and Association of Marker Chromosomes with Male Infertility

Background and objective A small supernumerary marker chromosome (sSMC) is a rare structurally abnormal chromosome in which no part can be identified by conventional cytogenetic banding technique. There is little known about the association of marker chromosomes with male infertility. We performed a molecular cyto-genetic characterization sSMCs and investigated their association with male infertility. Methods Among 4230 infertile male patients who underwent cytogenetic analyses from January 2008 to December 2018, the records of 15 patients who were diagnosed with sSMCs were reviewed. After ini-tial infertility evaluation, the patients received additional genetic tests including G-bands by trypsin using Giemsa (GTG-banding), Nucleolar organizer region (NOR) banding, Fluorescence in situ hybridization (FISH), and array comparative genomic hybridization (aCGH) analyses to further char-acterize and identify the origin of their marker chromosome. Testis biopsy was performed for the azoospermic patients to evaluate spermatogenic status. Results Among 15 infertile males with sSMCs, eight had nonobstructive azoospermia, five had severe oligozo-ospermia, and two had sperm concentrations above the lower normal limit. The marker chromosomes were identified as Y ring chromosomes (n&nbsp;=&nbsp;8), an isodicentric Y chromosome (n&nbsp;=&nbsp;1), a neocen-tromere Y chromosome (n = 1), a derivative chromosome 1 (n = 1), and an acrocentric short arms (n = 4).&nbsp; Conclusion Marker chromosomes appear to be a rare genetic cause of male infertility. Additional cytogenetic tests, including NOR banding, FISH, and aCGH, could help to characterize the origin of the marker chromosome. Appropriate genetic counseling for couples with infertility caused by a marker chromo-some should be recommended.&nbsp;

Mutation analysis and genomic imbalances of cells found in effusion fluids from patients with ovarian cancer.

Ovarian carcinomas and carcinosarcomas often cause malignant effusions, an accumulation within serous cavities of fluid containing cancer cells. Few studies have focused on the molecular alterations and genetic mechanisms behind effusion formation. The present study investigated the mutation status of TP53, PIK3CA, KRAS, HRAS, NRAS and BRAF in effusion fluids from 103 patients with ovarian cancer. In addition, array Comparative Genomic Hybridization (aCGH) analysis was performed on 20 effusions from patients with high-grade serous carcinoma (10 cases positive for TP53 mutation and 10 with TP53 wild-type). TP53 mutations, two of which were novel: c.826_830delCCTGT and c.475_476GC>TT, were identified in 44% of the cases. Mutations in KRAS, HRAS, and PIK3CA were identified in two, two and four cases, respectively. None of the effusions analysed showed NRAS or BRAF mutations. The aCGH analysis revealed highly imbalanced genomes similar to those described in primary ovarian carcinomas. No specific profile was indicated to distinguish tumors with TP53 mutations from those without. The molecular profiling of cells found in effusion fluids from patients with ovarian cancer thus showed considerable molecular heterogeneity. TP53 seems to be the most frequently mutated gene in these cells and may serve a leading role in the metastatic process.

SAT-LB17 Triplication of SHOX Downstream Region in Mild Short Stature

Deletions of the downstream flanking regions of the SHOX gene that contain conserved non-coding cis-regulatory DNA elements are a known cause of Leri-Weill dyschondrosteosis or idiopathic short stature (ISS). Functional and comparative genomic studies have demonstrated the existence of four CNE enhancers downstream of SHOX which have transcriptional activity, and a fifth has recently been suggested at approximately X:970,000. Duplications of these downstream regions have also been described in patients with ISS, although with a variable phenotype. We present a family with ISS and the first ever reported SHOX downstream triplication. The index case was a 12-year-old Caucasian female of unrelated parents, born with normal weight and length, and normal hearing/vision and development. At the age of 12, her height was -1.87 SD (-1.35 SD adjusted for parental height). Her growth chart showed a height on the 90th centile at birth, slowly crossing centiles for the first 5 years to the 2nd centile, which she then followed. She had normal proportions and head circumference, and no dysmorphic features apart from inverted nipples. Baseline investigations for short stature were normal, including IGF1, karyotype and celiac screen. Bone age was 1.5 year delayed. Family history: arthritis in the father, early menopause in the mother. Analysis of SHOX and its flanking regions was carried out using Multiplex Ligation-dependent Probe Amplification (MLPA) and direct sequencing of all coding exons. MLPA analysis showed a SHOX downstream triplication with a minimum size of 66kb (X:963670-1029779) and a maximum size of 428kb (X:899389-1327688). The size of the duplication was further defined using array comparative genome hybridization (ACGH) analysis which showed the minimum size to be ~325kb (X:907457-1232802). This triplication includes the proposed regulatory element at ~ X:970,000. The triplication was subsequently shown to be present in the proband’s mother and brother who also have mild ISS. Further analysis of these and other family members is ongoing. The triplication is of a single-sized fragment, and the fragment size is identical to one that has been seen in duplicated form in several other patients, increasing the likelihood that the triplicated fragments are in tandem. The precise mechanism of the pathogenic effect of the triplication is unknown, but it likely exerts a negative effect on SHOXtranscription, reducing its expression and ultimately resulting in SHOX haploinsufficiency. In conclusion, we describe a family with mild ISS and a downstream triplication of the proposed X:970,000 SHOX downstream regulatory element. This provides further evidence for this previously proposed regulatory element downstream of SHOX and adds additional proof that increased dosage of this regulatory element region is a cause of short stature.

Twist1 induces chromosomal instability (CIN) in colorectal cancer cells.

Twist1 is a basic helix-loop-helix transcription factor, essential during early development in mammals. While Twist1 induces epithelial-to-mesenchymal transition (EMT), here we show that Twist1 overexpression enhances nuclear and mitotic aberrations. This is accompanied by an increase in whole chromosomal copy number gains and losses, underscoring the role of Twist1 in inducing chromosomal instability (CIN) in colorectal cancer cells. Array comparative genomic hybridization (array CGH) analysis further shows sub-chromosomal deletions, consistent with an increased frequency of DNA double strand breaks (DSBs). Remarkably, Twist1 overexpression downmodulates key cell cycle checkpoint factors—Bub1, BubR1, Mad1 and Mad2—that regulate CIN. Mathematical simulations using the RACIPE tool show a negative correlation of Twist1 with E-cadherin and BubR1. Data analyses of gene expression profiles of patient samples from The Cancer Genome Atlas (TCGA) reveal a positive correlation between Twist1 and mesenchymal genes across cancers, whereas the correlation of TWIST1 with CIN and DSB genes is cancer subtype-specific. Taken together, these studies highlight the mechanistic involvement of Twist1 in the deregulation of factors that maintain genome stability during EMT in colorectal cancer cells. Twist1 overexpression enhances genome instability in the context of EMT that further contributes to cellular heterogeneity. In addition, these studies imply that Twist1 downmodulates nuclear lamins that further alter spatiotemporal organization of the cancer genome and epigenome. Notwithstanding their genetic background, colorectal cancer cells nevertheless maintain their overall ploidy, while the downstream effects of Twist1 enhance CIN and DNA damage enriching for sub-populations of aggressive cancer cells.

Interstitial chromosomal deletion of the tuberous sclerosis complex 2 locus is a signature for radiation-associated renal tumors in Eker rats.

Ionizing radiation can damage DNA and, therefore, is a risk factor for cancer. Eker rats, which carry a heterozygous germline mutation in the tumor‐suppressor gene tuberous sclerosis complex 2 (Tsc2), are susceptible to radiation‐induced renal carcinogenesis. However, the molecular mechanisms involved in Tsc2 inactivation are unclear. We subjected Fischer 344 × Eker (Long Evans Tsc2 +/−) F1 hybrid rats to gamma‐irradiation (2 Gy) at gestational day 19 (GD19) or postnatal day 5 (PND5) and investigated the patterns of genomic alterations in the Tsc2 allele of renal tumors that developed at 1 year after irradiation (N = 24 tumors for GD19, N = 10 for PND5), in comparison with spontaneously developed tumors (N = 8 tumors). Gamma‐irradiation significantly increased the multiplicity of renal tumors. The frequency of LOH at the chromosome 10q12 region, including the Tsc2 locus, was 38%, 29% and 60% in renal carcinomas developed from the nonirradiated, GD19 and PND5 groups, respectively. Array comparative genomic hybridization analysis revealed that the LOH patterns on chromosome 10 in renal carcinomas were classified into chromosomal missegregation, mitotic recombination and chromosomal deletion types. LOH of the interstitial chromosomal deletion type was observed only in radiation‐associated carcinomas. Sequence analysis for the wild‐type Tsc2 allele in the LOH‐negative carcinomas identified deletions (nonirradiated: 26%; GD19: 21%) and base‐substitution mutations (GD19: 4%). Reduced expression of Tsc2 was also observed in the majority of the LOH‐negative carcinomas. Our results suggest that interstitial chromosomal deletion is a characteristic mutagenic event caused by ionizing radiation, and it may contribute to the assessment of radiation‐induced cancer risk.

Iodine‑131 metabolic radiotherapy leads to cell death and genomic alterations through NIS overexpression on cholangiocarcinoma.

Cholangiocarcinoma (CC) is an aggressive liver tumor with limited therapeutic options. Natrium-iodide symporter (NIS) mediates the uptake of iodine by the thyroid, representing a key component in metabolic radiotherapy using iodine-131 (131I) for the treatment of thyroid cancer. NIS expression is increased in CC, providing the opportunity for a novel therapeutic approach for this type of tumor. Thus, in this study, we aimed to evaluate therapeutic efficacy of 131I in two human CC cell lines. Uptake experiments analyzed the 131I uptake profiles of the tumor cell lines under study. The cells were irradiated with various doses of 131I to evaluate and characterize the effects of metabolic radiotherapy. NIS protein expression was assessed by immunofluorescence methods. Cell survival was evaluated by clonogenic assay and flow cytometry was used to assess cell viability, and the type of death and alterations in the cell cycle. The genomic and epigenetic characterization of both CC cells was performed before and after irradiation. NIS gene expression was evaluated in the CC cells by RT-qPCR. The results revealed that CC cells had a higher expression of NIS. 131I induced a decrease in cell survival in a dose-dependent manner. With the increasing irradiation dose, a decrease in cell viability was observed, with a consequent increase in cell death by initial apoptosis. Karyotype and array comparative genomic hybridization (aCGH) analyses revealed that both CC cell lines were near-triploid with several numerical and structural chromosomal rearrangements. NIS gene expression was increased in the TFK-1 and HuCCT1 cells in a time-dependent manner. On the whole, the findings of this study demonstrate that the presence of NIS in cholangiocarcinoma cell lines is crucial for the decreased cell viability and survival observed following the exposure of cholangiocarcinoma cells to 131I.

Identification of PCDH19 Gene Mutations/Deletions in Patients with Early Onset Epilepsy.

Background and Aims:PCDH19 gene, which encodes protocadherin 19, is associated with epilepsy and intellectual disability, mainly in affected females. The clinical manifestations are heterogeneous and the main features include early onset seizure, generalized or focal seizures sensitive to fever, and brief seizures occurring in clusters. The disorders exhibit a unique and unusual X-linked pattern of expression. We aimed to investigate PCDH19 mutations/deletions in patients with epilepsy and describe the clinical/molecular features.Methods:PCDH19 gene was analyzed in 35 Turkish female patients from 34 families with early-onset epilepsy via direct sequencing and multiplex ligation-dependent probe amplification analysis. Additionally, array comparative genomic hybridization analysis was performed in patients with whole gene deletion.Results:We identified 2 different heterozygous mutations in 2 unrelated probands (5. 7%) which were located in exon 1. Additionally, whole gene deletions were detected in dizygotic twin girls (5. 7%), who had distinct clinical features and the deletion was inherited from the unaffected father. The second twin suffered more severe clinical manifestations including autistic features, behavioral problems, mild-moderate mental retardation and seizures, which were under control with multidrug regimen when compared with the first twin.Conclusion:PCDH19 is a major causative gene in patients with epilepsy and further data is required to gain a better understanding of phenotype-genotype correlation. In addition to gene sequencing, deletion/duplication analysis will improve the molecular diagnosis in patients with clinical findings.

Case report of 49, XXXXY syndrome: first case in Oman

Background: The incidence of sex chromosome aneuploidies is 1:400. Klinefelter syndrome is considered to be the most common type of sex chromosome aneuploidy, manifested as variants, such as 48,XXXY, 48,XXYY and 49,XXXXY. Nondisjunction of the X chromosome during meiosis I and II is considered as the cause of this type of aneuploidy. The classic clinical presentation of Klinefelter Syndrome is a triad of hypogonadism, mental retardation, and musculoskeletal anomalies. Case Presentation: In this case report, we describe a 2-year-old male child identified postnatally to have low birth weight, congenital heart defect, inguinal hernia, facial dysmorphism, and genital organs anomalies. Chromosomal analysis revealed a karyotype of 49,XXXXY and the comparative genomic hybridization (CGH) array analysis revealed a 155,065 kilo base pair duplication on chromosome Xp22.33q28 (168,546, −155,233,731) X4. Conclusion: 49,XXXXY karyotype is considered as the rarest sex chromosome aneuploidy syndrome. To our knowledge, this study is the first report of a patient with 49,XXXXY syndrome from Oman.

A case of de novo microdeletion with combination of 1q21.1 and 14q32.2q32.31

Background: Chromosomal microarray is considered as the first-line diagnostic genetic test in all individuals with intellectual disability (ID) and attention-deficit disorders. In recent years, the use of chromosomal microarrays routinely for this purpose has resulted in the identification of many new microdeletion and microduplication regions connected with these clinical situations, including the 1q21.1 and 14q32.2q32.31 microdeletions. Case Presentation: A 5-year-old male patient came to the clinic because of ID, hyperactivity, growth retardation, and speaking difficulty. We determined strabismus on both the eyes, and he was myopic. He had a high palate, little, and sparse teeth. On the right hand, there was a simian line. Both undescended testes were brought down with surgery. In addition, he had got an inward penis head. He had joint laxity in most of the joints. He had pes planus and talipes valgus. Therefore, we decided to make array-comparative genomic hybridization analysis and the result came 1368.001 kb deletion on 1q21.1 between chr1: 146023922 and 147391923 nucleotides and 992.003 kb deletion on 14q32.2q32.31 between chr14: 100453009 and 101445012 nucleotides according to �Human Genome Build 37� (The result was confirmed by a fluorescent in situ hybridization method performed to determine the particular deleted regions). Conclusion: Here, we report the first case presented with ID, hyperactivity, growth retardation, and speaking difficulty with other findings and has a combination of de novo 1q21.1 and 14q32.2q32.31 microdeletions. Although several research groups have reported similar results with similar regions separately, this study is the first of its kind revealing the effects of this combination to clinical outcome.