Comparative Genomic Hybridization Results Research Articles

Predictor(s) of Abnormal Array Comparative Genomic Hybridization Results in Patients With Cleft Lip and/or Palate.

Cleft lip and/or cleft palate (CL/P) occurs either as an isolated anomaly or as one manifestation of genetic syndromes. Chromosomal abnormalities from karyotype analysis are commonly seen in cases of nonisolated CL/P. This study was designed to evaluate the usefulness of clinical array comparative genomic hybridization (aCGH) testing in patients with CL/P. Our objectives were to identify the clinical phenotypes that are predicative of an abnormal aCGH result, correlate aCGH results with language outcome, and analyze the data in the abnormal aCGH results group. Nonisolated CL/P patients who had clinical aCGH testing performed between 2009 and 2012 in the University of Alabama at Birmingham cytogenetics lab were enrolled. The demographic data, clinical phenotypes, and speech outcome were collected. Two hundred forty-five nonisolated CL/P patients were studied, with 62 having an abnormal aCGH result compared to 183 patients with a normal aCGH result. The presence of developmental delay/intellectual disability (DD/ID), dysmorphic features, congenital anomalies, and/or family history of DD/ID were significantly higher in the abnormal aCGH group (P < .05). Neither the aCGH results nor the type of CL/P correlated with speech outcome. Finally, analysis of the abnormal aCGH result group revealed that DD/ID had a strong positive association with the copy number variation pathogenicity and the number of genes involved. This study demonstrated the diagnostic value of clinical aCGH testing in CL/P patients who present with DD/ID, dysmorphic features, other congenital anomalies, and/or family history of DD/ID.

Solitary fibrous tumors: loss of chimeric protein expression and genomic instability mark dedifferentiation

Solitary fibrous tumors, which are characterized by their broad morphological spectrum and unpredictable behavior, are rare mesenchymal neoplasias that are currently divided into three main variants that have the NAB2-STAT6 gene fusion as their unifying molecular lesion: usual, malignant and dedifferentiated solitary fibrous tumors. The aims of this study were to validate molecular and immunohistochemical/biochemical approaches to diagnose the range of solitary fibrous tumors by focusing on the dedifferentiated variant, and to reveal the genetic events associated with dedifferentiation by integrating the findings of array comparative genomic hybridization. We studied 29 usual, malignant and dedifferentiated solitary fibrous tumors from 24 patients (including paired samples from five patients whose tumors progressed to the dedifferentiated form) by means of STAT6 immunohistochemistry and (when frozen material was available) reverse-transcriptase polymerase chain reaction and biochemistry. In addition, the array comparative genomic hybridization findings were used to profile 12 tumors from nine patients. The NAB2/STAT6 fusion was detected in all of the tumors, but immunohistochemistry and western blotting indicated that chimeric protein expression was atypical or absent in 9 out of 11 dedifferentiated tumors. The comparative genomic hybridization results revealed that the usual and malignant solitary fibrous tumors had a simple profile, whereas the genome of the dedifferentiated tumors was complex and unstable, and suggested that 13q and 17p deletions and TP53 mutations may be present in malignant lesions before the full expression of a dedifferentiated phenotype. Solitary fibrous tumor dedifferentiation is associated with the loss of chimeric oncoprotein expression, genomic instability, and cell decommitment and reprogramming. The assessment of dedifferentiated solitary fibrous tumors is based on the presence of the fusion transcripts and, in principle, negative STAT6 immunohistochemistry should not rule out a diagnosis of solitary fibrous tumor.

Cryptic insertion of 3′FOXO1 into inverted chromosome arm 2q in the presence of two normal chromosome 13s and 13 small interstitial duplications in a patient with alveolar rhabdomyosarcoma

Alveolar rhabdomyosarcoma (ARMS) is a pediatric soft tissue neoplasm with a characteristic translocation, t(2;13)(q35;q14), which is detected in 70-80% of cases. This well-described translocation produces the gene fusion product PAX3-FOXO1. Cryptic rearrangements of this fusion have never before been reported in ARMS. Here we describe a patient with ARMS that showed, by fluorescence in situ hybridization and G-banded chromosomes, a cryptic insertion of 3'FOXO1 into inverted chromosome 2q. The inversion breakpoints were depicted by array comparative genomic hybridization as two small interstitial duplications, one of which involved the PAX3 gene. In addition, the array comparative genomic hybridization results revealed 1q gain, 16q loss, and 11 more small duplications, with one of them involving the FOXO1 gene. Although the pathogenesis in classic ARMS cases is thought to be driven by the 5'PAX3-3'FOXO1 fusion on derivative chromosome 13, here we report a novel cryptic insertion of 3'FOXO1 resulting in a pathogenic fusion with 5'PAX3 on inverted chromosome 2q.

Chromosomal meiotic segregation, embryonic developmental kinetics and DNA (hydroxy)methylation analysis consolidate the safety of human oocyte vitrification.

Oocyte vitrification has been introduced into clinical settings without extensive pre-clinical safety testing. In this study, we analysed major safety aspects of human oocyte vitrification in a high security closed system: (i) chromosomal meiotic segregation, (ii) embryonic developmental kinetics and (iii) DNA (hydroxy)methylation status. Fresh and vitrified sibling oocytes from young donors after intracytoplasmic sperm injection (ICSI) were compared in three different assays. Firstly, the chromosomal constitution of the fertilized zygotes was deduced from array comparative genomic hybridization results obtained from both polar bodies biopsied at Day 1. Secondly, embryo development up to Day 3 was analysed by time-lapse imaging. Ten specific time points, six morphokinetic time intervals and the average cell number on Day 3 were recorded. Thirdly, global DNA methylation and hydroxymethylation patterns were analysed by immunostaining on Day 3 embryos. The nuclear fluorescence intensity was measured by Volocity imaging software. Comprehensive chromosomal screening of the polar bodies demonstrated that at least half of the zygotes obtained after ICSI of fresh and vitrified oocytes were euploid. Time-lapse analysis showed that there was no significant difference in cleavage timings, the predictive morphokinetic time intervals nor the average cell number between embryos developed from fresh and vitrified oocytes. Finally, global DNA (hydroxy)methylation patterns were not significantly different between Day 3 embryos obtained from fresh and from vitrified oocytes. Our data further consolidate the safety of the oocyte vitrification technique. Nevertheless, additional testing in young and older sub-fertile/infertile patients and sound follow-up studies of children born after oocyte cryopreservation remain mandatory.

Validation of multiple annealing and looping-based amplification cycle sequencing for 24-chromosome aneuploidy screening of cleavage-stage embryos

To validate multiple annealing and looping-based amplification cycle (MALBAC) sequencing for 24-chromosome aneuploidy screening of cleavage embryos and to explore the chromosomal characteristics of embryos at the cleavage stage. The 24-chromosome aneuploidy analyses of the blastomeres included comparative genomic hybridization (CGH), single nucleotide polymorphism (SNP), and MALBAC sequencing. University-affiliated IVF center. Three couples who delivered babies from the same IVF cycle, which included 23 donated, frozen cleavage embryos. None. Three blastomeres were selected from each single embryo and subject to CGH, SNP, and MALBAC sequencing for 24-chromosome aneuploidy, respectively. The results of MALBAC sequencing were compared with the results of CGH and SNP. The chromosomal status and occurrence of the abnormal chromosomes were investigated. The relationship between the embryos' morphology and the euploid state was analyzed. Among the 23 donated embryos, the MALBAC sequencing results of 18 (78.26%) embryos were identical to those of CGH or SNP, including 8 embryos that had identical results by all three techniques. In terms of euploidy, only 6 of these 23 embryos (26.09%) were diploid. Blastomere abnormality was observed in all autosomes and sex chromosomes. In addition, the frequency of abnormality was different for certain chromosomes. With sequencing at 0.04× genome depth, MALBAC sequencing has been validated as a satisfactory method for 24-chromosome aneuploidy screening. The proportion of abnormal chromosomes was high in cleavage-stage embryos, and any chromosome could be abnormal.

DNA copy number alterations in pleomorphic leiomyosarcoma: A case report.

Pleomorphic leiomyosarcoma (P-LMS) is a rare morphological variant of LMS. The current study presents the cytogenetic data of a P-LMS that arose in the axillary region of a 31-year-old male. The results of array-based comparative genomic hybridization for the primary tumor showed DNA copy number alteration (DCNA) gains of 8ptel, 17ptel and 17q11.2 and losses of 2ptel, 7ptel, 7qtel, 10p15, 12p12-13.1, 13q14.2-14.3, 15q25-26 and Yq11. However, a metastatic lesion showed cytogenetic data different from the primary tumor DCNAs, with only the locus of 17ptel (282M15/SP6) in common between them. These observations add to the spectrum of DCNAs that have been reported in previous cases of LMS and provide novel cytogenetic data.

MP23-17 BELLINI DUCT CARCINOMAS HAVE A DIFFERENT GENETIC SIGNATURE COMPARED TO UROTHELIAL CARCINOMAS OF THE UPPER URINARY TRACT

You have accessJournal of UrologyKidney Cancer: Basic Research I1 Apr 2014MP23-17 BELLINI DUCT CARCINOMAS HAVE A DIFFERENT GENETIC SIGNATURE COMPARED TO UROTHELIAL CARCINOMAS OF THE UPPER URINARY TRACT Volker Jung, Frank Becker, Martin Parr, Arndt Hartmann, Susanne Füssel, Rainer Grobholz, Marieta Toma, Bernd Wullich, Arne Strauss, Carl Ludwig Behnes, Wolfgang Otto, Michael Stöckle, and Kerstin Junker Volker JungVolker Jung More articles by this author , Frank BeckerFrank Becker More articles by this author , Martin ParrMartin Parr More articles by this author , Arndt HartmannArndt Hartmann More articles by this author , Susanne FüsselSusanne Füssel More articles by this author , Rainer GrobholzRainer Grobholz More articles by this author , Marieta TomaMarieta Toma More articles by this author , Bernd WullichBernd Wullich More articles by this author , Arne StraussArne Strauss More articles by this author , Carl Ludwig BehnesCarl Ludwig Behnes More articles by this author , Wolfgang OttoWolfgang Otto More articles by this author , Michael StöckleMichael Stöckle More articles by this author , and Kerstin JunkerKerstin Junker More articles by this author View All Author Informationhttps://doi.org/10.1016/j.juro.2014.02.885AboutPDF ToolsAdd to favoritesDownload CitationsTrack CitationsPermissionsReprints ShareFacebookTwitterLinked InEmail INTRODUCTION AND OBJECTIVES Bellini duct carcinoma or collecting duct carcinoma (CDC) is a rare renal neoplasm that is associated with poor prognosis given its highly aggressive course and limited response to immuno- or chemotherapy. Histologically, CDC is defined as a subtype of renal cell carcinomas, but in some cases, it is difficult to differentiate from urothelial carcinomas (UC). Therefore the aim of this study was to determine genetic alterations of CDC in comparison to that of urothelial carcinomas of the upper urinary tract (UUT-UC) to clarify the histological origin of this rare tumor entity. METHODS Twenty-nine CDC samples were obtained from seven different German centers and compared with twenty-six urothelial carcinomas of the upper urinary tract. Comparative genomic hybridization (CGH) was used to investigate the genetic composition of patients’ tumors and allowed the detection of losses and gains of DNA copy numbers throughout the entire genome. The clinical data were correlated with CGH results. RESULTS CGH analysis of CDC revealed DNA aberrations in many chromosomes. DNA losses were more frequently observed than gains, while high-level amplifications were not detected. The mean frequency of CDC chromosomal aberrations (4.9/case) was slightly lower than that in UUT-UC (5.4/case). Recurrent CDC DNA losses occurred at 8p (n=9/29), 16p (9/29), 1p (n=7/29) and 9p (n=7/29), and gains occurred in 13q (n=9/29). In contrast to CDC, the most frequently detected UUT-UC DNA aberration was a loss at 9q (n=13/26). DNA losses at 9q, 13q and 8q as well as gains at 8p showed significant variations in UUT-UC compared to CDC. There was no correlation between the patients’ clinical course and the presence or absence of these recurrent genetic alterations. CONCLUSIONS For the first time we have shown that DBCs are characterized by a different genetic pattern compared to UUT urothelial carcinomas. Regarding also the published data on renal cell carcinoma, we conclude that CDC seems to be an unique entity of kidney carcinomas. © 2014FiguresReferencesRelatedDetails Volume 191Issue 4SApril 2014Page: e249-e250 Peer Review Report Advertisement Copyright & Permissions© 2014MetricsAuthor Information Volker Jung More articles by this author Frank Becker More articles by this author Martin Parr More articles by this author Arndt Hartmann More articles by this author Susanne Füssel More articles by this author Rainer Grobholz More articles by this author Marieta Toma More articles by this author Bernd Wullich More articles by this author Arne Strauss More articles by this author Carl Ludwig Behnes More articles by this author Wolfgang Otto More articles by this author Michael Stöckle More articles by this author Kerstin Junker More articles by this author Expand All Advertisement Advertisement PDF downloadLoading ...

Collecting Duct Carcinomas Represent a Unique Tumor Entity Based on Genetic Alterations

Collecting duct carcinoma (CDC) is a rare renal neoplasm that is associated with poor prognosis due to its highly aggressive course and limited response to immuno- or chemotherapy. Histologically, CDC is defined as a subtype of renal cell carcinomas, but in some cases, it is difficult to differentiate from urothelial carcinomas (UC). Therefore the aim of this study was to determine genetic alterations of CDC in comparison to that of urothelial carcinomas of the upper urinary tract (UUT-UC) to clarify the histological origin of this rare tumor entity. Twenty-nine CDC samples were obtained from seven different German centers and compared with twenty-six urothelial carcinomas of the upper urinary tract. Comparative genomic hybridization (CGH) was used to investigate the genetic composition of patients’ tumors and allowed the detection of losses and gains of DNA copy numbers throughout the entire genome. The clinical data were correlated with CGH results. CGH analysis of CDC revealed DNA aberrations in many chromosomes. DNA losses were more frequently observed than gains, while high-level amplifications were not detected. The mean frequency of CDC chromosomal aberrations (4.9/case) was slightly lower than that in UUT-UC (5.4/case). Recurrent CDC DNA losses occurred at 8p (n=9/29), 16p (9/29), 1p (n=7/29) and 9p (n=7/29), and gains occurred in 13q (n=9/29). In contrast to CDC, the most frequently detected UUT-UC DNA aberration was a loss at 9q (n=13/26). DNA losses at 9q, 13q and 8q as well as gains at 8p showed significant variations in UUT-UC compared to CDC. There was no correlation between the patients’ clinical course and the presence or absence of these recurrent genetic alterations. CDCs are characterized by a different genetic pattern compared to UUT-UC. Regarding the published data on renal cell carcinoma, we conclude that CDC appears to be a unique entity among kidney carcinomas.

Generation and application of dynamic standard reference intervals for analyzing results of comparative genomic hybridization

Generation and application of dynamic standard reference intervals for analyzing results of comparative genomic hybridization

Saethre–Chotzen syndrome with an atypical phenotype: identification of TWIST microdeletion by array CGH

Saethre–Chotzen syndrome is a very rare autosomal dominant congenital disorder characterized by craniosynostosis and acrocephalosyndactyly. It is caused by a mutation in TWIST1, located on chromosome 7p21. A shortage of functional TWIST1 protein affects the development and maturation of cells in the skull, face, and limbs. The patient described in this report displayed craniofacial features classic for Saethre–Chotzen syndrome, including craniosynostosis, low-set ears, small pinna with prominent crura, a high-arched palate, and a simian crease on the left hand. He did not have the limb anomalies commonly seen in patients with Saethre–Chotzen syndrome, and the results of conventional chromosome analysis were normal. However, results of a microarray-based comparative genomic hybridization (array CGH) study confirmed the karyotype of46,XY.7p21.1p15.3(15,957,375-20,331,837)x1, a region that includes TWIST1. Subsequent fluorescent in situ hybridization analysis confirmed this result. No other chromosome was involved in the rearrangement. This case illustrates the important contribution of array CGH to the identification of TWIST microdeletions, even in a patient not showing the phenotype typical of Saethre–Chotzen syndrome.

Abstract 4692: ARL4C, a novel ovarian cancer-related gene, its down-regulated predicts poor survival in ovarian cancer.

Abstract Purpose: To identify the genes involved in ovarian cancer progression and elucidate their biological actions in ovarian cancer. Experimental Design: Comparative genomic hybridization (CGH) was employed to screen the chromosome fragments with different copy numbers between the ovarian cancer cell line HO-8910 and its highly metastatic cell subline HO-8910PM. Fluorescent in situ hybridization (FISH) via biotin-labeled BAC clone probes was used to validate the results of CGH. Using gene expression microarray, differentially expressed genes on the target chromosome fragment were compared between the two cell lines. Lentiviral-mediated transfection was used to investigate the effects of candidate gene over-expression or knockdown on colony formation, proliferation and motility of ovarian cancer cells. Two hundred twelve epithelial ovarian cancer samples were analyzed for mRNA expression of candidate gene using RT-PCR. Associations of the gene expression with clinicopathological factors, progression free and overall survival were evaluated. Results: Chromosome fragment 2q36.1-37.3 was found hemizygous deletion in HO-8910PM cells by CGH and FISH. Comparing gene expression data, we identified ARL4C, located on chromosome 2q37.1, to be down-regulated in the HO-8910PM cells, compared with the HO-8910 cells. Western blot showed ARL4C expression was low/null in most ovarian cancer cell lines, including the HO-8910PM cells. ARL4C over-expression or knockdown had no obvious effect on colony formation and cell proliferation, but negatively regulated cell motility. ARL4C over-expression suppressed ovarian cancer cells migration and invasion. Inversely, ARL4C knockdown promoted ovarian cancer cells migration. Clinicopathologic studies further supported the relevance of ARL4C in ovarian cancers. Patients with medium or high levels of ARL4C mRNA were at lower risk for death, compared to those with low levels (HR, 0.45; 95% C.I., 0.27-0.76; P=0.003 for mid levels; and HR, 0.58; 95% C.I., 0.35-0.96; P = 0.035 for high levels) after adjusting for age, disease stage, tumor grade, histologic type and residual tumor size. Kaplan-Meier survival analysis demonstrated similar results. Conclusions: ARL4C is a novel ovarian cancer related gene that inhibits cell motility. It seems to play a protective role in cancer progression and is an independent prognostic factor for patient with ovarian cancer. More studies are needed to further elucidate the molecular mechanisms of ARL4C in ovarian cancer progression. Citation Format: Dan Su, Shenhau Xu, Haiyan Xu, Yun Gao, Jianguo Feng, Min Luo, Lisha Ying, Nengming Lin, Dionyssios Katsaros, Herbert Yu. ARL4C, a novel ovarian cancer-related gene, its down-regulated predicts poor survival in ovarian cancer. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 4692. doi:10.1158/1538-7445.AM2013-4692

Generation and application of dynamic standard reference intervals for analyzing results of comparative genomic hybridization

The present work was aimed at generating the dynamic standard reference intervals (DSRI) and their application for chromosomal-aberration (CA) analysis. The evaluation of the generated DSRI was performed using the DNA samples from four patients with already known CA. High-resolution comparative genomic hybridization analysis (HR-CGH) allowed us to not only identify all of the CAs, that were not revealed by CGH, but also to detect the breakpoints and to determine the size of chromosomal imbalance.

Localization of centromeric breaks in head and neck squamous cell carcinoma

Head and neck squamous cell carcinoma (HNSCC) have very complex karyotypes that show all types of structural rearrangements. The most frequent aberrations are whole-arm translocations, which appear to have their breakpoints in centromeric or pericentromeric regions. We aimed to pinpoint the exact location of the breakpoints of these marker chromosomes with high-resolution cytogenetic and genetic analyses using microarray comparative genomic hybridization (CGH), multiplex ligation-dependent probe amplification (MLPA), and fiber fluorescence in situ hybridization (FISH). Among the seven cell lines in this study, six (84%) harbored one or more centromeric breakpoints or whole-arm translocations. In total, microarray CGH identified 163 breakpoints, 47 (29%) of which were in centromeric regions. Microarray CGH and MLPA results indicated that the translocation breakpoints were localized between the microarray oligonucleotide clones and MLPA probes closest to the centromere. High-resolution fiber-FISH revealed adjacent or minimally overlapping signals of probes that recognize the pericentromeric sequences of the two participating chromosomes. This indicates that whole chromosome arm translocation breakpoints occur within the pericentromeric chromatin and not the centromere core sequences.

Defining the impact of maternal cell contamination on the interpretation of prenatal microarray analysis

PurposeTo understand the ability of microarray-based comparative genomic hybridization to detect copy-number variation in the presence of maternal cell contamination. MethodsTo simulate maternal cell contamination, normal female DNA was mixed at various levels with DNA carrying known copy-number variations. Mixtures were run on a whole-genome 135K oligonucleotide-based array. Data were analyzed with custom analysis software. ResultsThe array and software design allowed detection of larger copy-number variations at higher levels of maternal cell contamination than smaller copy-number variations. The smallest duplications and deletions were obscured at 22–31% and 55–58% maternal cell contamination, respectively. With male fetal samples, the sex chromosome ratios started showing observable shifts at ~10% maternal cell contamination. ConclusionAs knowledge of the maternal cell contamination level aids in interpretation of array results, we recommend concurrent, independent maternal cell contamination studies for all fetal samples for accurate and timely results. With male fetal samples in our laboratory, interfering levels of maternal cell contamination can be excluded when the sex chromosome plots appear normal. Thus, reportable male microarray-based comparative genomic hybridization results may be occasionally achieved without maternal cell contamination studies. Because the effects of maternal cell contamination on microarray results are dependent on array platforms, experimental techniques, and software algorithms, each laboratory should perform its own analysis to determine acceptable levels of maternal cell contamination for its assays.Genet Med 2012:14(11):914–921

Comparative Genomic Hybridization Analysis Shows Different Epidemiology of Chromosomal and Plasmid-Borne cpe-Carrying Clostridium perfringens Type A

Clostridium perfringens, one of the most common causes of food poisonings, can carry the enterotoxin gene, cpe, in its chromosome or on a plasmid. C. perfringens food poisonings are more frequently caused by the chromosomal cpe-carrying strains, while the plasmid-borne cpe-positive genotypes are more commonly found in the human feces and environmental samples. Different tolerance to food processing conditions by the plasmid-borne and chromosomal cpe-carrying strains has been reported, but the reservoirs and contamination routes of enterotoxin-producing C. perfringens remain unknown. A comparative genomic hybridization (CGH) analysis with a DNA microarray based on three C. perfringens type A genomes was conducted to shed light on the epidemiology of C. perfringens food poisonings caused by plasmid-borne and chromosomal cpe-carrying strains by comparing chromosomal and plasmid-borne cpe-positive and cpe-negative C. perfringens isolates from human, animal, environmental, and food samples. The chromosomal and plasmid-borne cpe-positive C. perfringens genotypes formed two distinct clusters. Variable genes were involved with myo-inositol, ethanolamine and cellobiose metabolism, suggesting a new epidemiological model for C. perfringens food poisonings. The CGH results were complemented with growth studies, which demonstrated different myo-inositol, ethanolamine, and cellobiose metabolism between the chromosomal and plasmid-borne cpe-carrying strains. These findings support a ubiquitous occurrence of the plasmid-borne cpe-positive strains and their adaptation to the mammalian intestine, whereas the chromosomal cpe-positive strains appear to have a narrow niche in environments containing degrading plant material. Thus the epidemiology of the food poisonings caused by two populations appears different, the plasmid-borne cpe-positive strains probably contaminating foods via humans and the chromosomal strains being connected to plant material.

Results of array comparative genomic hybridization (aCGH) combined with multiplex PCR for preimplantation genetic diagnosis (PGD) of both single gene disorders and aneuploidy

Results of array comparative genomic hybridization (aCGH) combined with multiplex PCR for preimplantation genetic diagnosis (PGD) of both single gene disorders and aneuploidy

A Linear Karyotypic Association between PB-I, PB-II and Blastomere Using Sequentially Performed Comparative Genome Hybridization with No Association Established between Karyotype, Morphologic, Biochemical (sHLA-G Expression) Characteristics, Blastocyst Formation and Subsequent Pregnancy Outcome

Background: The importance of oocyte/embryo ploidy to achieve implantation and a subsequent pregnancy. Aim: To correlate first and second polar bodies and day-3 blastomere ploidy, embryo morphology and biochemical (sHLA-G) characteristics with blastocyst development and in vitro pregnancy outcome. Materials and Methods: All oocytes/zygotes and embryos were individually cultured to the blastocyst stage. PB-I, PB-II and blastomeres underwent complete karyotyping and sHLA-G expression was measured on day 2. Results: 57 mature (MII) donor oocytes were obtained, 33/57 (57.9%) were aneuploid, 21/57 (36.8%) were euploid, and 3/57 (5%) were ‘inconclusive’. No correlation was found between comparative genomic hybridization (CGH) status of PB-I, PB-II and the graduated embryo score. Furthermore, no correlation was established between PB-I CGH results and blastocyst morphology grade. There was a significant correlation between PB-I CGH and blastomere CGH results. Euploid and aneuploid PB-I developed into 58 and 67% blastocysts, respectively. ĸ statistics (>0.7) revealed a positive correlation between the ploidy of PB-I, PB-II and the blastomeres. Conclusion: Following ICSI and sequential genetic karyotyping of the oocyte/zygote and subsequent blastomeres, the majority of oocytes fertilized and subsequent zygotes developed into blastocysts, despite their ploidy status. We therefore conclude that blastocyst development is not associated with ploidy.

Marked genetic differences between BRAF and NRAS mutated primary melanomas as revealed by array comparative genomic hybridization

Somatic mutations of BRAF and NRAS oncogenes are thought to be among the first steps in melanoma initiation, but these mutations alone are insufficient to cause tumor progression. Our group studied the distinct genomic imbalances of primary melanomas harboring different BRAF or NRAS genotypes. We also aimed to highlight regions of change commonly seen together in different melanoma subgroups. Array comparative genomic hybridization was performed to assess copy number changes in 47 primary melanomas. BRAF and NRAS were screened for mutations by melting curve analysis. Reverse transcription PCR and fluorescence in-situ hybridization were performed to confirm the array comparative genomic hybridization results. Pairwise comparisons revealed distinct genomic profiles between melanomas harboring different mutations. Primary melanomas with the BRAF mutation exhibited more frequent losses on 10q23-q26 and gains on chromosome 7 and 1q23-q25 compared with melanomas with the NRAS mutation. Loss on the 11q23-q25 sequence was found mainly in conjunction with the NRAS mutation. Primary melanomas without the BRAF or the NRAS mutation showed frequent alterations in chromosomes 17 and 4. Correlation analysis revealed chromosomal alterations that coexist more often in these tumor subgroups. To find classifiers for BRAF mutation, random forest analysis was used. Fifteen candidates emerged with 87% prediction accuracy. Signaling interactions between the EGF/MAPK-JAK pathways were observed to be extensively altered in melanomas with the BRAF mutation. We found marked differences in the genetic pattern of the BRAF and NRAS mutated melanoma subgroups that might suggest that these mutations contribute to malignant melanoma in conjunction with distinct cooperating oncogenic events.

Supernumerary marker chromosomes derived from chromosome 6: Cytogenetic, molecular cytogenetic, and array CGH characterization

Supernumerary marker chromosomes (SMC) are relatively common in prenatal diagnosis. As the clinical outcomes vary greatly, a better understanding of the karyotype-phenotype correlation for different SMCs will be important for genetic counseling. We present two cases of prenatally detected de novo, small SMCs. The markers were present in 80% of amniocyte colonies in Case 1 and 38% of the colonies in Case 2. The SMCs were determined to be derived from chromosome 6 during postnatal confirmation studies. Although the sizes and the chromosomal origin of the SMCs in these two cases appeared to be similar, the clinical outcomes varied. The clinical manifestations observed in Case 1 included small for gestational age, feeding difficulty at birth, hydronephrosis, deviated septum and dysmorphic features, while the phenotype is apparently normal in Case 2. Array comparative genomic hybridization (CGH) was performed and showed increase in dosage for approximately 26 Mb of genetic material from the proximal short and long arms of chromosome 6 in Case 1. Results of array CGH were uninformative in Case 2, either due to mosaicism or lack of detectable euchromatin. The difference in the clinical presentation in these two patients may have resulted from the difference in the actual gene contents of the marker chromosomes and/or the differential distribution of the mosaicism.

Array CGH analysis shows that aneuploidy is not related to the number of embryos generated

This study retrospectively analysed array comparative genomic hybridization (CGH) results of 7753 embryos from 990 patients to determine the frequency of embryonic euploidy and its relationship with the cohort size (i.e. the number of embryos available for biopsy and array CGH analysis). Linear regression analysis was performed to assess the effect of cohort size on euploidy rate adjusted for the effect of female age. While increasing female age was associated with a significant decrease in euploidy rate of day-3 and day-5 embryos (P<0.001 for both groups), cohort size was not significantly associated with euploidy rate. Logistic regression analysis was performed to assess the effect of cohort size, adjusted for maternal age, on the likelihood of having at least one euploid embryo available for transfer. The odds of having at least one euploid embryo in an assisted cycle was significantly decreased by increasing female age (P<0.01 for both day-3 and day-5 embryos) and was significantly increased by every additional embryo available for analysis (P<0.001 for both day-3 and day-5 embryos).Array comparative genomic hybridization (CGH) enables assessment of the entire chromosomal component of human embryos, which was not possible with the formerly used preimplantation genetic diagnosis (PGD) technology. We analysed array CGH PGD results of 7753 embryos from 990 women to determine the frequency of embryonic euploidy and its relationship with the number of embryos available for biopsy (hereafter called cohort size) in an assisted reproduction cycle. While increasing female age was associated with a significant decrease in euploidy rate of day-3 and day-5 embryos (P<0.001 for both groups), cohort size was not significantly associated with euploidy rate. Logistic regression analysis was performed to assess the effect of cohort size, adjusted for maternal age, on the likelihood of having at least one euploid embryo available for transfer. The odds of having at least one euploid embryo in an assisted cycle was significantly decreased by increasing female age (P<0.01 for both day-3 and day-5 embryos) and was significantly increased by every additional embryo available for analysis (P<0.001 for both day-3 and day-5 embryos). In conclusion, increasing the number of embryos available in a conventionally stimulated cycle increases the chance of having an euploid embryo available for transfer.