Dissertations / Theses: 'Array CGH'

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Shah, Sohrab P. "Model based approaches to array CGH data analysis." Thesis, University of British Columbia, 2008. http://hdl.handle.net/2429/2808.

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DNA copy number alterations (CNAs) are genetic changes that can produce adverse effects in numerous human diseases, including cancer. CNAs are segments of DNA that have been deleted or amplified and can range in size from one kilobases to whole chromosome arms. Development of array comparative genomic hybridization (aCGH) technology enables CNAs to be measured at sub-megabase resolution using tens of thousands of probes. However, aCGH data are noisy and result in continuous valued measurements of the discrete CNAs. Consequently, the data must be processed through algorithmic and statistical techniques in order to derive meaningful biological insights. We introduce model-based approaches to analysis of aCGH data and develop state-of-the-art solutions to three distinct analytical problems. In the simplest scenario, the task is to infer CNAs from a single aCGH experiment. We apply a hidden Markov model (HMM) to accurately identify CNAs from aCGH data. We show that borrowing statistical strength across chromosomes and explicitly modeling outliers in the data, improves on baseline models. In the second scenario, we wish to identify recurrent CNAs in a set of aCGH data derived from a patient cohort. These are locations in the genome altered in many patients, providing evidence for CNAs that may be playing important molecular roles in the disease. We develop a novel hierarchical HMM profiling method that explicitly models both statistical and biological noise in the data and is capable of producing a representative profile for a set of aCGH experiments. We demonstrate that our method is more accurate than simpler baselines on synthetic data, and show our model produces output that is more interpretable than other methods. Finally, we develop a model based clustering framework to stratify a patient cohort, expected to be composed of a fixed set of molecular subtypes. We introduce a model that jointly infers CNAs, assigns patients to subgroups and infers the profiles that represent each subgroup. We show our model to be more accurate on synthetic data, and show in two patient cohorts how the model discovers putative novel subtypes and clinically relevant subgroups.

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Mohrmann, Inga [Verfasser]. "Array-CGH bei Patienten mit Intelligenzminderung / Inga Mohrmann." Lübeck : Zentrale Hochschulbibliothek Lübeck, 2014. http://d-nb.info/1046429280/34.

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Lee, Sansan. "Genetic counseling perspectives on prenatal array CGH testing." Waltham, Mass. : Brandeis University, 2009. http://dcoll.brandeis.edu/handle/10192/23259.

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VIDETTA, ALESSANDRO DAVIDE. "Molecular analysis: an invaluable approach to improve diagnosis and tailor therapy." Doctoral thesis, Università di Siena, 2017. http://hdl.handle.net/11365/1011505.

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Neoplastic transformation can start in nearly every cell type in the human body. It is recognisable as cells acquiring the ability to divide uncontrollably and to escape aging mechanisms and naturally occurring cell death, resulting in the growth of a tumour. Tumours have different features, depending on the organ of origin and the level of differentiation of the tumour cells. At certain points in development, a tumour will be influencing its microenvironment, ensuring, among other things, vascularisation and cooperation with the immune system. A tumour can progress further, evolving into malignant disease, by invading the surrounding tissue, disseminating into the bloodstream or lymphatic channels, and establishing metastases in other parts of the body, often with fatal consequences for the affected individual. The diversity of cancer, in both biological and clinical terms, is well acknowledged and has been extensively studied. Today, with increasingly sophisticated technologies at our disposal, highly detailed molecular features of individual tumours can be described. Such features are often referred to as being layered, occurring at the genomic (DNA), transcriptomic (mRNA) and functional proteomic (protein) levels. Proteins are the key functional elements of cells, resulting from transcription of a gene into mRNA, which is further translated into a protein. This simplistic way of describing the relationship between the layers has gradually changed during the past decades of functional and molecular insight. Protein synthesis is no longer perceived as a linear process, but as an intricate network of a multitude of operational molecules. Astonishing progress has been made in the discovery of molecules that are able to influence transcription and translation, such as DNA-modifying enzymes and non-translated RNAs, and of mechanisms that are able to control the processing, localisation and activation of proteins. A picture is emerging of individual cells within a tumour that can differ at the genomic, epigenomic and transcriptional levels, as well as at the functional level. Mutations and epigenetic alterations create the required phenotypic diversity that, under the influence of shifting selective pressures imposed by the environment, determines the sub-clonal expansion and selection of specific cells. The development of solid tumours thus follows the same basic principles as Darwinian evolution. Most single nucleotide polymorphism (SNP) variants that arise in human evolution are neutral with respect to survival advantage; over a period of time, these variants are typically fixed in or die out from the genome according to chance. Other variants provide a survival advantage and will, over time, dominate the cell population, leading to distinct haploid signatures. Cancer may involve hundreds or thousands of mutations, with each mutation potentially contributing to tumour fitness. Most of these mutations are assumed to be passengers, but a limited number have driver capability, sometimes only in a sub-population of cells. There is an intricate interplay between sub-populations of tumour cells and among tumour and normal cells in the microenvironment, and tumour topology is likely to play a role in this context. Our knowledge of molecular mechanisms in cancer development and progression are mainly derived from model systems such as in vitro cell cultures and animal models, as well as from descriptive molecular analyses of tissue samples. Model systems have been crucial for understanding molecular interactions and their implications in cancer, but they cannot fully mimic tumour conditions in vivo. Tissue samples, on the other hand, contain both a microenvironment and sub-populations of cancer cells, but they represent only a snapshot in an individual tumour’s life history. Until recently, cancer studies mainly considered only one or a few molecular levels at a time. Altered protein expression can have several causes; it can be due to copy number gain, a translocation event that combines the gene with an active promoter, alteration of factors that modify DNA or influence the transcription machinery, or modifications of mRNA or the protein itself. Revealing the various downstream effects of such alterations is potentially useful for tumour classification and for prediction of treatment response and prognosis.

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Rocha, Ana Laís Bignotto da. "Sequenciamento direto dos genes SIX3, SHH, TGIF1, ZIC2 e array-CGH no estudo de pacientes com holoprosencefalia." Universidade de São Paulo, 2013. http://www.teses.usp.br/teses/disponiveis/61/61132/tde-12112013-150520/.

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Objetivos: Analisar por meio da técnica de sequenciamento direto a presença de alterações moleculares nos genes SHH, SIX3, ZIC2 e TGIF1 em indivíduos com diagnóstico clínico de HPE. Analisar por meio da técnica de CGH-array a presença de alterações moleculares em indivíduos com diagnóstico clínico de HPE previamente submetidos à análise por sequenciamento direto. Local: Laboratório de Genética e Citogenética Humana HRAC/USP, Bauru-SP. Casuística e metodologia: Foram selecionados 50 indivíduos, de ambos os sexos com idades entre 03 meses a 50 anos com diagnóstico clínico para HPE. Todos foram analisados por meio da técnica de sequenciamento direto para os genes SHH e TGIF1 completamente e para os genes ZIC2 e SIX3 parcialmente. Dentre os indivíduos que não apresentaram alterações na técnica de sequenciamento oito indivíduos com fenótipo mais grave foram selecionados para a análise por CGH-array. Resultados e discussão: Foram analisados 50 indivíduos por meio da técnica de sequenciamento direto dos gene SHH e TGIF1, foram encontradas duas variantes patogênicas na análise do gene SHH, no caso 1 a variante p.24G>P foi identificada, e no caso 2 foi identificada a variante c.1031del C. No gene TGIF1 foram encontrados cinco polimorfismos já descritos na literatura. Foi identificada uma nova variante silenciosa no éxon 1 do gene ZIC2 p.Q46Q (c. 431 G>A) e um polimorfismo já descrito na literatura em dois indivíduos no gene SIX3. A análise por CGH-array revelou a presença de uma microdeleção no caso 37, de 1,5Mb no cromossomo 17p12 entre as posições genômicas 14,052,279-15,102,307. A mesma deleção foi encontrada na mãe, sendo que esta região nunca foi associada a HPE. Conclusão: A técnica de sequenciamento direto é uma ferramenta muito importante no diagnóstico molecular da HPE, a padronização do sequenciamento direto para os genes ZIC2 e SIX3 poderá auxiliar em diagnósticos mais precisos em estudos futuros dentro do HRAC/USP. O emprego de novas técnicas como CGH-array pode indicar novas relações entre regiões cromossômicas e os múltiplos fatores envolvidos na formação da HPE.
Objective: Analyze through direct sequencing technique the presence of molecular changes on the genes SHH, SIX3, ZIC2 and TGIF1 on individuals with clinical diagnosis of HPE. Analyze through array-CGH technique the presence of molecular changes on individuals with clinical diagnosis of HPE previously submitted to the direct sequencing analyzes. Local: Genetics and Human Cytogenetics Laboratory, HRAC/USP, Bauru-SP. Methods: Were selected 50 individuals from both genders with ages between 03 months and 50 years clinically diagnosed with HPE. Everyone was analyzed through the direct sequencing technique for the genes SHH and TGIF1 completely and for the genes ZIC2 and SIX3 partially. From those individuals which did not have shown changes on the direct sequencing technique, eight individuals with more severe phenotype were selected to the analysis through array-CGH. Results an Discussion: Were analyzed 50 individuals through the technique of direct sequencing of the genes SHH and TGIF1, were found two pathogenic variants in the analysis of SHH gene, in the case 1, the variant p.G24P was identified, and in the case 2 was identified the variant c.1031delC. On the TGIF1 gene were found five polymorphisms already described on the literature. Was identified a new silent variant on the exon 1 of the ZIC2 gene p. Q46Q(c.431G>A) and a polymorphism already described in the literature in two individuals on the gene SIX3. The analysis through array-CGH revealed the presence of one microdeletion in the case 37, of 1,5 Mb on the region 17p12 between the genomic positions 14,052,279-15,102,307. The same deletion was detected in the mother, though this region was never associated to the HPE. Conclusion: The direct sequencing technique is a very important tool for the molecular diagnosis of the HPE, and the direct sequencing standardization for the genes ZIC2 and SIX3 might help in more precise diagnostics on HRAC/USP future studies. The employ of new techniques such as array-CGH may indicate new relations between chromosomal regions and the multiple hit involved in the development of HPE.

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Marioni, John Carlo. "Statistical methods for array CGH and copy number variation experiments." Thesis, University of Cambridge, 2008. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.611877.

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Sporns, Peter [Verfasser]. "Korrelation von Array-CGH-Befunden und klinischem Phänotyp / Peter Sporns." Kiel : Universitätsbibliothek Kiel, 2015. http://d-nb.info/1065669992/34.

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Grzesiuk, Juliana Dourado. "Caracterização Citogenética Molecular de Rearranjos Cromossômicos Aparentemente Equilibrados Associados ao Fenótipo de Infertilidade." Universidade de São Paulo, 2012. http://www.teses.usp.br/teses/disponiveis/17/17135/tde-22042013-151132/.

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A translocação recíproca é o rearranjo equilibrado mais comum em humanos. Frequentemente, indivíduos com rearranjos equilibrados não apresentam manifestações clínicas, entretanto, na meiose, o pareamento entre cromossomos translocados forma uma figura quadrivalente em forma de cruz que torna a disjunção cromossômica incerta e dependendo do rearranjo, o individuo pode vir a ser infértil, apresentar um risco aumentado de abortamento espontâneo e/ou da prole apresentar alterações fenotípicas. Neste projeto, investigamos duas famílias de pacientes inférteis, portadores de translocações cromossômicas. O objetivo foi caracterizar as alterações citogenéticas e citogenômicas relacionadas à infertilidade masculina em pacientes portadores de rearranjos aparentemente equilibrados, associando técnicas de citogenética clássica (bandeamento GTG), citogenética molecular (FISH) e citogenômica (array-CGH). Foram estudados sete indivíduos da família 1, sendo diagnosticados três portadores da translocação (X;22), sendo um deles azoospérmico. Nesta família foram ainda detectados dois casos de mosaicismo para síndrome de Turner. A família 2 foi composta por dois irmãos oligozoospérmicos, portadores de translocação (8;13). Com a aplicação da técnica de FISH, definimos o cariótipo final dos portadores dos rearranjos como 46,XX ou 46,XY,t(X;22)(p22.3;q11.2) para a família 1 e 46,XY,t(8;13)(q13;q14)para a família 2. A técnica de array-CGH (plataforma 2x400K, Agilent) detectou alterações no número de cópias de alguns genes candidatos relacionados ao fenótipo de infertilidade, sendo a sequência 132 de piRNAs, os genes DDX11, Jagged 2 e ADAM18 na família 1 e os genes candidatos ADAM18 e POTE nos pacientes da família 2.
Reciprocal translocations are the most common balanced rearrangement in humans. Often individuals with balanced rearrangements show no clinical findings. However, in meiosis, the pairing between translocated chromosomes forms a quadrivalent cross-shaped figure which has the effect of making chromosome disjunction uncertain and, depending on the rearrangement, and on the segregation of the unbalanced chromosomes, the individual can be infertile, can present with an increased risk of spontaneous abortions or can have an offspring with abnormal phenotype. We have studied two families of infertile patients, who were carriers of chromosomal translocations. The objective was to characterize the cytogenetic and cytogenomic alterations related to male infertility in patients with apparently balanced rearrangements using classical cytogenetic techniques (GTG banding), molecular cytogenetics (FISH) and cytogenomics (array-CGH). Seven subjects of the family 1 were studied, including three carriers of translocation (X;22), one azoospermic. Two cases of mosaicism for Turner syndrome were detected in this family. The second family consisted of two oligozoospermic brothers with translocation (8;13). FISH was used to characterize the karyotypes as 46, XX or 46,XY, t(X;22)(p22.3;q11.2) for the members of the family 1 and 46,XY,t(8;13)(q13;q14) for family 2. Array-CGH was also performed using the Agilent platform 2x400K, to detect associated copy number variations of some of the candidate genes that could be related to infertility. In the family 1 the candidate genes were 132 piRNAs sequences and DDX11,Jagged 2 and ADAM18 genes. The candidate genes for the family 2 were ADAM18 and POT.

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Castells, Sarret Neus. "Array CGH com a primera opció per al diagnòstic genètic postnatal." Doctoral thesis, Universitat Autònoma de Barcelona, 2015. http://hdl.handle.net/10803/325159.

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La citogenètica convencional detecta un 3-5% dels pacients amb retard global del desenvolupament / discapacitat intel·lectual (RGD / DI) i / o malformacions congènites (MC). L'amplificació de sondes múltiples dependents de lligació (MLPA) permet incrementar la taxa diagnòstica entre 2,4-5,8%. Actualment els arrays d'hibridació genòmica comparada (aCGH) constitueixen l'eina diagnòstica amb major rendiment en pacients amb RGD / DI, MC i trastorns de l'espectre autista (TEA). L'objectiu del present treball ha estat avaluar l'eficiència de l'ús dels aCGH com a tècnica de primera opció diagnòstica en aquestes i altres indicacions (epilèpsia, talla baixa). Per assolir aquest objectiu, s'han estudiat 1.000 pacients afectats de les patologies abans esmentades mitjançant la tècnica de aCGH, utilitzant una estratègia d'hibridació pacient versus pacient i afegint el suport de la tècnica de MLPA en el 50% dels pacients estudiats. En primer lloc es va validar la tècnica i es va escollir la plataforma d’oligoarrays. Per tal de minimitzar costos i incrementar la eficiència, es va utilitzar l’estratègia d’hibridació de pacient versus pacient amb MLPA confirmatòria i es van establir criteris de diagnòstic per optimitzar la detecció de desequilibris patogènics. Per facilitar la interpretació dels resultats es va dissenyar un programa informàtic EasyArray. Es van detectar desequilibris d'efecte patogènic en un 14% dels pacients (140 / 1.000). En funció del fenotip es van diagnosticar un 18,9% dels pacients afectats de RGD / DI; un 13,7% de les MC; un 9,75% de les patologies psiquiàtriques, 1 7,01% dels casos amb epilèpsia i un 13,3% dels pacients amb talla baixa. Dins de les MC destaquen les del sistema nerviós central amb un 14,9% i les cardiopaties congènites amb un 10,6% de diagnòstics. Dins de les patologies psiquiàtriques, destaquen els pacients amb TEA amb un 8.9% de diagnòstics. Podem concloure que la tècnica d’arrayCGH té un rendiment diagnòstic molt superior al del cariotip amb bandes G. La seva utilització com a eina diagnòstica de primera opció juntament amb el disseny d’estratègies d’hibridació no estàndards, suposa una reducció considerable de costos. Els nostres resultats demostren l'efectivitat i eficiència de la utilització de l’arrayCGH com a primera opció en el diagnòstic genètic dels pacients amb sospita de desequilibris genòmics. Tot això avala la seva inclusió dins del Sistema Nacional de Salut.
Conventional cytogenetics diagnoses 3-5% of patients with unexplained developmental delay / intellectual disability (DD / ID) and / or multiple congenital Anomalies (MCA). Multiplex ligation probes Amplification (MLPA) increases diagnostic rate between 2.4 to 5.8%. Currently the array comparative genomic Hybridization (CGH) is the highest performing diagnostic tool in patients with DD / ID, MC and autism spectrum disorders. Our aim was to evaluate the efficiency of the use of aCGH as first-line test replacing the karyotype and MLPA in these and other pathological indications (epilepsy, short stature). A total of 1000 patients referred by one or more of the above mentioned disorders were analysed by aCGH using a methodology / strategy hybrid alternative patient versus patients adding support MLPA technique in 50% of patients studied. Following a validation period, an oligoarray platform was chosen. In order to minimize costs and increase efficiency, a patient versus patient hybridization strategy plus MLPA confirmation was used, and analysis criteria were set to optimise detection of pathogenic imbalances. In order to facilitate interpretation of results, a database application named Easy Array was designed. Pathogenic genomic imbalances were detected in 14% of the cases (140/1000), with a variable distribution of diagnosis according to the phenotypes: 18.9% of patients with DD / ID, 13.7% of MCAS, 9.75% of Psychiatric pathologies, 7.01% of patients with Epilepsy and 13.3% of patients with short Stature. Within the MCA, central nervous system abnormalities and congenital heart diseases accounted for 14.9% and 10.6% of diagnosis respectively. Among the Psychiatric disorders, patients with ASD accounted for 8.9% of diagnosis. We can conclude that Array-CGH provides a substantially higher diagnostic yield tan G-banded chromosomes analysis. Its use as first line test and the development of non-standard hybridization strategies reduces consumable costs considerably. Our results demonstrate the effectiveness and efficiency of the use of arrayCGH as the first line test in genetic diagnosis of patients suspected of genomic imbalances, supporting its inclusion within the National Health System.

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Zhang, Yunyu. "Hidden Markov Model inference copy number change in array-CGH data." Thesis, Massachusetts Institute of Technology, 2005. http://hdl.handle.net/1721.1/33086.

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Thesis (S.M.)--Harvard-MIT Division of Health Sciences and Technology, 2005.
Includes bibliographical references (p. 56-57).
Cancer development and progression typically features genomic instability frequently resulting in genomic changes involving DNA copy number gains or losses. Identifying the genomic location of these regional alterations provides important opportunities for the discovery of potential novel oncogenes and tumor suppressors. Recently, array based competitive genomic hybridization (array-CGH) has become available as a powerful approach for genome-wide detection of DNA copy number changes. Array-CGH assesses DNA copy number in tumor samples through competitive hybridization on microarrays containing probes for thousands of genes. The datasets generated are complex and require statistical methods to accurately define discrete and uniform copy number from the data and to identify transitions between genomic regions with altered copy number. Several approaches based on different statistical frameworks have been developed. However, a fundamental informatic issue in array-CGH analysis remains unsolved by these methods. In particular, sample-specific data compression, a result of tumor cells being commonly admixed with normal cells in many tumor types, must be accounted for in each sample analyzed. Additionally, in order to accurately assess deviations from normal copy number, the copy number readout must be shifted to faithfully represent the baseline copy number in each tumor sample. Failure to appropriately address these issues reduces the accuracy of the data in hard-threshold based high-level analysis.
(cont.) By using the natural framework Hidden Markov Models (HMM) to model the distribution of array-CGH signals, a method infer the absolute copy number and identify change points has been developed to address the above problems. This method has been validated on independent dataset and its utility in inference on array-CGH data is demonstrated here.
by Yunyu Zhang.
S.M.

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Jaillard-Herrebrecht, Sylvie. "Place de la CGH-array dans l'étude des anomalies du développement." Rennes 1, 2010. http://www.theses.fr/2010REN1B141.

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Kreuz, Markus. "Entwicklung und Implementierung von Auswertungswerkzeugen für Hochdurchsatz-DNA-Kopienzahl-Analysen und deren Anwendung auf Lymphomdaten." Doctoral thesis, Universitätsbibliothek Leipzig, 2015. http://nbn-resolving.de/urn:nbn:de:bsz:15-qucosa-161664.

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Aberrationen in der DNA-Kopienzahl sind häufige genetische Veränderungen bei malignen Lymphomerkrankungen. Zugewinne sowie Deletionen stellen dabei Mechanismen zur Onkogen-Aktivierung sowie Tumorsuppressorgen-Inaktivierung dar und tragen somit zur Pathogenese der Erkrankung bei. Array-CGH und SNP-Array sind Messplattformen, die die genomweite Bestimmung von Kopienzahlaberrationen in einem Experiment ermöglichen. Die bei der Analyse entstehenden Datensätze sind komplex und erfordern automatische Methoden zur Unterstützung der Analyse und Interpretation der Messergebnisse. In dieser Promotionsarbeit wurden Methoden entwickelt, welche die Analyse von Array-CGH- und SNP-Array-Messungen ermöglichen. Diese Methoden wurden für die Auswertung umfangreicher Datensätze von malignen Non-Hodgkin-Lymphomen verwendet. Dabei wurden Lymphome der Entitäten Burkitt-Lymphom, diffus großzelliges B-Zell-Lymphom, Mantelzelllymphom, primäres ZNS-Lymphom und peripheres T-Zell-Lymphom – nicht anderweitig spezifiziert – analysiert. Für die untersuchten Lymphom-Entitäten konnten hierbei zahlreiche neue rekurrente Kopienzahlaberrationen sowie uniparentale Disomien gezeigt werden, die neue Einblicke in die Pathogenese der jeweiligen Erkrankungen erlauben. Darüber hinaus erfolgte ein Vergleich beider Messplattformen anhand eines Datensatzes mit gepaarten Array-CGH- und SNP-Array-Daten. Für die eingesetzten Plattformen (2800k-BAC-Array vs. Affymetrix 250k-Sty-SNP-Array) konnte eine circa zwölffach höhere effektive Auflösung der SNP-Array-Plattform gezeigt werden. Die wesentlichen Ergebnisse dieser Arbeit sind in sieben Publikationen eingeflossen.

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Paiva, Greicy Helen Gambarini [UNESP]. "Genes candidatos a marcadores tumorais na progressão do adenocarcinoma de próstata indentificados por análise de HR-CGH e CGH-ARRAY." Universidade Estadual Paulista (UNESP), 2009. http://hdl.handle.net/11449/102718.

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Made available in DSpace on 2014-06-11T19:32:14Z (GMT). No. of bitstreams: 0 Previous issue date: 2009-02-01Bitstream added on 2014-06-13T19:02:40Z : No. of bitstreams: 1 paiva_ghrg_dr_botib.pdf: 1701322 bytes, checksum: d1fa5b5c562a2a6ce8ad0d14ab948d4a (MD5)
Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)
O câncer de próstata (CaP) é a neoplasia mais comumente diagnosticada entre homens no ocidente. Embora tratamentos efetivos para a doença localizada estejam disponíveis atualmente, não há terapia curativa para tumores metastáticos. Além disso, os marcadores diagnósticos utilizados na clínica não conseguem discriminar totalmente a evolução diferencial da doença. Desta forma, o conhecimento das diferenças biológicas entre tumores primários confinados ao órgão e metástases é essencial para o desenvolvimento de novos marcadores e identificação de alvos terapêuticos. Neste estudo a análise baseada na metodologia de HR-CGH cromossômico foi realizada para identificar alterações de ganhos e perdas genômicas em três grupos de amostras: o grupo I, que compreende amostras pareadas de tumor primário e respectivas metástases (11 casos); o grupo II, constituído de pacientes que apresentaram seguimento clínico favorável por mais de 10 anos (5 casos); e o grupo III, constituído por diferentes biópsias do mesmo paciente (5 pacientes com 2 biópsias cada). As amostras foram microdissecadas (amostras a fresco: a partir de lâminas de referência; em blocos de parafina: a laser) e após a obtenção de DNA foram amplificadas (amostras de arquivo: PCR-SCOMP) ou marcadas por nick-translation para a realização de HR-CGH. Os resultados de HR-CGH foram comparados com os dados obtidos da análise de CGH-array num subgrupo de amostras e revelaram concordâncias significativas. Os resultados obtidos na presente investigação revelaram perdas dos cromossomos 1p, 2, 3q, 4p, 5q, 7, 8, 9q, 10q, 11q, 12q, 14q, 15q, 16q, 17q, 18q, 19, 20q e 22q em 80% dos casos avaliados. Além disso, perdas em 17q11.2-25, por exemplo, foram detectadas exclusivamente nos tumores do grupo I e nas suas metástases, e não nos tumores do grupo II, sugerindo que esta alteração deve ser importante...
Prostate cancer (PCa) is the most commonly diagnosed non-cutaneous malignancy and the second leading cause of cancer mortality in men from Occident. Although effective treatments for the localized disease are available, there is no efficient therapy for metastatic tumors. Additionally, clinical diagnostic markers are not able to completely discriminate the differential evolution of the disease. The knowledge of biological differences between localized primary tumors and metastasis can establish new molecular markers and therapeutic targets. In this study, an analysis based on HR-CGH methodology was performed to identify imbalances genomic in three groups of samples: group I, paired samples of primary tumors and its metastasis (11 cases); group II, patients that exhibited favorable follow-up over 10 years (5 cases); and group III, different biopsies from the same patient (5 patients with 2 biopsies each). The tumor samples were submitted to microdissection procedures (fresh samples: from reference slides; paraffin embedded samples: laser), DNA extracted and amplified (archive sample: PCR-SCOMP) or labeled by nick-translation to HR-CGH. The HRCGH results were compared with data obtained from CGH-array analysis of a subgroup of samples and revealed significant concordances. In the present investigation, there were observed losses on chromosomes 1p, 2, 3q, 4p, 5q, 7, 8, 9q, 10q, 11q, 12q, 14q, 15q, 16q, 17q, 18q, 19, 20q and 22q in 80% of the cases. Losses in 17q11.2-25, for instance, were detected exclusively in tumor from group I and its metastasis, but were not found in tumors from group II, suggesting that this alteration must be important in the progression of the disease. Five genes were selected after the comparison between the HR-CGH and CGH-array data. The tumor suppressor genes ARID1A, MTSS1, NME1 and S100A4 and TOP2A (oncogenes) were evaluated by quantitative real time... (Complete abstract click electronic access below)

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Paiva, Greicy Helen Gambarini. "Genes candidatos a marcadores tumorais na progressão do adenocarcinoma de próstata indentificados por análise de HR-CGH e CGH-ARRAY." Botucatu : [s.n.], 2009. http://hdl.handle.net/11449/102718.

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Orientador: Silvia Regina Rogatto
Banca: Spencer L. M. Payão
Banca: Carla Rosemberg
Banca: José Carlos de S. Trindade
Banca: Maria Aparecida M. Rodrigues
Resumo: O câncer de próstata (CaP) é a neoplasia mais comumente diagnosticada entre homens no ocidente. Embora tratamentos efetivos para a doença localizada estejam disponíveis atualmente, não há terapia curativa para tumores metastáticos. Além disso, os marcadores diagnósticos utilizados na clínica não conseguem discriminar totalmente a evolução diferencial da doença. Desta forma, o conhecimento das diferenças biológicas entre tumores primários confinados ao órgão e metástases é essencial para o desenvolvimento de novos marcadores e identificação de alvos terapêuticos. Neste estudo a análise baseada na metodologia de HR-CGH cromossômico foi realizada para identificar alterações de ganhos e perdas genômicas em três grupos de amostras: o grupo I, que compreende amostras pareadas de tumor primário e respectivas metástases (11 casos); o grupo II, constituído de pacientes que apresentaram seguimento clínico favorável por mais de 10 anos (5 casos); e o grupo III, constituído por diferentes biópsias do mesmo paciente (5 pacientes com 2 biópsias cada). As amostras foram microdissecadas (amostras a fresco: a partir de lâminas de referência; em blocos de parafina: a laser) e após a obtenção de DNA foram amplificadas (amostras de arquivo: PCR-SCOMP) ou marcadas por nick-translation para a realização de HR-CGH. Os resultados de HR-CGH foram comparados com os dados obtidos da análise de CGH-array num subgrupo de amostras e revelaram concordâncias significativas. Os resultados obtidos na presente investigação revelaram perdas dos cromossomos 1p, 2, 3q, 4p, 5q, 7, 8, 9q, 10q, 11q, 12q, 14q, 15q, 16q, 17q, 18q, 19, 20q e 22q em 80% dos casos avaliados. Além disso, perdas em 17q11.2-25, por exemplo, foram detectadas exclusivamente nos tumores do grupo I e nas suas metástases, e não nos tumores do grupo II, sugerindo que esta alteração deve ser importante... (Resumo completo, clicar acesso eletrônico abaixo)
Abstract: Prostate cancer (PCa) is the most commonly diagnosed non-cutaneous malignancy and the second leading cause of cancer mortality in men from Occident. Although effective treatments for the localized disease are available, there is no efficient therapy for metastatic tumors. Additionally, clinical diagnostic markers are not able to completely discriminate the differential evolution of the disease. The knowledge of biological differences between localized primary tumors and metastasis can establish new molecular markers and therapeutic targets. In this study, an analysis based on HR-CGH methodology was performed to identify imbalances genomic in three groups of samples: group I, paired samples of primary tumors and its metastasis (11 cases); group II, patients that exhibited favorable follow-up over 10 years (5 cases); and group III, different biopsies from the same patient (5 patients with 2 biopsies each). The tumor samples were submitted to microdissection procedures (fresh samples: from reference slides; paraffin embedded samples: laser), DNA extracted and amplified (archive sample: PCR-SCOMP) or labeled by nick-translation to HR-CGH. The HRCGH results were compared with data obtained from CGH-array analysis of a subgroup of samples and revealed significant concordances. In the present investigation, there were observed losses on chromosomes 1p, 2, 3q, 4p, 5q, 7, 8, 9q, 10q, 11q, 12q, 14q, 15q, 16q, 17q, 18q, 19, 20q and 22q in 80% of the cases. Losses in 17q11.2-25, for instance, were detected exclusively in tumor from group I and its metastasis, but were not found in tumors from group II, suggesting that this alteration must be important in the progression of the disease. Five genes were selected after the comparison between the HR-CGH and CGH-array data. The tumor suppressor genes ARID1A, MTSS1, NME1 and S100A4 and TOP2A (oncogenes) were evaluated by quantitative real time... (Complete abstract click electronic access below)
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Fuhrmann, Christine. "Entwicklung der Array-CGH zur hochauflösenden, genomweiten Untersuchung von DNA-Veränderungen einzelner Tumorzellen." Diss., lmu, 2008. http://nbn-resolving.de/urn:nbn:de:bvb:19-89535.

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Fuhrmann, Christine. "Entwicklung der Array-CGH zur hochauflösenden, genomweiten Untersuchung von DNA-Veränderungen einzelner Tumorzellen." kostenfrei, 2008. http://edoc.ub.uni-muenchen.de/8953/.

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Piard, Juliette. "Déficience intellectuelle : identification de nouveaux gènes par une approche multicentrique." Thesis, Bourgogne Franche-Comté, 2018. http://www.theses.fr/2018UBFCE005.

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La déficience intellectuelle (DI) touche 1 à 3% de la population générale avec un excès de sujets de sexe masculin. Cette affection est caractérisée par une extrême hétérogénéité clinique et génétique rendant son élucidation complexe. La révolution technologique des outils permettant l’analyse du génome intervenue depuis les années 2000 avec l’analyse chromosomique sur microréseau et singulièrement depuis 2010 avec les applications du séquençage à haut débit a considérablement facilité l’identification de nouveaux gènes. Nous avons tiré avantage de ce phénomène pour identifier trois affections neurologiques à caractère familial Nous avons procédé selon une méthodologie structurée pour conduire, grâce à la mise en place d’un réseau de collaborations, à la découverte ou à la confirmation de l’existence de nouvelles formes de DI. 1.Séquençage de l'exome couplé à la recherche de variations du nombre de copies 2. Mise à jour d’une altération de séquence génique potentiellement causale retrouvée chez le cas index et chez les autres sujets atteints de la famille 3.Extension des résultats à d’autres familles par la constitution d’une cohorte de réplication 4.Élaboration d’une série d’études fonctionnelles venant conforter l’hypothèse de causalité par la création d’un modèle animal et/ou la réalisation d’études biochimiques spécifiquesL’application de cette méthodologie nous a permis de conduire à terme trois projets : L’individualisation d’une forme syndromique de DI récessive autosomique associée à une malformation du rachis cervical et liée aux mutations bi-alléliques de CDK10. La caractérisation d’une encéphalopathie récessive autosomique létale associée à une hypertonie sévère et à une arthrogrypose distale liée aux mutations bi-alléliques d’ATAD1. L’implication de FRMPD4 dans une nouvelle forme de DI non syndromique liée à l’X
Intellectual disability (ID) impacts 1 to 3% of the general population with an excess of affected males. This condition is characterized by an extreme clinical and genetic heterogeneity making the deciphering of its causes more complex. The technological revolution that took place in the study of the genome over the last two decades has provided a useful tool for identification of new genetic entities. This is particularly true for chromosomal micro-array analysis since early 2000s and for next generation sequencing since 2011. We took advantage of this by identifying the molecular basis of three singular conditions. We applied a structured methodology and created a network of collaborations to define or confirm these new ID syndromes. 1. Whole exome sequencing alongside with array-CGH 2.Identification of a candidate gene sequence alteration in the index case and other affected patients of the family 3.Constitution and study of a replication cohort 4.Biochemical studies and/or animal models in order to support the assumption of causalityBased on this research strategy, we were able to complete the following projects : Discovery of a syndromic form of autosomal recessive ID associated with cervical spine defects due to bi-allelic CDK10 mutations. Identification of an ATAD1-related profound and lethal autosomal recessive encephalopathy with stiffness and distal arthrogryposis. Characterization of a FRMPD4-related X-linked non-syndromic ID

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Coe, Bradley P. "The role of specific genomic alterations in small cell lung cancer aggressiveness." Thesis, University of British Columbia, 2008. http://hdl.handle.net/2429/2283.

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Small Cell Lung Cancer (SCLC) is a very aggressive neuroendocrine tumour of the lung, which demonstrates a 5 year survival of only 10% for extensive stage disease (20-30% for limited stage), with only modest improvement over the last few decades. Identification of new molecular diagnostic and therapeutic targets is thus imperative. Previous efforts in identifying molecular changes in SCLC by gene expression profiling using microarrays have facilitated disease classification but yielded very limited information on SCLC biology. Previous DNA studies have been successful in identifying several loci important to SCLC. However the low resolution of conventional chromosomal Comparative Genomic Hybridization (CGH) has limited the findings to large chromosomal regions with only a few specific candidate genes discovered to date. Thus, to further understand the biological behaviour of SCLC, better methods for studying the genomic alterations in SCLC are necessary. This thesis highlights the development of array CGH technology for the high resolution dissection of aneuploidy in cancer genomes and the application of this new technology to the study of SCLC. I present the development of the first whole genome CGH array which offered unprecedented resolution in the profiling of cancer genomes allowing fine mapping of genes in a single experiment. Through application of DNA based analysis in conjunction with integrated expression analysis and comparison of SCLC to less aggressive non-small cell lung tumours I have identified novel patterns of pathway disruption specific to SCLC. This included alteration to Wnt pathway members and striking patterns of cell cycle activation through predominantly downstream disruption of signalling pathways including direct activation of the E2F transcription factors, which are normally repressed by the Rb gene. Analysis of targets of the E2F/Rb pathway identified EZH2 as being specifically hyper-activated in SCLC, compared to NSCLC. EZH2 is a polycomb group gene involved in the control of many cellular functions including targeted DNA methylation and escape from senescence in hematopoietic stem cells. Taken together these results suggest that in SCLC, downstream disruption may replace multiple upstream alterations leading to activation independent of a specific mitogenic pathway, and that EZH2 represents a potentially important therapeutic target.

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Móz, Luis Eduardo Silva. "Caracterização genômica do Edema de Reinke." Botucatu, 2017. http://hdl.handle.net/11449/149913.

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Orientador: Patrícia Pintor dos Reis
Resumo: Introdução: o Edema de Reinke (ER) é uma lesão laríngea considerada benigna relacionada ao tabagismo. Dados em literatura relatam associações entre o ER e a detecção de diferentes graus de displasia e carcinoma in situ, bem como alterações na imunoexpressão de proteínas tumorais como a p53. Alguns autores classificam o ER entre as lesões pré-malignas, com risco de transformação e progressão para carcinoma de laringe. Não havendo consenso na literatura, torna-se necessária a realização de estudos moleculares. Objetivos: caracterizar o perfil genômico global de alterações no número de cópias do DNA em amostras de pacientes com ER. Métodos: oito amostras removidas por microcirurgia foram submetidas à extração do DNA. Os perfis de alteração no número de cópias genômicas e os genes candidatos associados foram analisados pela metodologia da hibridação genômica comparativa (CGH array), utilizando-se a plataforma de 4x180K (Agilent Technologies). Os dados de microarranjos foram analisados utilizando o programa CytoGenomics v4.0.2.21 (Agilent Technologies). As alterações no número de cópias (CNAs) obtidas foram comparadas com o banco de dados Database of Genomic Variants (DGV). A classificação dos genes selecionados para análise foi realizada baseada em dados descritos no National Center for Biotechnology Information (NCBI). Resultados: Foram encontrados perdas, ganhos ou deleções em 54 genes, um RNA não codificador longo intergênico (lincRNA), seis sequências hipotéticas e 10 microRN... (Resumo completo, clicar acesso eletrônico abaixo)
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Delahaye-Duriez, Andrée. "Identification de nouveaux gènes impliqués dans des maladies ophtalmologiques rares en utilisant la CGH-array." Paris 7, 2011. http://www.theses.fr/2011PA077066.

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L'objectif de ce travail était de caractériser par CGH (Comparative Genomic Hybridization) array des régions chromosomiques impliquées dans des maladies ophtalmologiques rares, pour identifier de nouveaux gènes. Dans le premier volet de ma thèse, 65 patients présentant une malformation oculaire syndromique ont été explorés par CGH-array. Une anomalie probablement causale a été identifiée chez 15% d'entre eux. Quatre patients ont une délétion emportant un gène déjà connu dans le développement de l'œil (FOXC1 ou OTX2) et 4 autres patients, une délétion pathogénique non classiquement associée à une atteinte oculaire : del(17)(p!3. 3pl3. 3), del(10)(pl4p!5. 3) et del(16)(pll. 2pll. 2). En collaboration avec d'autres équipes nous avons rassemblé des patients pour étudier les corrélations génotype-phénotype des délétions 6p25 et 17pl3. 3. La deuxième partie de ce travail s'est centrée sur l'étude d'un gène candidat : ARHGEF26. L'étude de la ségrégation dans la famille index et le séquençage de ce gène chez des patients à phénotype commun ne nous ont pas permis de conclure à l'implication d'ARHGEF26 dans ce phénotype. Cette deuxième partie souligne les difficultés et les limites de la stratégie d'identification de nouveaux gènes par CGH-array. Au total, ces résultats montrent que l'analyse chromosomique par CGH-array au-delà de son intérêt diagnostique et pour le conseil génétique, permet d'établir de nouvelles corrélations génotype-phénotype et d'identifier de nouvelles régions potentiellement impliquées dans des pathologies ophtalmologiques rares
The karyotype detects a chromosomal anomaly in 7. 7% to 10% of neonates with ocular birth defect. The introduction of microarray technology showed a very high rate of rearrangements below the resolution of karyotyping. My objectives in this work were to characterize using comparative genome hybridisation-based microarray analysis (array-CGH) chromosomal regions involved in rare ophthalmologic disorders, and then to identify new genes. In the first part of my work, we performed array-CGH in 65 patients presenting syndromal ocular developmental anomalies. A causal or potentially causal anomaly was found for 15% of them. Four had a pathogenic deletion involving a gene known to be involved in ocular anomalies (FOXC1 or OTX2}, while 4 others had a pathogenic deletion not classically associated with ocular malformations: del(17)(pl3. 3p!3. 3), del(10)(pl4p!5. 3) and del(16)(pl 1. 2pl 1. 2). In collaboration with other teams, we gathered patients to study genotype-phenotype correlations for 6p25 and 17pl3. 3 deletions. The second part of my work focused on a candidate gene study: ARHGEF26. Sequencing this gene in other patients with similar phenotype and studying the index patient family segregation, we could not demonstrate the ARHGEF26 involvement in this phenotype. This second part highlights the limits and difficulties of gene identification using array-CGH. These results demonstrate that array-CGH-based chromosomal analysis, beyond its importance for diagnosis and genetic counselling, can help to establish new genotype-phenotype correlations for chromosomal anomalies as well as identify potential new regions involved in rare ophthalmologic disorders

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Lohan, Silke [Verfasser]. "Analyse von genomischen Aberrationen mit hochauflösender Array-CGH bei Patienten mit Fehlbildungen der Extremitäten / Silke Lohan." Berlin : Freie Universität Berlin, 2013. http://d-nb.info/1031190546/34.

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Machado, Isabela Nelly. "Detecção de instabilidade genômica por hibridização genômica comparativa baseada em microarranjos (array CGH) em fetos dismórficos." [s.n.], 2010. http://repositorio.unicamp.br/jspui/handle/REPOSIP/311739.

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Orientador: Ricardo Barini
Tese (doutorado) - Universidade Estadual de Campinas. Faculdade de Ciências Médicas
Made available in DSpace on 2018-08-16T03:43:19Z (GMT). No. of bitstreams: 1 Machado_IsabelaNelly_D.pdf: 3476920 bytes, checksum: d5a5716fb5a528c323d17a22ef6c28d0 (MD5) Previous issue date: 2010
Resumo: Introdução: Para uma parcela significativa de fetos com defeitos congênitos o diagnóstico sindrômico permanece indefinido, dificultando a abordagem perinatal, o estabelecimento de prognóstico e o aconselhamento genético. A incapacidade de detecção de pequenas instabilidades genômicas, atualmente apontadas como provável fator causal nestas condições dismórficas, é a principal limitação do estudo cromossômico microscópico pelo bandamento G (cariótipo convencional). A hibridização genômica comparativa (comparative genomic hybridization-CGH) é capaz de identificar perdas e ganhos de material genômico com alta resolução, sem envolver o cultivo celular e o conhecimento prévio da região genômica envolvida. Objetivo: Avaliar a aplicabilidade da técnica de array CGH em sangue fetal para o diagnóstico de perdas e ganhos genômicos em um grupo de fetos dismórficos. Sujeitos/Método: Foi realizado um estudo prospectivo descritivo a partir de amostras sanguíneas de fetos dismórficos e com cromossomos numericamente normais ao bandamento G, admitidos no Setor de Medicina Fetal do Centro de Atenção Integral à Saúde da Mulher (CAISM) da Universidade Estadual de Campinas (Unicamp). Foi realizada a caracterização da amostra estudada e uma análise descritiva dos achados moleculares através da técnica de array CGH. Resultados: Foram incluídos no estudo 50 fetos, dos quais 49 preencheram os critérios de qualidade da técnica. A taxa de detecção de alterações cromossômicas pela técnica de array CGH não detectadas pelo cariótipo convencional foi de 93,7% (45 fetos), e 27% foram consideradas significativas dos pontos de vista citogenético e clínico. Entre os fetos com alterações do número de cópias, 87% apresentaram pelo menos um clone para o qual já estão descritas variações do número de cópias (CNV) em indivíduos fenotipicamente normais. Adicionalmente, a técnica mostrou-se eficaz para o esclarecimento diagnóstico da origem, exata localização e dimensionamento do material adicional encontrado em um feto com anomalia cromossômica estrutural. Conclusões: A caracterização do perfil genômico por array CGH de fetos com defeitos congênitos permitiu complementar o diagnóstico citogenético convencional, aumentando a definição diagnóstica e a identificação de regiões cromossômicas associadas a algumas anomalias congênitas
Abstract: Introduction: A great number of fetuses with congenital defects remain without definitive diagnosis, making difficult the perinatal management, the prognosis establishment and the genetic counseling. The incapacity of detection of short sequence copy number changes, pointed as a probable etiology factor for some congenital defects, is the main limitation of routine G-banding. The Comparative Genomic Hybridization (CGH) overcome this limitation, and also does not require cellular culture or prior knowledge of the involved genomic region. Objective: To evaluate the applicability of the CGH method on fetal material for genomic gains and losses in a group of malformed fetuses. Methods: On a prospective descriptive study, fetal blood samples were collected from malformed fetuses with numerically normal chromosomes at G-banded karyotype, at the Fetal Medicine Unit of the Centro de Atenção Integral à Saúde da Mulher (CAISM) of the Universidade Estadual de Campinas (UNICAMP). Sample characterization and a descriptive analysis of the CGH-based technique results were accomplished. Results: Fifty fetuses were included in this study and 49 were considered optimal according to adopted quality control criteria. The detection rate of fetuses with copy number imbalances not detected by the G-banded karyotype was 93.7% (45 fetuses), with 27% of cytogenetic and clinical significance. Among fetuses with copy number imbalances, 87% presented at least one abnormal clone encompassing CNVs described for phenotipically normal individuals. Additionally, the array CGH showed to be effective for the identification of the additional genetic material origin, with its precise location and size, presented in one fetus with structural chromosomal anomaly. Conclusions: The genomic profile characterization of malformed fetuses through array CGH allowed complementing the conventional cytogenetic diagnosis, obtaining a higher precise diagnosis and the identification of chromosomal regions associated with some congenital anomalies
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Tocoginecologia
Doutor em Tocoginecologia

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23

Pinato, Claudia. "ARRAY-CGH COME ESAME DI PRIMO LIVELLO NELLA DIAGNOSI MOLECOLARE DI RITARDO MENTALE E ANOMALIE CONGENITE." Doctoral thesis, Università degli studi di Padova, 2015. http://hdl.handle.net/11577/3424644.

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The array-CGH technique has emerged in recent years as a powerful tool for the identification of molecular causes underlying complex phenotypes characterized by intellectual disability, autism, epilepsy, psychiatric disorders and multiple congenital anomalies. Over the past 10 years, it has become clear that the conventional cytogenetic analysis is unable to detect rearrangements less than 5-10Mb which can be responsible for these clinical phenotypes. This limit has been exceeded with array-CGH technique which has increased by 15-20% the detection rate of cryptic chromosomal imbalances (deletions or duplications). The possibility to have a genome wide technique with a high resolution led to the proposal, in 2010, by the International Standard Cytogenomic Array (ISCA) Consortium, to use this technique as the first-line test in individuals with intellectual disabilities and congenital anomalies. From studies with microarray technology it has become clear that there are chromosomal regions in which aberrant recombination are particularly frequent, due to the presence of segments with elevated sequence homology, that cause a high degree of genomic instability. Moreover the use of the array-CGH showed the presence in the genome of a large number of structural variations, larger than 1Kb, defined copy number variation or CNVs, that does not always represent a direct cause of disease because they have also been identified in healthy individuals. This complexity in the interpretation of pathogenic CNVs is even more relevant in prenatal diagnosis because it leads to uncertainty in terms of prognosis for the fetal health. For this reason, despite the advantages of the technique, the array-CGH analysis in prenatal diagnosis is considered as a second-line test to be used in association to conventional cytogenetics analysis. In this study were evaluated with array-CGH analysis, 1051 patients with mental and / or developmental disabilities, autism, multiple congenital anomalies and dimorphisms. The main purpose was to verify the presence of cryptic chromosomal rearrangements in order to demonstrate the utility of genomic microarray as first-line test for the characterization of the molecular causes underlying the phenotype of individuals. Then the mechanisms of formation of anomalies were hypothesized by the analysis of the breakpoints, to verify the presence of regions of homology that may have predisposed to the rearrangement. So it was examined whether the mechanism of formation and the clinical significance of the identified CNVs could be related to the pattern of inheritance, the type or the size of the imbalance. The results show that 15.8% of the patients has at least one pathological anomaly or VOUS (variant of uncertain significance) that is likely pathological, and that these are more frequently deletions and CNVs arisen de novo. It was also highlighted that both the clinical significance of CNVs and their mechanism of formation may be related to the size of the imbalance. It was later analyzed the distribution of CNVs in different chromosomes and it was found that in some of them the density of anomalies is greater than the others. The application of the array-CGH in a high number of patients has also allowed to estimate the sensitivity to detect mosaicism, although they have a frequency less than 1% in individuals with learning disabilities. It was observed that the technique is able to detect anomalies present in up to 10% of cells. Finally some fetal samples of chorionic villi and amniotic fluid were analyzed to evaluate the possible use of genomic microarray in prenatal diagnosis. The small number of analyzed samples did not allow us to draw conclusions, but the difficulties in the interpretation of the clinical significance of CNVs, make it a second-line test to be used in association with standard karyotype.
La tecnica di array-CGH si è affermata negli ultimi anni come un potente strumento per l’identificazione delle cause molecolari alla base di fenotipi complessi caratterizzati da disabilità intellettive, autismo, epilessia, disordini psichiatrici e anomalie congenite multiple. Negli ultimi 10 anni, infatti, è emerso sempre più chiaramente che l’analisi citogenetica convenzionale non è in grado di rilevare riarrangiamenti inferiori alle 5-10 Mb che possono essere responsabili di tali fenotipi clinici. Questo limite è stato superato dall’array-CGH che ha aumentato del 15-20% la detection rate di sbilanciamenti cromosomici criptici (delezioni o duplicazioni). La possibilità di avere una tecnica di tipo genome wide ad elevata risoluzione ha portato alla proposta, nel 2010, da parte dell’International Standard Cytogenomic Array (ISCA) Consortium, dell’utilizzo di tale tecnica come esame di primo livello in individui con disabilità intellettive e anomalie congenite. Dagli studi effettuati con tecnologia microarray è risultato evidente che esistono regioni cromosomiche in cui sono particolarmente frequenti ricombinazioni aberranti, dovute alla presenza di segmenti con un’elevata omologia di sequenza che causano un alto grado di instabilità genomica. L’utilizzo dell’array-CGH ha inoltre rivelato la presenza nel genoma di un elevato numero di variazioni strutturali, di dimensioni maggiori di 1Kb, chiamate copy number variations (CNVs), le quali, essendo state identificate anche in individui sani, non sempre rappresentano una causa diretta di malattia. Questa difficoltà nell’interpretazione della patogenicità delle CNVs è ancora più rilevante in diagnosi prenatale poiché si traduce in una incertezza in termini prognostici sulla salute del feto. Per questo motivo, nonostante i vantaggi dati dalla tecnica, l’array-CGH in diagnosi prenatale viene, al momento, considerato un test di secondo livello da utilizzare in associazione all’analisi citogenetica convenzionale. In questo studio sono stati valutati, mediante array-CGH, 1051 pazienti che presentano ritardo mentale e/o dello sviluppo, autismo, anomalie congenite multiple e dimorfismi. L’obiettivo principale è stato quello di verificare la presenza di riarrangiamenti cromosomici criptici in modo da dimostrare l’utilità dell’impiego di microarray genomici come esame di primo livello per la caratterizzazione delle cause molecolari alla base del fenotipo patologico degli individui. Sono stati poi ipotizzati i meccanismi di formazione delle anomalie verificando, mediante l’analisi dei breakpoints, la presenza di regioni di omologia che possano aver predisposto al riarrangiamento. Quindi è stato valutato se il meccanismo di formazione e il significato clinico delle CNVs identificate possano essere correlati al pattern di ereditarietà, al tipo o alle dimensioni dello sbilanciamento. I risultati ottenuti mostrano che il 15.8% dei casi analizzati è portatore di una anomalia patologica o VOUS (variant of uncertain significance) verosimilmente patologica, e che queste sono più frequentemente delezioni e CNVs insorte de novo. È stato inoltre evidenziato che sia il significato clinico delle CNVs sia il loro meccanismo di formazione possano essere correlati alle dimensioni degli sbilanciamenti. È stata successivamente analizzata la distribuzione delle CNVs nei diversi cromosomi ed è emerso che in alcuni di essi la densità di anomalie riscontrate è maggiore rispetto agli altri. L’applicazione dell’array-CGH in un elevato numero di pazienti ha permesso, inoltre, di stimarne la sensibilità nell’identificazione di mosaicismi, sebbene abbiano una frequenza inferiore all’1% in individui con disabilità intellettive. È stato osservato che la tecnica è in grado di rilevare anomalie che coinvolgono anche un numero limitato di cellule, fino al 10%. Infine sono stati analizzati alcuni campioni fetali di villi coriali e liquido amniotico per valutare il possibile utilizzo dei microarray genomici in diagnosi prenatale. Il numero esiguo di campioni analizzati non ci ha permesso di trarre delle conclusioni, tuttavia, per le difficoltà che si riscontrano nell’interpretazione del significato clinico delle CNVs, è da ritenere al momento un test di secondo livello da utilizzare in associazione al cariotipo standard.

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Dimassi, Sarra. "Identification de gènes responsables d'épilepsies de l'enfant." Thesis, Lyon, 2017. http://www.theses.fr/2017LYSE1114.

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L'épilepsie est une affection neurologique chronique qui se définit par la répétition de crises épileptiques, signe de l'hyperactivité paroxystique d'un groupe de neurones corticaux. Ces dernières années, plusieurs gènes responsables d'épilepsies monogéniques ont été mis en évidence. Notre travail avait pour objectif l'identification d'anomalies génétiques responsables ou favorisants certaines formes d'épilepsies de l'enfant. Ce travail est composé de quatre études complémentaires. La première était l'exploration pangénomique d'une cohorte de 47 patients porteurs d'épilepsie à paroxysme rolandique (EPR) par CGH array, à la recherche de variations de nombre de copies (CNV) récurrentes. Nous avons ainsi pu mettre en évidence plusieurs CNVs emportant des gènes impliqués dans l'épilepsie, dont PRRT2 et GRIN2A. La deuxième reposait sur la même approche appliquée à une cohorte de 8 patients tunisiens présentant des spasmes infantiles. Elle a permis d'identifier une délétion 9q34.3 emportant le gène EHMT1, responsable du syndrome de Kleefstra et une duplication 15q13.1, région impliquée dans des troubles du neurodéveloppement. Pour la troisième étude, nous avons comparé deux techniques de capture pour séquençage à haut débit d'un panel de gènes impliqués dans les épilepsies de l'enfant, à partir des échantillons de 24 patients épileptiques. Cette approche nous a permis de mettre au point un logiciel d'analyse de couverture, que nous avons nommé DeCovA. Lors de la dernière étude, nous avons appliqué une stratégie de séquençage d'exome en trio pour explorer 10 patients porteurs des spasmes infantiles. Nous avons ainsi pu mettre en évidence des variants pathogènes de novo chez quatre patients,impliquant les gènes KCNQ2, SCN1A, NR2F1 et ALG13. Nos résultats confirment ainsi la place importante de la génétique et l'intérêt majeur des nouvelles technologies dans l'exploration étiologique des épilepsies de l'enfant
Epilepsy is a chronic neurological disorder characterized by repeated epileptic seizures, a sign of cortical neurons paroxysmal hyperactivity. In recent years, several monogenic genes involved in epilepsy have been identified. The aim of our work is to identify new genetic abnormalities responsible for childhood epilepsies. This work is divided into four complementary studies. First, we searched copy number variation (CNV) by pangenomic exploration of a cohort of 47 patients with Rolandic epilepsy (RE) using CGH array. We identified several CNVs carrying genes involved in epilepsy, including PRRT2 and GRIN2A (genes). Secondly, the same approach was applied to a cohort of 8 Tunisian patients with infantile spasms. It allowed the identification of a 9q34.3 deletion includingEHMT1, implicated in Kleefstra syndrome and a 15q13.1 duplication, known to be involved in neurodevelopment disorders. For the third study, we compared two library-building methods for a gene-targeted panel for the diagnosis of Monogenic childhood epilepsies, in a cohort of 24 epileptic patients. This approach allowed us to develop a coverage analysis software, which we named DeCovA. In the last study, we used a trio-based exome-sequencing approach to look for de novo mutations in 10 patients with infantile spasms. We found de novo pathogenic variants in four patients, involving KCNQ2, SCN1A, NR2F1, and ALG13. Our results confirm the increasing role of genetics and the major interest of new technologies in the etiological exploration of childhood epilepsy

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Baker, Serena F. "Assessment of aCGH Clustering Methodologies." BYU ScholarsArchive, 2010. https://scholarsarchive.byu.edu/etd/2644.

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Array comparative genomic hybridization (aCGH) is a technique for identifying duplications and deletions of DNA at specific locations across a genome. Potential objectives of aCGH analysis are the identification of (1) altered regions for a given subject, (2) altered regions across a set of individuals, and (3) clinically relevant clusters of hybridizations. aCGH analysis can be particularly useful when it identifies previously unknown clusters with clinical relevance. This project focuses on the assessment of existing aCGH clustering methodologies. Three methodologies are considered: hierarchical clustering, weighted clustering of called aCGH data, and clustering based on probabilistic recurrent regions of alteration within subsets of individuals. Assessment is conducted first through the analysis of aCGH data obtained from patients with ovarian cancer and then through simulations. Performance assessment for the data analysis is based on cluster assignment correlation with clinical outcomes (e.g., survival). For each method, 1,000 simulations are summarized with Cohen's kappa coefficient, interpreted as the proportion of correct cluster assignments beyond random chance. Both the data analysis and the simulation results suggest that hierarchical clustering tends to find more clinically relevant clusters when compared to the other methods. Additionally, these clusters are composed of more patients who belong in the clusters to which they are assigned.

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Oliveira, Jakeline Santos. "Determinação das alterações genômicas em pacientes com malformações congênitas." Botucatu, 2018. http://hdl.handle.net/11449/180587.

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Orientador: Danilo Moretti-Ferreira
Resumo: As ACs são alterações visíveis nos cromossomos, classificadas como numéricas e estruturais. Atualmente o grande desafio da genética clínica é classificar e associar a relevância clínica dos desequilíbrios genéticos ao fenótipo dos portadores. O trabalho tem como objetivo principal caracterizar desequilíbrio genômico sem diagnóstico sindrômico previamente descritos pelas técnicas de citogenética clássica, molecular visando apurar os pontos de quebras e genes inseridos na região cromossômica alterada por meio da citogenômica em estudos de casos. Foram feitas análises por citogenética (bandamento GTG), citogenética molecular (FISH) e citogenômica (array-CGH) em três pacientes com malformações congênitas não-sindrômicas para definição diagnóstica e maior conhecimento sobre a correlação genótipo-fenótipo. Foram redigidos estudos de casos de três pacientes portadores de MCs, atraso do desenvolvimento e deficiência intelectual. O primeiro copilado de caso trata-se de paciente do sexo feminino com anomalias esqueléticas, deficiência intelectual e atraso do desenvolvimento. O cariótipo da paciente é 46,XX[11]/47,XX,+mar[9]. A análise de array-CGH revelou dois ganhos/duplicações nas bandas cromossômicas 6p11.2q12 (10.335 Mb de tamanho) e 6q14.1q14.3 (10.765 Mb de tamanho). Por meio da técnica da FISH e os resultados do array-CGH a região duplicada 6q14.1q14.3 encontra-se inserida em um cromossomo marcador, oriundo do cromossomo 6. Os sinais clínicos descritos na paciente foram semelhan... (Resumo completo, clicar acesso eletrônico abaixo)
Abstract: The ACs are visible changes in the chromosomes, classified as numerical and structural. Currently the great challenge of clinical genetics is to classify and associate the clinical relevance of genetic imbalances with the phenotype of the carriers. The main objective of this work is to characterize genomic imbalance without syndromic diagnosis previously described by the classical and molecular cytogenetic techniques, in order to determine the breakpoints and genes inserted in the chromosomal region altered by cytogenetics in case studies. Cytogenetics (GTG banding), molecular cytogenetics (FISH) and cytogenetics (array-CGH) were performed in three patients with non-syndromic congenital malformations for diagnostic definition and greater knowledge on genotype-phenotype correlation. Case studies of three patients with MCs, developmental delay and intellectual disability were written. The first case file is a female patient with skeletal anomalies, intellectual disability and developmental delay. The patient's karyotype is 46, XX [11] / 47, XX, + sea [9]. The array-CGH analysis revealed two gains / doublings in the chromosomal bands 6p11.2q12 (10,335 Mb in size) and 6q14.1q14.3 (10,765 Mb in size). Through the FISH technique and the results of the array-CGH the duplicate region 6q14.1q14.3 is inserted in a chromosome marker, coming from chromosome 6. The clinical signs described in the patient were similar to other patients with duplication of the region 6q14. The genes PGM3, M... (Complete abstract click electronic access below)
Mestre

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RASSU, STEFANIA. "Utilità clinica dell’array-CGH nello studio di pazienti in età pediatrica con Leucemia Linfatica Acuta." Doctoral thesis, Università degli Studi di Cagliari, 2016. http://hdl.handle.net/11584/266634.

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Acute leukemia represent the most common malignancy in children, with the 80% of cases of lymphoblastic type. Most patients with acute lymphocytic leukemia are reported to have acquired chromosomal abnormalities in their leukemic bone marrow cells. Multiple chromosome rearrangements have been described, and their associations with specific clinical, biologic, and prognostic features are well defined. Conventional cytogenetic analysis is critical in the diagnosis of LLA, identifying characteristic chromosomal abnormalities associated with a given prognosis, therein facilitating optimized treatment. We investigated the utility of array comparative genomic hybridization (array-CGH) for detection of chromosomal abnormalities compared to standard clinical evaluation with karyotype and fluorescent in-situ hybridization (FISH). In the present study 19 LLA pediatric bone marrows were analyzed, 12 diagnosis and 7 relapse sample. Array-CGH detected unbalanced chromosome rearrangements in all cases except testicular relapses. The most recurrently altered chromosome regions were 9p ( deletion of CDKN2A/B, JAK2, PAX5 genes) and 21q (amplification of RUNX1 gene). The complementary use of microarray and conventional cytogenetics would allow for more sensitive, comprehensive, and accurate analysis of the underlying genetic profile, with concomitant improvement in prognosis and treatment for pediatric LLA.

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Carracedo, Marsiñach Alma. "Caracterización genética de dos subtipos de tumores mamarios (ER+PR+ vs ER+PR-) mediante la técnica de CGH-array." Doctoral thesis, Universitat Autònoma de Barcelona, 2011. http://hdl.handle.net/10803/79088.

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El desarrollo y la progresión de los cánceres de mama, y especialmente de aquellos que son hormono-dependientes están ampliamente influenciados y determinados por los receptores de estrógenos y progesterona (ER y PR respectivamente). Aproximadamente un 70% de todos los cánceres mamarios son ER+ y más de la mitad también son PR+. Está aceptado ampliamente que el estado del ER es un importante factor predictivo de buena respuesta a las terapias endocrinas (TEs), sin embargo la positividad del ER no es una garantía de la sensibilidad al tratamiento ya que algunos tumores ER+ no presentan una buena respuesta. Las observaciones clínicas indican que los tumores ER+PR- presentan una pobre respuesta a la TE y un fenotipo más agresivo que los ER+PR+. El objetivo de este estudio es identificar diferencias genéticas entre los subtipos de tumores ER+PR+ y ER+PR-. La técnica de CGH-array fue aplicada a 25 tumores ER+PR+ y 23 tumores ER+PR-. Los genes de interés obtenidos a partir del análisis de los resultados de dicha técnica fueron analizados por la técnica de hibridación in situ fluorescente (FISH) en una serie de validación formada por 50 tumores ER+PR+ y 50 tumores ER+PR- localizados en varios arrays de tejidos (TMAs). Como resultado se observó que los tumores ER+PR- tienen un ligero pero diferente perfil genético respecto a los tumores ER+PR+. Los tumores ER+PR- presentaban un mayor perfil genómico aberrante, con los cromosomas 17 y 20 como los más diferentemente alterados con ganancias solapadas, y los cromosomas 3, 8, 9, 14, 17, 21 y 22 como los más diferentemente alterados con pérdidas solapadas respecto al grupo ER+PR+. Las regiones ganadas solapadas 17q23.2-q23.3 y 20q13.12, y las regiones perdidas solapadas 3p21.32-p12.3, 9pter-p13.2, 17pter-p12 y 21tel-q21.1 estaban alteradas de forma estadísticamente significativa en los tumores ER+PR-. Las regiones de pérdida incluyen genes (RASSF1A, FHIT, CDKN2A, TP53 y BTG3) con funciones supresoras de tumores y están involucrados en apoptosis, mitosis, angiogénesis y dispersión celular. Mientras que las regiones de ganancia incluyen genes (MAP3K3, RPS6KB1 y ZNF217) involucrados en el control del ciclo celular, angiogénesis, resistencia a la apoptosis, metástasis y dispersión celular y en la activación de las vías de señalización de la PI3K/Akt/mTOR. Todas estas alteraciones podrían contribuir, al menos en algunos casos, a explicar la mayor inestabilidad genómica, la pérdida de la expresión del PR, el fenotipo más agresivo y la mayor resistencia a las ETs, todo ello tradicionalmente observado en los tumores ER+PR-.
Development and progression of all types of breast cancer, and especially the hormone-dependent ones are widely influenced and determined by estrogen and progesterone receptors (ER and PR, respectively). Approximately 70% of all breast cancers are ER+ and more than half of them are also PR+. It is widely accepted that ER status is a strong predictive factor of good response to endocrine therapy (ET), but ER positivity is not a guarantee of sensitivity to the treatment and some tumors fail to respond. Clinical observations indicate that ER+PR- breast cancers present a poorer response to ET and more aggressive phenotype than ER+PR+ ones. The aim of this study was to identify genetic differences between ER+PR+ and ER+PR- subgroups. An array CGH technique was applied to 25 ER+PR+ breast tumors and 23 ER+PR- ones. Genes of interest were analyzed by Fluorescence in situ hybridization (FISH) in a validation series composed by 50 ER+PR+ tumors and 50 ER+PR- ones on TMAs. As a result, it was observed that ER+PR- breast tumors have a smaller but different genetic profile. ER+PR- group presented a higher genomic aberrant profile with chromosomes 17 and 20 as the most differently altered with overlapped gains, and chromosomes 3, 8, 9, 14, 17, 21 and 22 as the most differently altered with overlapped losses respect to ER+PR+ group. The overlapped gained regions 17q23.2-q23.3 and 20q13.12, and the overlapped lost regions 3p21.32-p12.3, 9pter-p13.2, 17pter-p12 and 21tel-q21.1 were found in a significant way in ER+PR- breast tumors. Significant lost regions included genes (RASSF1A, FHIT, CDKN2A, TP53 and BTG3) with tumor suppressor functions and involved in apoptosis, mitosis, angiogenesis and cell spreading. Significant gained regions included genes (MAP3K3, RPS6KB1 and ZNF217) involved in cell cycle control, angiogenesis, resistance to apoptosis, metastasis and cellular spread, and activation of PI3K/Akt/mTOR pathways. All these alterations could contribute, at least in some cases, to explain the higher genomic instability, loss of PR expression, more aggressive phenotype and higher resistance to ETs, traditionally observed in ER+/PR- tumors.

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Morice-Picard, Fanny. "Etude clinique et génétique de l’albinisme oculocutané : développement d’outils de diagnostic moléculaire et recherche de nouveaux gènes." Thesis, Bordeaux 2, 2013. http://www.theses.fr/2013BOR22101/document.

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Notre travail s’est intéressé à l’albinisme oculocutané en étudiant ses aspects clinico- moléculaires. Malgré l’analyse approfondie des gènes connus d’albinisme oculocutané, 15 % des patients restent sans mutation identifiée indiquant que les mutations sont situées dans des régions géniques non analysées par les techniques classiques de diagnostic moléculaire, ou qu’il existe d’autres gènes d’albinisme oculocutané. Nous avons établi une base de données clinico- biologiques décrivant les caractéristiques de plus de 400 patients analysés. Des outils de diagnostic moléculaire ont été développés à la recherche de mutations situées dans les introns et les régions régulatrices et de réarrangements géniques. Différentes stratégies ont également été utilisées pour rechercher des gènes candidats. La puce à façon a permis l’identification de grands réarrangements dans les gènes TYR, OCA2 et SLC45A2 et un réarrangement complexe du gène OCA2 chez 2 patients non apparentés. L'analyse de gènes candidats nous a permis d'identifier, chez 5 patients non apparentés présentant un albinisme oculocutané non syndromique, des mutations dans le gène SLC24A5, très récemment associé à l’AOC6. Le séquençage d’exome de 6 patients a mis en évidence des gènes candidats pour lesquels des analyses complémentaires sont poursuivies afin de confirmer leur implication dans la pathogenèse de l’AOC.Les résultats de ce travail permettent de redéfinir les aspects cliniques et moléculaires de l’AOC, d’identifier de nouveaux mécanismes moléculaires à l’origine de l’AOC ainsi que des gènes candidats dont la fonction dans le développement pigmentaire reste à élucider. L’identification de nouveaux gènes impliqués dans l’AOC pourrait permettre de mieux comprendre et de mieux prendre en charge les patients avec un AOC
Our work focused on oculocutaneous albinism (OCA) by studying its clinical and molecular aspects. Despite a thorough analysis of the known genes involved in oculocutaneous albinism, 15% of patients remain without diagnostic at the molecular level indicating that mutations are located in unexplored regions and are undetected by standard techniques or that other genes are involved in albinism. We established a clinicomolecular database describing more than 400 patients and developped molecular tools in order to improve molecular diagnostic including a custom high resolution array-CGH dedicated to the four OCA genes (TYR, OCA2, TYRP1 and SLC45A2). We also used different strategies to identify new genes. Array-CGH allows us to detect large deletion in TYR, OCA2 and SLC45A2 and a complexe rearrangement in OCA2 in 2 unrelated patients. We identified, in 5 patients presenting with a non syndromic OCA, mutations in SLC24A5, recently associated with OCA6. Exome sequencing of 6 different patients allows us to identify candidate genes, for which further studies are required to confirm their involvement in OCA pathogenesis. The results of this work allowed us to delineate clinical and genetics aspects of more than 400 OCA patients and to identify new molecular mechanisms leading to OCA and candidates genes for which exact nature of their functions has to be understood. Giving the complexity of pigmentary system development and its regulation, identification of new genes leading to OCA could help to better understand OCA and take care of patients

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Birnbaum, David. "Altérations moléculaires dans l'adénocarcinome du pancréas." Thesis, Aix-Marseille, 2014. http://www.theses.fr/2014AIXM5088.

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Le cancer du pancréas est un problème majeur de santé publique. Le mauvais pronostic de l'adénocarcinome du pancréas est lié à un diagnostic tardif, une progression rapide, et à une mauvaise réponse aux traitements médicaux disponibles. Une caractérisation complète des altérations génétiques moléculaires sont aujourd'hui nécessaires pour permettre une détection plus précoce et identifier/élaborer de nouvelles thérapies ciblées dans le traitement de l'ADK du pancréas. En utilisant la technique d'hybridation génomique comparative, nous avons étudié les altérations du génome de 39 ADK. Plusieurs pertes récurrentes ont été observées, ainsi que des gains de matériel génétique. A partir de ces résultats, nous avons voulu aller plus loin en identifiant les gènes qui pourraient avoir des conséquences au niveau transcriptomique. A partir de données publiques, nous avons comparé les profils d'expression d'ADK (n=419) et de pancréas normal (n=105). Parmi les gènes trouvés amplifiés et/ou gagnés par aCGH, 170 (48%) étaient surexprimés dans les ADK par rapport au tissus normal. Les principales voies de signalisation impliquées touchaient la régulation du cycle cellulaire, les voies TP53 et TGFß. Parmi les gènes délétés en aCGH, 141 (41%) étaient sous exprimés dans les ADK du pancréas par rapport au tissus normal et étaient essentiellement liés au « métabolome » de la cellule pancréatique. Enfin, nous avons identifié une dizaine de gènes dont l'expression pourrait être liée à la survie des patients. Certains de ces gènes pourraient être des candidats à tester en tant que biomarqueurs pronostic ou cibles pour le développement de nouvelles thérapeutiques
Pancreatic adenocarcinoma (PDCA) is a major public health problem in France and worldwide. The inoperability and the poor prognosis of the PDCA are due to late diagnosis, rapid tumor progression (>80% of patients displayed metastases at diagnosis), early recurrences after resection, and poor response to available therapies. Innovative approaches and a comprehensive characterization of molecular genetic alterations are dearly needed to help develop techniques of early detection, identify new molecular targets and devise novel targeted-therapies (Hidalgo, 2010). Using high-resolution array-comparative genomic hybridization (aCGH), we studied the genome alterations of 39 fine-needle aspirations from PDCA. Recurrent losses were observed and comprised several known tumor suppressor genes. We identified frequent genetic gains. With this study, we decided to go one step further by identifying genes that might also be deregulated at the transcriptomic level. We started our analysis with a population of PDCA (n=419) versus normal pancreas (n = 105). Among the 352 genes found amplified and/or gained by aCGH, 170 (48%) were up regulated at the transcriptional level in PDCA compared to normal pancreatic tissues. Major pathways involved were cell cycle, TP53 and TGFß. Among the genes located in regions of losses, 141 (41%) were down regulated in PDCA compared to normal tissues. Furthermore, some genes were found related to a patients' survival With this study, we highlighted novels genes associated to PDCA oncogenesis. Some of those candidates should be further investigated as prognosis markers or as potential targets for new therapeutic approaches

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Joaquim, Tatiana Mozer. "Correlação cariótipo-genótipo-fenótipo de rearranjo cromossômico estrutural familiar envolvendo as regiões 4p e 12q." Universidade de São Paulo, 2016. http://www.teses.usp.br/teses/disponiveis/17/17135/tde-04012017-114707/.

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Rearranjos cromossômicos estruturais estão potencialmente associados ao desenvolvimento de doenças genéticas devido à disrupção, inativação ou alteração da dosagem gênica. O objetivo deste projeto foi realizar a caracterização genômica de duas pacientes e seus familiares portadores de rearranjo cromossômico estrutural envolvendo o braço curto do cromossomo 4 e o braço longo do cromossomo 12, associando técnicas de citogenética clássica (bandamento GTG), citogenética molecular (FISH) e citogenômica (array-CGH), para definição diagnóstica e maior conhecimento sobre os fatores envolvidos na correlação cariótipo-genótipo-fenótipo. Foram avaliados seis indivíduos, duas pacientes, primas em primeiro grau que apresentavam alterações fenotípicas, assim como seus familiares, portadores de translocação aparentemente equilibrada e fenótipo normal. Apesar das duas pacientes apresentarem alteração cromossômica comum, derivativo do cromossomo 4 [der(4)], foram observados achados fenotípicos distintos. A investigação permitiu a definição do diagnóstico de deleção 4p16 e trissomia 12qter para as duas pacientes com fenótipo alterado e cariótipo 46,XX,der(4)t(4;12)(p16;q24.3), a definição precisa dos pontos de quebra em 4p16.3 e 12q24.31->q24.33, assim como a determinação da origem parental do rearranjo e a definição do diagnóstico citogenético final de quatro portadores de translocação aparentemente equilibrada e cariótipo t(4;12)(4pter->4p16.3::2q24.31->12qter;12qter->12q24.31::4p16.3->4pter),direcionando o aconselhamento genético para a família. Nas duas pacientes, a técnica de array-CGH (Plataforma 2x400K, Agilent®) detectou uma diferença sutil de tamanho entre as perdas e ganhos referentes aos cromossomos envolvidos no rearranjo, sendo diagnosticado em P1 uma perda de 2.707.221 pb na citobanda 4p16.3, além de um ganho de 12.405.205 pb em 12q24.31->q24.33. A paciente 2 apresentou uma perda de 2.710.969 pb em 4p16.3 e um ganho de 12.393.885 pb em 12q24.31->q24.33. Ambas as regiões de desequilíbrio genômico incluem genes que podem ser relevantes para manifestação fenotípica observada nas pacientes, entre eles: WHSC1, NELFA, LETM1, FGFRL1 e SPON2. Os resultados da investigação citogenômica indicaram, ainda, a presença de translocação equilibrada nos quatro indivíduos portadores, não sendo detectadas perdas e/ou ganhos genômicos nas regiões dos pontos de quebra cromossômica. Os resultados obtidos na investigação do padrão de metilação dos genes FGFRL1 e SPON2 não permitiram afirmar que uma provável repressão da expressão gênica devido ao imprinting materno e paterno esteja associada às características fenotípicas distintas observadas nas duas pacientes. Embora tenha sido possível a indicação de genes correlacionados ao fenótipo das pacientes, a correlação entre a alteração genética e o fenótipo das mesmas pode depender da ação sinérgica dos mais de 190 genes envolvidos neste rearranjo cromossômico estrutural familiar.
Structural chromosomal rearrangements are potentially associated with the development of genetic disorders due to disruption, inactivation or gene dosage alterations. The objective of this project was to perform the genomic characterization of a familial structural chromosomal rearrangement involving the short arm of chromosome 4 and the long arm of chromosome 12 in two patients and carriers. The experimental approach involved using a combination of classical cytogenetic techniques (GTG banding), molecular cytogenetics (FISH) and cytogenomics (array-CGH), to provide a diagnostic definition and a better understanding of how changes in the karyotype and genotype may be associated with the phenotype. Six individuals were evaluated, two patients with phenotypic abnormalities, as well as the carriers of an apparently balanced 4p;12q translocation with normal phenotypes. Although the two patients showed a common chromosomal abnormality, the derivative chromosome 4 [der (4)], they presented distinct phenotypic findings. The investigation provided a definition of the diagnosis of 4p16 deletion and trisomy 12qter for the two patients with abnormal phenotypes and a karyotype 46,XX,der(4)t(4;12)(p16;q24.3). In addition a precise definition of the breakpoints at 4p16.3 and 12q24.31->q24.33, and the parental origin of the rearrangement was determined. A precise definition of the cytogenetic diagnosis of four carriers with an apparently balanced translocation and karyotype t(4;12)(4pter->4p16.3::2q24.31->12qter; 12qter 12q24.31->4pter::4p16.3), facilitated the genetic counseling for the family. In both patients, the array-CGH technique (2x400K Platform, Agilent®) detected a subtle difference in size between losses and gains in the chromosomal regions involved in the rearrangement. Patient 1 presented a loss of 2,707,221 bp in the cytoband 4p16.3, and a gain of 12,405,205 bp in 12q24.31->q24.33. Patient 2 had a loss of 2,710,969 bp in 4p16.3 and a gain of 12,393,885 bp in 12q24.31 -> q24.33. Both regions of genomic imbalance included genes that may be relevant to phenotypic findings observed in our patients, including: WHSC1, NELFA, LETM1, FGFRL1 and SPON2. Genomic findings also confirmed the presence of a balanced translocation in four carriers, with no genomic losses and/or gains in the regions of chromosome breakpoints. The results of the investigation of the methylation pattern of FGFRL1 and SPON2 genes could not demonstrate that repression of gene expression due to maternal and paternal imprinting was associated with the distinct phenotypes observed in the two patients. Although it has been possible to indicate genes related to the phenotype of the patients, the correlation between the genetic alteration and phenotype may depend on the synergistic action of multiple genes from more than the 190 involved in this familial chromosomal rearrangement.

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Grzesiuk, Juliana Dourado. "Investigação genômica de pacientes inférteis com oligozoospermia." Universidade de São Paulo, 2016. http://www.teses.usp.br/teses/disponiveis/17/17135/tde-30032017-162247/.

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A infertilidade afeta aproximadamente 15% dos casais, sendo atualmente reconhecido o envolvimento de fatores masculinos em metade dos casos. Alterações nas análises seminais são detectadas na maioria dos homens inférteis e a mais frequente é a baixa concentração de espermatozoides no ejaculado, conhecida como oligozoospermia. Vários estudos mostram uma forte relação entre fatores genéticos e a infertilidade, incluindo alterações cromossômicas e microdeleções do cromossomo Y, porém as causas da oligozoospermia ainda permanecem obscuras. O desenvolvimento de novas tecnologias de investigação vem possibilitando a detecção de alterações a nível genômico, como mutações e variações no número de cópias (CNVs). O presente trabalho teve por objetivo a caracterização genômica de homens com oligozoospermia sem causa definida, visando estabelecer correlação entre alterações no número de cópias e perdas de heterozigosidade (LOHs) e o fenótipo de infertilidade. Foram selecionados 18 pacientes após rigorosa avaliação clínica e investigação do histórico reprodutivo, sendo excluídos pacientes portadores de alterações cromossômicas e portadores de microdeleções do cromossomo Y. Seis homens comprovadamente férteis foram selecionados para o grupo controle. A investigação genômica de ambos os grupos, amostral e controle, foi realizada pela técnica de hibridação genômica comparativa em microarranjos (aCGH) utilizando a plataforma de resolução 180K (Agilent®,US), analisada pelo software Nexus 8.0. Foram detectadas alterações possivelmente patogênicas no cromossomo Y, no cromossomo X e em autossomos. Um ganho na região de AZFc envolvendo apenas os genes DAZ1 e DAZ4 foi detectado em nove pacientes e em quatro controles, sendo classificado como alteração benigna. Porém, alterações na região de AZFc possivelmente relacionadas ao fenótipo de oligozoospermia foram detectadas em três pacientes e incluíram extensas duplicações e deleções envolvendo, entre outros genes, as quatro cópias do gene DAZ. Após comparação de regiões selecionadas com a literatura e com diferentes bancos de dados genéticos, sugerimos que os genes PLEC, SPATC1, COL1A1, MOV10L1, SYCE3 e ODF3B possam estar associados a alterações na produção espermática. Adicionalmente, entre os doze miRNAs presentes em regiões de LOH possivelmente relacionadas ao fenótipo de infertilidade, dez têm como alvo genes com funções relacionadas à espermatogênese e reprodução humana. Estudos adicionais a nível de expressão e sequenciamento gênico são necessários para confirmar a correlação entre o genótipo e o fenótipo de oligozoospermia.
Infertility affects about 15% of the couples, and it is currently recognized, that male factors are involved in about 50% of cases. Changes in seminal parameters are detected in most infertile men and the most common alteration, known as oligozoospermia, is a low concentration of sperm in the ejaculate. Several studies show a strong relationship between genetic factors and infertility, including chromosomal abnormalities and microdeletions of Y chromosome, however, the causes of oligozoospermia remain unclear. The development of new research technologies has allowed the detection of changes at genomic levels, such as mutations and copy number variations (CNVs). This study aimed to perform a genomic characterization of patients with idiopathic oligozoospermia to determine whether there is a correlation between changes of copy number and losses of heterozygosity (LOHs) in relation to the phenotype of infertility. Eighteen patients were selected for the cases after rigorous clinical examination and investigation of their reproductive history. Patients with chromosomal abnormalities or microdeletions of the Y chromosome were excluded. Six proven fertile men comprised the control group. Genomic investigation of both groups was performed by microarray comparative genomic hybridization (aCGH) using 4X180K platform (Agilent, US) analysed by Nexus 8.0 software. Potential pathogenic changes were detected on Y chromosome, as well as on the X and autosome chromosomes. A gain in AZFc region involving only DAZ1 and DAZ4 genes was detected in nine patients and four controls, and was considered as benign. However, changes in AZFc region, that could be related to the oligozoospermia phenotype were detected in three patients. These changes included extensive duplications and deletions involving the four copies of the DAZ gene together with copy number changes affecting other genes. After comparing the selected regions with the literature and with different databases, we suggest that changes such as LOH affecting PLEC, SPATC1, COL1A1, MOV10L1, SYCE3 and ODF3B genes may influence sperm production. Our analysis indicates that, ten out of the twelve miRNAs present in LOH regions could be involved in the infertility phenotype and could have target genes with functions related to spermatogenesis and human reproduction. Additional studies involving gene sequencing and expression analysis are needed to confirm the the correlation between the genotype and oligozoospermia phenotype.

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Rousseau, Audrey. "Approche diagnostique et pronostique dans les tumeurs gliales : de l'immunohistochimie à l'hybridation génomique comparative." Paris 6, 2007. http://www.theses.fr/2007PA066256.

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Les 2 principaux types de tumeurs gliales sont les gliomes diffus et les épendymomes. L’étude immunohistochimique de l’expression de marqueurs de lignage astrocytaire (GFAP, YKL-40, ApoE) et oligodendroglial (Olig2, Sox10, ASCL1, NKX2-2) a porté sur 55 gliomes diffus. Les molécules GFAP, YKL-40, ApoE, ASCL1 et NKX2-2 permettaient de distinguer les oligodendrogliomes des astrocytomes et représentent donc des marqueurs tumoraux prometteurs. Les anomalies les plus fréquentes détectées en CGH array dans 45 épendymomes étaient un gain des chromosomes (chr) 7, 9, 12, 15, et une perte du chr22. L’association de ces 5 altérations était corrélée aux épendymomes médullaires tandis que les formes myxopapillaires du cône présentaient un profil génomique caractéristique, associant un gain des chr5, 7, 9, 16 et 18. L’identification d’altérations génomiques spécifiques d’un siège tumoral donné suggère que les épendymomes issus de régions différentes représentent des maladies génétiquement distinctes

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Potluri, Keerti. "Improving DNA quality using FFPE tissues for Array Comparative Genomic Hybridization to find Single Nucleotide Polymorphisms (SNPs) in Melanoma." Wright State University / OhioLINK, 2015. http://rave.ohiolink.edu/etdc/view?acc_num=wright1438267267.

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PANZERI, ELENA. "Mutazioni nel gene tau associate ad instabilità cromosomica: un nuovo ruolo della proteina tau. Studio nell'uomo." Doctoral thesis, Università degli Studi di Milano-Bicocca, 2010. http://hdl.handle.net/10281/17980.

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Tau is a microtubule associated protein that promotes assembly and stabilization of cytoskeleton microtubules (MT) required for morphogenesis and axonal transport. It is mostly expressed in neuronal and glial cells but it is also present in non-neuronal cells such as fibroblast, lymphocytes and lymphoblasts where it can play further functions. Six major tau isoforms are expressed in adult human brain, as a result of alternative mRNA splicing of exons 2, 3 and 10 of MAPT, a gene located on 17q21.1 where it occupies over 100 kb. Tau is subjected to different post-translational modifications, among which phosphorylation is known to negatively regulate its ability to bind to MT. In fact, in cycling cells, tau has a low level of phosphorylation and is mostly bound to MT during interphase, while at the onset of mitosis, it became highly phosphorylated reducing its affinity to MT and permitting their dinamic instability required for the rapid mitotic movements. Abnormal hyperphosphorylation of tau causes disruption of cytoskeleton and deposition of hyperphosphorylated tau filaments in neuron and glial cells, resulting in cerebral atrophy and dementia, hallmarks of Alzheimer’s disease as well as of the so-called tauopathies, adult-onset diseases that can be of genetic origin and caused by dominant mutations in MAPT, among which frontotemporal dementia (FTD) is the most frequent. Missense, deletion and silent mutations have been reported in the coding region of MAPT, with additional mutations in the intronic region close to the 5’ splice site of exon 10. Very recently it has been proposed an involvement of tau in chromosome stability, based on its localization both in the interphasic nucleus and along mitotic chromosomes and on the presence of several chromosome and chromatin aberrations in fibroblasts and lymphocytes of patients affected by FTD carrying P301L mutation (Rossi G et al., 2008). In the present study, in order to establish whether chromosomal aberrations are a general phenomenon in tauopathies rather than a unique feature of the P301L mutation, we investigated these alterations in somatic cells of patients bearing different MAPT mutations with standard (karyotype) and molecular (array-CGH) cytogenetic techniques. Furthermore, since the observed chromosome instability resembled some features of what is seen in syndromes caused by mutations in genes involved in the DNA repair systems, we tested the response of mutated lymphoblastoid cell lines to genotoxic agents, able to induce single- and double-strand breaks. Lymphocytes and fibroblasts of patients showed a significantly higher level of structural aberrations and total aneuploidy compared to controls. We observed also a high level of premature chromatid division (pcd), that is correlated with chromosome segregation defects and aneuploidies, and abnormal nuclei. Condensations defects were also widely present in the patients and not in the controls, thus a role of tau in condensation or decondensation of chromatin may be hypothesized. Analyzing lymphoblastoid cell lines treated with genotoxic agents, we have not yet demonstrated an involvement of the protein in DNA repair systems, but these results need to be confirmed. Our findings indicate that tau may be involved in chromosome segregation and stabilization of chromatin structure most likely through interactions with both microtubules and chromatin itself. The occurrence of numerical chromosome aberrations in FTD patients is likely due, at least in part, to misfunction of tau as a MT associated protein, since the mutations studied decrease the binding of the protein to MT, producing an instable spindle. Fragility and alterations in chromosome structure due to mutated nuclear tau may slowly accumulate in postmitotic cells and lead to abnormal regulation of gene expression with subsequent neuronal damage and death. The evidence of a role of tau in the functioning of mitotic spindle and stability of chromatin points to this protein as a possible component of the complex system guarding genome stability.

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FUSCO, ILEANA. "Search for genomic micro-rearrangements through array-CGH in patients with Intellectual Disability and Multiple Pituitary Hormone Deficiency, with standard and custom-design array platforms." Doctoral thesis, Università del Piemonte Orientale, 2015. http://hdl.handle.net/11579/115568.

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Guediche, Narjes. "Caractérisation par cytogénétique moléculaire des chromosomes marqueurs surnuméraires et étude de leur implication dans le développement et la reproduction humaine." Thesis, Paris 11, 2012. http://www.theses.fr/2012PA11T024.

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Les chromosomes marqueurs surnuméraires (CMS) sont définis comme des chromosomes de structure anormale qui ne peuvent pas être identifiés ni caractérisés de façon non ambigüe par les techniques de cytogénétique conventionnelle seules et qui sont de taille égale ou plus petits qu’un chromosome 20 de la même métaphase. La prévalence de cette anomalie chromosomique est estimée à 0,071% en post-natal, 0,075% en diagnostic prénatal et 0,288% chez les patients atteints de retard mental et/ou du développement. Chez les patients infertiles, la fréquence des CMS est estimée à 0,122% et varie selon le sexe. Les CMS sont sans conséquence clinique dans 70% des cas. Dans un tiers des cas, ils peuvent être responsables de nombreuses anomalies du développement et de la reproduction humaine. A ce jour, il existe très peu d’études de caractérisation des CMS permettant une cartographie précise des gènes présents. Dans ce travail, nous avons étudié une série de huit CMS par cytogénétique conventionnelle, FISH (fluorescent in situ hybridization) et CGH array (array comparative genomic hybridization). Nous avons établi une cartographie des gènes présents dans ces CMS. L’étude de la relation génotype-phénotype des patients nous a permis de proposer l’implication de certains gènes candidats dans des anomalies du développement et de la reproduction humaine. Notre étude de l’implication des CMS dans les anomalies du développement humain s’est basée sur l’étude cytogénétique de trois fœtus. Les deux premiers fœtus étaient porteurs d’un CMS(20) en anneau. Le sujet 1 présentait un retard de croissance intra-utérin (RCIU) et une dysmorphie cranio-faciale. Le sujet 2 n’avait pas d’anomalies particulières à part une obésité diagnostiquée à l’âge de quatre mois. La taille de ces CMS(20) était de 13,6 Mb pour le sujet 1 et 4,8 Mb pour le sujet 2. Le gène SSTR4 présent sur le CMS(20) du sujet 2 code pour un récepteur de la somatostatine. Cette hormone joue un rôle dans le comportement alimentaire. Le troisième fœtus présentait un hygroma kystique et un RCIU associé à un CMS(13) néocentromérique. Les explorations par CGH array ont révélé un gain chromosomique de la région 13q21.1qter de 39 Mb contenant 80 gènes dont GPC5, GPC6, SPRY2, EFNB2, SOX1 et DZIP1. La modification d’expression de ces gènes est susceptible d’être responsable du phénotype des sujets étudiés.Notre étude de l’implication des CMS dans les anomalies de la reproduction humaine s’est basée sur l’étude cytogénétique de cinq patients qui présentaient des troubles de la fertilité (anomalies de la spermatogenèse, insuffisance ovarienne prématurée, syndrome des ovaires polykystiques et fausses couches spontanées). Les CMS explorés par CGH array correspondaient aux régions chromosomiques 15q11.2 (3,6 Mb), 21p11.2 (0,266 Mb), 6p11.2q12 (9 Mb) et 20p11.21 (3,3 Mb). Le CMS d’une des patientes ne contenait pas d’euchromatine et une autre patiente était porteuse de deux CMS d’origines chromosomiques différentes. Plusieurs gènes candidats (POTE B, BAGE et THBD) ont pu être identifiés. La modification de leur expression ainsi que des effets mécaniques ou biochimiques perturbant la méiose et la maturation des gamètes pourraient être responsables des troubles de la fertilité observés chez ces patients. L’étude des CMS par CGH array nous a permis de caractériser précisément les points de cassure des CMS, leur taille et leur composition génétique afin de cartographier les gènes présents dans les CMS et d’établir des relations entre le génotype et le phénotype des patients
Small supernumerary marker chromosomes (sSMC) are defined as structurally abnormal chromosomes which cannot be unambiguously identified or characterized by conventional banding cytogenetic techniques alone and are generally equal in size or smaller than a chromosome 20 of the same metaphase spread. sSMC frequency is estimated at 0.071% in postnatal cases, 0.075% in prenatal cases, and 0.288% for mentally and/or development retarded patients. In infertile patients cases, sSMC frequency is estimated at 0.122% and is different in male (0.165%) and female infertility (0.022%). sSMC have no clinical consequences in 70% of the cases. In one third of the cases, they can be responsible for various human development and reproduction anomalies. To date, only a few studies precisely characterizing the sSMC contents have been performed.In this study, we used conventional cytogenetics, FISH (fluorescent in situ hybridization) and array CGH (array comparative genomic hybridization) to characterize eight sSMC and to precisely localize the genes included. The study of the genotype-phenotype correlations of the patients led us to suppose the implication of some candidate genes in human development and reproduction anomalies.Our study of the implication of sSMC in human development anomalies was based on the cytogenetic study of three fetuses. The first two fetuses carried a ring sSMC(20). Case 1 presented with intrauterine growth retardation and craniofacial dysmorphism. Case 2 had a normal phenotype except for obesity diagnosed at the age of four months. The size of these sSMC(20) was approximately 13,6 Mb for case 1 and 4,8 Mb for case 2. The SSTR4 gene located on the case 2 sSMC(20) is coding for one of the somatostatin receptor. This hormone has multiple effects on variable cells and is implicated in the regulation of food behavior, which could explain the obesity of case 2. Case 3 presented with intrauterine growth retardation and a cystic hygroma associated with a neocentric sSMC(13). Array CGH investigations showed a 32.9 Mb gain from 13q31.1 to 13qter region containing 80 genes. Among these genes, six genes could be involved in the phenotype of the proband (GPC5, GPC6, SPRY2, EFNB2, SOX1 and DZIP1). The expression modification of these genes could be responsible for the phenotype observed.Our study of the implication of sSMC in human reproduction anomalies was based on the cytogenetic study of five patients presenting fertility troubles (spermatogenesis impairment, ovarian insufficiency, polycystic ovary syndrome and repeated abortions). The sSMC explored by array CGH corresponded to the 15q11.2 region (3.6 Mb), the 21p11.2 region (0.266 Mb), the 6p11.2q12 region (9 Mb) and 20p11.21 region (3.3 Mb). The sSMC of one of the patients did not contain euchromatin and one patient carried two sSMC derived from two different chromosomes. Among the genes present on the sSMC, some candidate genes (POTE B, BAGE and THBD) have been identified. The modification of their expression and mechanical or biochemical effects of the sSMC impeding meiosis could be directly responsible for the fertility trouble observed in these patients. A detailed molecular cytogenetic investigation using array CGH allowed us to precisely characterize the chromosomal breakpoints, the size and genomic constitution of sSMC. This study may be helpful to address genotype–phenotype correlations and for medical and genetic counseling

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Smith, Marissa B. "A description of genetic counselors' views and current practice with regard to the use of array-CGH for prenatal diagnosis." Cleveland, Ohio : Case Western Reserve University, 2009. http://rave.ohiolink.edu/etdc/view?acc%5Fnum=case1246977726.

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Redon, Sylvia. "Variations structurales du génome et pathologies humaines : recherche de nouveaux marqueurs génétiques impliqués dans les ischémies cérébrales du sujet jeune." Thesis, Brest, 2012. http://www.theses.fr/2012BRES0006.

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Ce travail de thèse a permis, dans un premier temps, de mettre en évidence de nouveaux grandsréarrangements dans trois pathologies étudiées au laboratoire : la mucoviscidose, la pancréatitechronique et l’hémochromatose. En particulier, ces travaux ont permis de trouver de nouveaux CNVs(Copy Number Variations) pathologiques dans le gène CFTR (Cystic Fibrosis Transmembraneconductance Regulator), de mieux comprendre les mécanismes d’un remaniement complexe dansPRSS1 (Protease Serine 1) et d’aider à caractériser finement un réarrangement dans HFE(Hemochromatosis). Ces études ont donc servi de preuve de concept pour l’utilisation de puces à ADN àl’échelle d’un gène et dans des zones difficiles car riches en séquences répétées.Dans un second temps, la recherche de facteurs de susceptibilité génétiques aux infarctus cérébraux(AICs) du sujet jeune a été réalisée chez 168 cas et 200 témoins âgés de moins de 40 ans. Dans notrepopulation, l’hypertension, les migraines, le tabac, et la prise de stupéfiants sont des facteurs de risqueimportants, multipliant respectivement par 35, 3,8, 4 et 2,8 le risque d’AIC. Notre étude pangénomiquepar CGH-array (Comparative Genomic Hybridization-array) a mis en évidence 98 régionspolymorphiques dans le génome humain. Parmi elles, la délétion d’une partie du gène NOTCH2, pourraitjouer un rôle protecteur dans la survenue des AICs (OR=0,11 [0,01-0,87] ; p=0,013) mais qui ne dépassepas le seuil fixé par la correction de Bonferroni). Ce travail a également mis en évidence environ 400CNVs rares, dont deux récurrents chez les cas, l'un portant les gènes CRELD2 (cysteine-rich with EGFlikedomains 2) et AGL12 (asparagine-linked glycosylation 12, alpha-1, 6-mannosyltransferase) (p=0,02)et le deuxième situé en 5’ du gène VBP1 (von Hippel-Lindau binding protein 1) (p=0,04). Enfin, uneapproche gènes candidats a été effectuée sur les gènes NOTCH2 et ALOX5AP (5-lipoxygenaseactivating protein) sans donner de résultats significatifs. Ceci a également été réalisé sur les mutationsprincipales de trois gènes de la coagulation (Facteur II, Facteur V Leiden et MTHFR). Une associationsignificative a été mise en évidence entre la C677T du gène MTHFR (5,10-methyltetrahydrofolate) et lesinfarctus cérébraux du sujet jeune (OR=2,39, p=0,02 pour le génotype TT). Ce travail de thèse a permisde confirmer l’existence de facteurs de risque environnementaux et génétiques déjà connus mais surtoutd’émettre de nouvelles hypothèses génétiques dans la survenue des AICs du sujet jeune
The use of locus-specific array-CGH (Comparative Genomic Hybridization) has allowed us to detect largerearrangements in three pathologies of our laboratory: cystic fibrosis, chronic pancreatitis andhemochromatosis. We successfully observed new pathological CNV (Copy Number Variations) in theCFTR (Cystic Fibrosis Transmembrane conductance Regulator) gene and characterized complex eventsin PRSS1 (Protease Serine 1) and HFE (Hemochromatosis) genes, showing that the use of thistechnique is possible even in regions with high sequence homologies.We also confirmed that hypertension, migraine, tobacco and drugs are high significant risk factors forischemic strokes (IS) in young population (under 40 years) (OR=35, 3.8, 4 and 2.8, respectively). Then,we tried to identify new genetic susceptibility loci using a pangenomic approach. Among the 98 copynumber polymorphisms (CNP) observed, an interstitial NOTCH2 deletion is candidate for a protective rolein IS (OR=0.11 [0.01-0.87] ; p=0.013 before Bonferonni correction). We also observed approximately 400uncommon CNV, two of them being particularly reccurent in patients: a 22q13.31 duplication containingCRELD2 (cysteine-rich with EGF-like domains 2) and AGL12 (asparagine-linked glycosylation 12, alpha-1, 6-mannosyltransferase) genes (p=0.02) and a Xq28 deletion localised in the 5’ region of the VBP1 (vonHippel-Lindau binding protein 1) gene (p=0.04). We also applied a candidate-gene approach onNOTCH2, ALOX5AP (5-lipoxygenase activating protein) and coagulation genes (Factor II, Factor VLeiden and MTHFR). A significant association was found for the C677T in the MTHFR gene (5,10-methyltetrahydrofolate) and young ischemic strokes (OR=2.39, p=0.02 for TT genotype). In conclusion,this study confirmed the implication of environmental and genetic factors in ischemic strokes before 40years and suggests new genetic risk factors for IS

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Praulich, Inka [Verfasser]. "Untersuchung genomischer Alterationen bei myelodysplastischen Syndromen (MDS) und juveniler myelomonozytärer Leukämie (JMML) im Kindesalter mittels hochauflösender array-CGH / Inka Praulich." Hannover : Technische Informationsbibliothek und Universitätsbibliothek Hannover (TIB), 2013. http://d-nb.info/1035436914/34.

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Baretto, Nathacha. "Análise de CNVs e indicação clínica em indivíduos com deficiência intelectual e outros distúrbios do desenvolvimento diagnosticados por CGH array." reponame:Repositório Institucional da UFSC, 2015. https://repositorio.ufsc.br/xmlui/handle/123456789/132484.

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Dissertação (mestrado) - Universidade Federal de Santa Catarina, Centro de Ciências Biológicas, Programa de Pós-Graduação em Biologia Celular e do Desenvolvimento, Florianópolis, 2015.
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A deficiência intelectual (DI) é caracterizada por limitações significativas no funcionamento intelectual e no comportamento adaptativo, origina-se antes dos 18 anos de idade e afeta cerca de 1 a 3% da população do Mundo e 1,37% da população brasileira. As causas etiológicas da DI são variadas e de difícil identificação, devido a sua heterogeneidade. Entre as causas genéticas, a variação no número de cópias (CNVs) de trechos do DNA no genoma vem sendo amplamente estudada em distúrbios do desenvolvimento, através da técnica de hibridização comparativa por arrays (CGH array). CNVs são comumente classificadas como benignas (CNVs comum), de significado clínico incerto (raras), potencialmente patogênicas (raras) e patogênicas (raras). O presente estudo teve como objetivo avaliar as CNVs encontradas em indivíduos com DI e distúrbios do desenvolvimento e interpretar a sua contribuição para o aparecimento do fenótipo. Foram analisados 109 resultados de exames de CGH array, realizados pelo Laboratório de Genética Humana Neurogene, em Santa Catarina, com as informações clínicas e fenotípicas fornecidas através do preenchimento de questionários pelos médicos responsáveis pela solicitação dos exames. Houve uma prevalência de dois terços do sexo masculino na população estudada, sendo que os principais fenótipos que levaram a solicitação da investigação foram ADNPM, dismorfias de cabeça e face e DI. Oesclarecimento diagnóstico foi maior para DI severa, DI leve e hiperatividade associada a outros distúrbios. Os indivíduos testados apresentaram um total de 276 CNVs (187 microduplicações e 89 microdeleções); 81,2% benignas, 7,2% de significado incerto, 9,4% patogênicas e 2,2% potencialmente patogênicas. Os cromossomos 18, 19 e 21 apresentaram o menor número de CNVs. Não foi encontrada nenhuma CNV rara nos cromossomos 5, 10, 12, 19, 21 e Y. Seis casos potencialmente patogênicos foram descritos em mais detalhe, um desses casos representa uma deleção em 3 p13-p14.1 de 1.9MB, que confirma que a haploinsuficiência do gene MITF é suficiente para causar surdez congênita. Este estudo vem destacar a importância do CGH array para o diagnóstico de distúrbios do desenvolvimento inclusive para que seja inserido nos programas de saúde pública como um primeiro teste diagnóstico a ser ofertado pelo SUS.

Abstract : Intellectual disability (ID) is characterized by significant limitations in intellectual functioning and in adaptive behavior. It originates before the age of 18, and affects about 1-3% of the world population and about 1.37% of the Brazilian population (CENSO 2010). ID has different levels of severity: mild, moderate, severe or profound, depending on the degree of intellectual impairment combined with adaptive behaviour. The etiological causes of ID are varied and difficult to identify due to clinical and genetic heterogeneity. Among the genetic causes, copy number variation (CNV) of DNA stretches in the genome has been widely studied in developmental disorders. CNVs are commonly classified as benign (common CNVs), of uncertain clinical significance (rare), potentially pathogenic (rare), and pathogenic (rare). This study aims to evaluate the CNVs found in patients with ID and developmental disorders and classify them according to their contribution to the appearance of the phenotype. We analyzed 109 results for CGH array investigation to obtain the phenotype of each individual. We observed: (1) a higher prevalence of males in the population studied, (2) that the major phenotypes observed were developmental delay, dysmorphic face and head and ID, and (3) the higher diagnostic rates were obtained for individuals with severe ID, mild ID, and hyperactivity. An overall of 276 CNVs (187 microduplications and 89 microdeletions) were observed. Ofthese changes 81.2% were benign, 7.2% of uncertain significance, 2.2% potentially pathogenic and 9.4% pathogenic. Chromosomes 18, 19 and 21 had the lowest number of CNVs. Rare CNVs were not observed in chromosomes 5, 10, 12, 19, 21 and Y. Six cases of potentially pathogenic CNVS were studied in more detail, and in one of these cases a deletion of 1.9MB in 3p13-p14.1 was observed, confirming that the haploinsufficiency of the MITF gene is sufficient to cause congenital deafness. This study highlights the importance of the investigation of CNVs in the diagnosis of developmental disorders, underscoring the importance to include array CGH as first investigation for ID in the public health system (SUS).

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Rigon, C. "Caratterizzazione molecolare mediante array-CGH e origine parentale di anomalie cromosomiche strutturali in pazienti con ritardo mentale/psicomotorio/autismo e/o anomalie comportamentali." Doctoral thesis, Università degli studi di Padova, 2011. http://hdl.handle.net/11577/3425326.

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The study of structural chromosomal abnormalities has emerged in recent years as a powerful tool for the identification of molecular causes underlying disorders responsible for genomic complex phenotypes such as mental retardation, autism, epilepsy, psychiatric disorders and multiple congenital anomalies. For nearly 10 years it has increasingly become clear that the conventional cytogenetic analysis is unable to detect chromosomal abnormalities less than 5-10 Mb, that may be associated with phenotypic abnormalities and mental retardation. This limit has been exceeded in recent years by the introduction of a molecular cytogenetic technique, array-CGH, which allows complete and precise analysis of DNA copy number variations and allows to evaluate with high specificity many chromosomal regions in order to detect chromosomal imbalances. Over the last decade with the introduction of genome wide array, it became clear that the molecular mechanisms at the basis of the genomic disorders are related to rearrangements of some regions of the genome, particularly predisposed to aberrant recombination. Several studies have indeed revealed the presence of some segments (SINE, LINE, LCRS) that cause a high degree of genomic instability leading to chromosomal rearrangements. The parental origin of chromosome abnormalities is of considerable interest because it may help to understand their formation mechanism. Many studies show that male gametogenesis appears susceptible to the formation of structural chromosome abnormalities. This is generally attributed to the much larger number of premeiotic cell divisions undergone by male germ cells in comparison with female germ cells. In this study 66 subjects with mental retardation and / or development, autism, multiple congenital anomalies and dimorphisms were assessed by array CGH, in order to verify the presence of cryptic chromosomal rearrangements and to characterize more precisely the chromosomal abnormalities identified by high-resolution chromosome examination. It was then analyzed the parental origin of chromosome rearrangements by use of polymorphic markers (RFLP or STR) to determine whether there is a different mutation rate in both sexes. Finally the breakpoints were analyzed to verify the presence of homologous regions which may predisposed to rearrangements. The results obtained in this study show that 16% of patients with clinical signs and a normal karyotype have a cryptic deletion / duplication. In 20% of patients with karyotypic alterations, previously identified by standard cytogenetic, array- CGH detected other abnormalities. The analysis of the breakpoints revealed the presence of homologous regions that may have predisposed the rearrangement confirming that the architecture of the genome play a major role for genomic instability causing chromosomal rearrangements. In contrast to the literature there are no significant differences between the sexes in the formation of chromosomal rearrangements.
Lo studio delle anomalie cromosomiche strutturali si è affermato negli ultimi anni come un potente mezzo per l’identificazione delle cause molecolari alla base di disordini genomici responsabili di quadri fenotipici complessi quali ritardo mentale, autismo, epilessia, disordini psichiatrici e anomalie congenite multiple. Da circa 10 anni è emerso sempre più chiaramente che l’analisi citogenetica convenzionale non è in grado di rilevare anomalie cromosomiche inferiori a 5-10 Mb che, seppur di dimensioni submicroscopiche, possono associarsi a ritardo mentale e anomalie fenotipiche. Questo limite è stato da qualche anno superato dall’introduzione di una tecnica di citogenetica molecolare, l’array-CGH, che permette un’analisi completa e precisa delle variazioni del numero di copie delle sequenze di DNA e consente di valutare contemporaneamente e con alta specificità più regioni cromosomiche in modo da poter evidenziare sbilanciamenti. Nell'ultimo decennio con l'introduzione di array genome wide, è risultato evidente che i meccanismi molecolari alla base dei disordini genomici sono correlati a riarrangiamenti di particolari regioni del genoma, suscettibili più di altre ad andare incontro a ricombinazioni aberranti. Diversi studi hanno evidenziato infatti la presenza di alcuni segmenti (sequenze SINE, LINE, LCRs) che causano un alto grado di instabilità genomica portando a riarrangiamenti cromosomici. L’origine parentale delle anomalie cromosomiche è di considerevole interesse in quanto potrebbe aiutare a capire il loro meccanismo di formazione. Gli studi fatti fino ad ora riportano nella gametogenesi maschile c’è una maggiore tendenza alla formazione di riarrangiamenti cromosomici conseguente a un maggior numero di divisioni premeiotiche delle cellule germinali maschili rispetto a quelle femminili. In questo studio sono stati valutati mediante array CGH 66 soggetti che presentano ritardo mentale e/o dello sviluppo, autismo, anomalie congenite multiple e dimorfismi con lo scopo di verificare la presenza di riarrangiamenti criptici e caratterizzare in modo più preciso le anomalie identificate grazie all’esame cromosomico ad alta definizione. E’ stata quindi determinata l'origine parentale dei riarrangiamenti mediante l'utilizzo di marcatori polimorfici (STR o RFLP) per definire se esiste un diverso tasso di mutazione nei due sessi; infine sono stati analizzati i breakpoints per verificare la presenza di regioni di omologia che possano aver predisposto al riarrangiamento. I risultati ottenuti in questo studio mostrano che il 16% dei pazienti con fenotipo patologico e cariotipo normale è portatore di una delezione/duplicazione criptica; inoltre nel 20 % dei pazienti in cui erano state precedentemente individuate alterazioni del cariotipo, l’array-CGH ha evidenziato ulteriori anomalie. L’analisi dei breakpoints ha evidenziato la presenza di regioni di omologia che possono aver favorito il riarrangiamento confermando che l’architettura del genoma agisce come catalizzatore dell’instabilità cromosomica causando riarrangiamenti genomici, tuttavia al contrario di quanto riportato in letteratura non ci sono differenze significative tra i due sessi nella formazione di riarrangiamenti cromosomici.

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Bestetti, I. "GENOME WIDE ANALYSIS IN A COHORT OF 46,XX PATIENTS AFFECTED BY AN EXTREME PHENOTYPE OF PRIMARY OVARIAN INSUFFICIENCY: AN EFFICIENT TOOL TO IDENTIFY NEW GENES INVOLVED IN OOCYTE MATURATION AND DIFFERENTIATION." Doctoral thesis, Università degli Studi di Milano, 2016. http://hdl.handle.net/2434/363638.

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Primary Ovarian Insufficiency (POI) is a heterogeneous group of disorders with an incidence of 1:10,000 women by age 20, 1:1,000 by age 30, 1:100 by age 40. POI describes the progression toward the cessation of ovarian function and can occur in the most serious form as primary amenorrhea (PA), with absent pubertal development and/or ovarian dysgenesis (OD), or in milder phenotype with post-pubertal onset and secondary amenorrhea (SA). Several are the etiological causes that may induce ovarian dysfunction, among which the genetic component is considered prevalent (as supported by the occurrence of families with more than one affected women and the existence of several idiopathic cases) but highly heterogeneous. 46,XX non-syndromic women showing the most severe phenotype, characterized by the absence of pubertal development with PA and OD, are very rare but the search for genetic variations in this extreme phenotype may be more effective in identifying novel pathogenic mechanisms. Hence, using high resolution array-CGH we searched for rare high-penetrant CNVs involving genes essential for ovarian function in a cohort of 67 46,XX non-syndromic patients affected by PA, namely 53 sporadic (79.1%) and 14 familial (20.9%) cases. 28 out of 67 women resulted positive to array-CGH analysis because having at least one rare “ovarian” CNV: a total number of 32 CNVs involving 37 ovarian genes was selected. Population from Database of Genomic Variants (DGV) was used to evaluate the rarity of POI CNVs, but it does not match to the ideal set of controls for POI disease (neither age nor gender of DGV controls are known). Thus, to better understand the CNVs contribution in disease onset, the rare “ovarian” CNVs found in patients according to DGV were searched in an ad hoc control cohort, previously screened by array-CGH, consisting in 140 healthy women with normal reproductive life and physiological menopause. 28 out of 32 rare “ovarian” CNVs detected in patients were not found in the control group thus supporting their role in the POI’s pathogenesis. Moreover, to evaluate the presence of a significant enrichment in ovarian genes in the POI group, array-CGH of the ad hoc control cohort were analyzed with the same approach adopted for patients cohort and 49 CNVs involving 54 ovarian genes were selected. Several statistical analyses were performed comparing patients’ to controls’ data and revealed no significant differences. Nevertheless, the CNVs found in the POI cohort containing ovarian genes are more harmful respect to the CNVs identified in the controls cohort. The 37 genes perturbed or possibly perturbed by POI CNVs are implicated in several ovarian processes (e.g., regulation of cytoskeleton dynamics for oocytes asymmetric division, maintenance of oocytes genomic integrity, ovarian differentiation, follicular development, and meiotic resumption), thus supporting their involvement in POI etiology. Validation and characterization of selected CNVs, as well as the study of a possible gene perturbation at mRNA level, was also crucial in order to perform a correct genotype-phenotype correlation and to propose new candidate genes for POI disease (e.g. TP63, VLDLR). 39 out of 67 women resulted negative for rare “ovarian” CNVs (58.2%) suggesting to combine different genomic approaches in order to increase the detection rate of the disorder. Hence, 17 out of 67 collected patients, were submitted to a preliminary WES analysis searching for rare SNVs in a total of 191 genes selected from array-CGH data, and literature regarding POI and ovary. The WES preliminary analysis allowed to confirm the importance of some array CGH new candidate genes in POI onset (e.g. VLDLR) and the complex heterogeneity of POI. The combination of these molecular evidences, with major or minor contribution, might have been at the basis of POI supporting the existence of a disease genetic model characterized by oligogenic heterozygosity (i.e., the simultaneous presence in a single patient of multiple heterozygous quantitative variants/rare mutations, both de novo and/or inherited, affecting multiple genes). The present approach using both array-CGH and WES techniques, resulted an efficient tool to identify rare variants (CNVs and SNVs) involving both genes already reported in POI, and new candidate genes with a role in oocyte maturation and differentiation. The results of this study are promising to expand the knowledge about the molecular pathways involved in POI pathogenesis and probably provide the basis for a more accurate genetic diagnosis of POI patients.

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Santos, Mauren Fernanda Moller dos. "Estudo genético de síndromes associadas à obesidade." Universidade de São Paulo, 2014. http://www.teses.usp.br/teses/disponiveis/41/41131/tde-26082014-155459/.

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A obesidade se tornou uma das maiores preocupações de saúde pública. É um distúrbio neuroendócrino, no qual fatores ambientais e genéticos agem em conjunto, levando ao excesso de armazenamento de energia na forma de gordura corporal. A síndrome de Prader-Willi (PWS) é a mais freqüente das síndromes que possui a obesidade como uma de suas características, com incidência de 1:25.000 nascimentos. É caracterizada por hipotonia neonatal com dificuldade de sucção, atraso do desenvolvimento neuropsicomotor (DNPM), hiperfagia, obesidade, baixa estatura em adolescentes, mãos e pés pequenos, hipogonadismo, distúrbios do sono, características faciais dismórficas, deficiência intelectual leve a moderada e comportamento obsessivo-compulsivo. Pacientes com atraso do DNPM e/ou dificuldade de aprendizado, distúrbios de comportamento, obesidade e/ou hiperfagia, com teste negativo para PWS, foram estudados com plataformas de SNP array, “The GeneChip® Mapping 500K Set” da Affymetrix, ou array-CGH, CytoSure ISCA 4x180k da OGT, para identificar genes relacionados a obesidade e hiperfagia, assim como, novas regiões genômicas implicadas na etiologia de síndromes genéticas associadas à obesidade. Dentre os 31 pacientes estudados, oito apresentaram variações de número de cópias (CNVs) em seu genoma: deleção em 1p22.1p21.2; deleção em 3q25.33q26.1 e deleção em 13q31.2q32.1; duplicação em 7q36.2; deleção em 8p23.3p23.1 e duplicação em 12p13.33p13.31; duplicação 16p13.11p12.3; duplicação em 17q11.2; deleção em 20p12.1; duplicação em 21q22.13. Duas dessas alterações foram herdadas de pais fenotipicamente normais. Algumas dessas CNVs sobrepõem regiões genômicas previamente relacionadas com obesidade, incluindo a microdeleção de 1p21.3 e as duplicações dos cromossomos 12 e 21. Identificamos genes anteriormente descritos como associados à obesidade (PTBP2, DPYD, MIR137, GNB3 e PPM1L), ou possivelmente envolvidos com este fenótipo (HTR5A e KCNJ6), mapeados em várias dessas CNVs. Além disso, os genes RNF135, NF1, DPP6, GPC5, DYRK1A e MACROD2 são os prováveis causadores da deficiência intelectual, atraso do desenvolvimento neuropsicomotor, dificuldades de aprendizagem, distúrbios de comportamento e outras características clínicas encontrados nos pacientes. O diagnóstico e prognóstico dos pacientes e o Aconselhamento Genético aos pais e familiares é fornecido
Obesity has become a major concern for public health. It is a neuroendocrine disorder, in which genetic and environmental factors act together, leading to excessive storage of energy as fat. Prader-Willi syndrome (PWS) is the main obesity-related syndrome with a birth incidence of 1:25,000. It is characterized by neonatal hypotonia, poor sucking, developmental delay, hyperphagia, obesity, short stature in adolescents, small hands and feet, hypogonadism, sleep disturbance, dysmorphic facial features, mild to moderate intellectual disability and obsessive-compulsive behavior. Patients with psychomotor developmental delay and/or learning disabilities, behavior disorders, obesity and/or hyperphagia, who tested negative for PWS, were studied by chromosomal microarray analysis, including the SNP-based platform “The GeneChip® Mapping 500K Set” (Affymetrix), and the array-CGH platform “CytoSure ISCA 4x180k (OGT)”, to identify genes related to hyperphagia and obesity, as well as new genomic regions implicated in the etiology of genetic syndromes associated with obesity. Of 31 patients studied, eight had copy number variants (CNVs) in the genome: 1p22.1p21.2 deletion; 3q25.33q26.1 deletion and 13q31.2q32.1 deletion; 7q36.2 duplication; 8p23.3p23.1 deletion and 12p13.33p13.31 duplication; 16p13.11p12.3 duplication; 17q11.2 duplicaton; 20p12.1 deletion; 21q22.13 duplication. Two of these CNVs were inherited from an unaffected father. Some of these CNVs overlap genomic regions that have previously been related to obesity, including the 1p21.3 microdeletion and the duplications of chromosomes 12 and 21. Furthermore, we identified genes previously described as associated with obesity (PTBP2, DPYD, MIR137, GNB3 and PPM1L), or possibly involved with this phenotype (HTR5A and KCNJ6), mapped to several of these CNVs. In addition, the genes RNF135, NF1, DPP6, GPC5, DYRK1A and MACROD2 are likely implicated in intellectual disability, developmental delay, learning disabilities, behavioral disorders and other clinical features found in patients. The diagnosis and prognosis of patients and genetic counseling to parents and families is provided

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Zaccaria, Julia [Verfasser], and Tina [Akademischer Betreuer] Buchholz. "Pilotstudie zum Nachweis von Aneuploidien in der Frühgravidität an transzervikal gewonnenen Trophoblasten mittels Array-CGH-Analyse / Julia Zaccaria ; Betreuer: Tina Buchholz." München : Universitätsbibliothek der Ludwig-Maximilians-Universität, 2018. http://d-nb.info/1175381713/34.

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Zaccaria, Julia Patrizia [Verfasser], and Tina [Akademischer Betreuer] Buchholz. "Pilotstudie zum Nachweis von Aneuploidien in der Frühgravidität an transzervikal gewonnenen Trophoblasten mittels Array-CGH-Analyse / Julia Zaccaria ; Betreuer: Tina Buchholz." München : Universitätsbibliothek der Ludwig-Maximilians-Universität, 2018. http://d-nb.info/1175381713/34.

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Chêne, Gautier. "Les dysplasies tubo-ovariennes : contribution à une meilleure compréhension de la carcinogenèse ovarienne." Thesis, Clermont-Ferrand 1, 2011. http://www.theses.fr/2011CLF1MM09/document.

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Introduction : l’analyse histopathologique des pièces d’annexectomie prophylactique(pBSO) pour risque génétique (mutations BRCA) a pu révéler des anomalies cytologiques etarchitecturales interprétées comme potentiellement pré-cancéreuses et dénommées « dysplasieovarienne et tubaire ». Nous proposons d’étudier les aspects morphologiques,immunohistochimiques et moléculaires des dysplasies tubo-ovariennes.Matériels & Méthodes : l’analyse morphologique a été réalisée dans un premier grouped’annexectomies après stimulation de l’ovulation (protocole de Fécondation in vitro).L’évaluation morphologique et immunohistochimique (expression de Ki67, p53, Bcl2, PAX2et ALDH1) a par la suite concerné 111pBSO, 42 annexectomies exposées au Tamoxifène et116 témoins non cancéreux et spontanément fertiles (nBSO). Les analyses ont été réaliséespar deux pathologistes en aveugle. Les cellules épithéliales d’intérêt ovariennes et tubairesprovenant du groupe pBSO ont été microdisséquées par laser ; l’ADN extrait a été étudié parhybridation génomique comparative (CGH array). La longueur des télomères a été évaluéepar PCR quantitative en temps réel.Résultats : les scores moyens de dysplasie ovarienne et tubaire étaient significativement plusélevés dans les groupes stimulation de l’ovulation et génétique par rapport aux témoins. Seulle score de dysplasie tubaire était supérieur aux témoins pour le groupe Tamoxifène. Onretrouvait une surexpression de ALDH1 dans les groupes pBSO et tBSO alors que Ki67, p53,bcl2 et PAX2 étaient faiblement exprimés dans les groupes pBSO et tBSO. D’ailleurs,l’expression d’ALDH1 était faible dans l’épithélium non dysplasique, forte dans la dysplasieet constamment faible dans les carcinomes occultes. De subtiles altérations génomiques et desraccourcissements télomériques ont été mis en évidence au niveau des dysplasies génétiques.Conclusions : les scores élevés de dysplasie, la forte expression d’ALDH1 et les altérationsmoléculaires provenant du groupe à risque génétique pourraient supporter le conceptd’instabilité génétique. La dysplasie tubo-ovarienne pourrait être une étape importante etprécoce de la carcinogenèse ovarienne. Nos résultats suggèrent également qu’un certainnombre de cancers de l’ovaire pourrait avoir pour origine la trompe de Fallope. Le marqueurde cellules souches ALDH1 pourrait constituer une cible dans la prévention et le diagnosticprécoce des cancers de l’ovaire
Background: Histopathological examination of material from prophylactic salpingooophorectomies(pBSO) performed in patients at genetic risk has revealed frequentabnormalities interpreted as possible pre-cancerous “ovarian and tubal dysplasia” lesions. Wesought to study the morphologic, immunohistochemical and molecular features in ovarian andtubal dysplasiaMaterials and methods : Morphologic analysis was evaluated in 37 oophorectomies afterovulation induction (iBSO). Morphologic features and immunohistochemical expressionpatterns of Ki-67, p53, Bcl2, PAX2 and ALDH1 (an enzyme significantly associated withearly-stage ovarian cancer) were evaluated in 111 pBSO, 42 salpingo-oophorectomiesexposed with Tamosifen (tBSO) and 116 normal salpingo-oophorectomies (nBSO).Representative slides from formalin-fixed, paraffin-embedded tissue blocks were read blindlyby two gynaecological pathologists. Tubal and ovarian epitheliums from normal anddysplastic tissues (from pBSO) were laser microdissected and studied by comparativegenomic hybridization (array CGH). Telomere length was performed using quantitative realtimePCR.Results: Mean ovarian and tubal dysplasia score were significantly higher in the ovulationinduction group and in the genetic risk group than in controls. Only tubal dysplasia score wassignificantly higher in the Tamoxifen group. Increased ALDH1 expression was observed inpBSO and tBSO compared with nBSO whereas expression patterns of Ki67, p53, bcl2 andPAX2 were low at moderate in pBSO and tBSO group. Interestingly, ALDH1 expression waslow in non dysplastic epithelium, high in dysplasia and constantly low in the carcinoma foundincidentally on pBSO. Subtle genomic alterations were found in the dysplastic ovarian andtubal epitheliums. Shortened telomeres were found in dysplastic tissues from pBSO.Conclusion: The increased dysplasia scores, the strong ALDH1 expression and the geneticalterations in ovaries and tubes from BRCA 1/2 carriers could support the genetic instabilityof dysplasia and might be consistent with progression towards neoplastic transformation andcould justify the use of the term “dysplasia”. Ovarian and tubal dysplasia may be a premalignant,non-invasive histopathological abnormalitie that could be an important step in11early ovarian neoplasia. Our results suggest that a greater percentage of ovarian cancers thanoriginally thought may actually have a fallopian origin with metastasis to the ovary. The stemcell marker ALDH1 activation in pBSO could be considered as a target for early diagnosisand prevention of ovarian cancers

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Huynh, Minh Tuan. "Apport de l'analyse chromosomique sur différents microréseaux d'ADN dans l'identification de nouvelles mutations et la caractérisation de gènes candidats impliqués dans la déficience intellectuelle." Thesis, Université de Lorraine, 2013. http://www.theses.fr/2013LORR0129/document.

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Anomalies de structure du génome et Déficience Intellectuelle : Recherche des gènes candidats de Déficience intellectuelle en utilisant l'analyse chromosomique sur microréseau d'ADN pangénomique 180K et l'analyse chromosomique sur microréseau d'ADN de haute résolution 1M ciblée des gènes candidats de Déficience intellectuelle. L'analyse chromosomique sur microréseau d'ADN (ACM) de haute résolution est une innovation technologique puissante afin de détecter les aberrations chromosomiques concernant les variations du nombre de copies. En utilisant l'ACM 180K, l'ACM 1M et la PCR quantitative, nous avons identifié les 5 variations du nombre de copies (CNV) intragéniques pathogènes de novo impliquant les gènes : RUNX1T1, KIAA1468, FABP7, ZEB2 (syndrome de Mowat-Wilson) et ANKRD11 (syndrome de KBG). Les 5 patients ayant une DI et une dysmorphie faciale. L'ACM 180K a révélé une délétion d'une taille de 92 kb emportant le gène KIAA1468 candidat pour le syndrome de West chez un enfant de 3 ans présentant une DI sévère et une encéphalopathie épileptique infantile précoce. Le criblage des mutations du gène KIAA1468 a été réalisé chez 35 patients atteints de syndrome de West. Un variant intronique c.2761-7 T>C et un variant faux-sens hérité de la mère c.3547 G>A avec signification clinque inconnue ont été identifiés. En utilisant des approches par l'ACM 1M de haute résolution chez 45 patients atteints de DI idiopathique modérée à sévère, un seul CNV causal a été identifié, une délétion intragénique d'une taille de 28.37 kb du gène ZEB2. Notre étude confirme une fréquence très faible des délétions/duplications intragéniques avec la détection d'une seule aberration chromosomique (1/45). En conclusion, si la fréquence des mutations ponctuelles est élevée, nous avons également souligné l'application de la technique de séquençage à haut débit avec un rendement diagnostique jusqu'à 45%-55% des cas de DI sévère idiopathique chez lesquels aucun CNV n'a été détecté sur ACM
Chromosomal structural abnormalities and Intellectual Disability : In search of intellectual disability candidate genes by using pangenomic comparative genomic hybridization 180 K and high resolution comparative genomic hybridization 1M targeting intellectual disability candidate gene.High resolution microarray-based comparative genomic hybridization (a-CGH) has been a powerful technical innovation in order to detect submicroscopic chromosomal aberrations related to copy number variations. By using a-CGH 180K, 1M high resolution a-CGH and quantitative PCR, we have identified 5 pathogenic intragenic copy number variations (CNVs) de novo : RUNX1T1, KIAA1468, FABP7, ZEB2 (Mowat-Wilson syndrome) and ANKRD11 (KBG syndrome). All five patients had intellectual disability (ID) and facial dysmorphism. Interestingly, a-CGH 180K has revealed a 92 kb deletion of a candidate gene KIAA1468 for West syndrome in a 3 year-old boy with severe ID and early infantile epileptic encephalopathy. Mutational screening for candidate gene KIAA1468 was performed in 35 patients with West syndrome. An intronic variant c.2761-7 T>C and a non synonymous maternally inherited variant c.3547 G>A with unknown clinical significance were identified. By using 1M high-resolution a-CGH approach in 45 patients presenting moderate to severe idiopathic ID, only one causal CNV was identified, a 28.37 kb intragenic ZEB2 deletion. Our study has confirmed the low frequency of intragenic deletion/duplication with the detection of only one chromosomal aberration (1/45). In conclusion, providing that the high frequency of intragenic point mutation, we also stressed the application of next-generation sequencing technology with 45-55% diagnostic yield in patients with idiopathic severe ID in case of no apparent CNV(s) on high-resolution a-CGH

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Costa, Claudia Ismania Samogy. "Copy number variations (CNVs) in Brazilian patients with autism spectrum disorder (ASD)." Universidade de São Paulo, 2018. http://www.teses.usp.br/teses/disponiveis/41/41131/tde-20092018-124809/.

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Autism Spectrum Disorder (ASD) is a heterogeneous group of neurodevelopmental disorders that affects about 1% of the worldwide population and has a strong genetic component. Stereotyped behavior and restricted interests, as well as problems of social interaction and communication characterize ASD. Moreover, in 10% of cases, ASD occurs as a secondary condition in addition to a syndrome, such as Phelan-McDermid syndrome (PMS), which is associated with a great clinical variability. Among genetic factors, copy number variations (CNVs) are one of the most important. However, the clinical significance of many CNVs remains nuclear and there is an underrepresentation of small CNVs associated with ASD in the literature. In this context, this project aimed to 1) characterize large and small CNVs in Brazilian patients with ASD using an array-CGH previously customized in our laboratory. 2) Clinically and genetically describe a cohort of Brazilian patients with PMS, as well as to determine the frequency of this syndrome among Brazilian patients with ASD and other neurodevelopmental disorders. In result, we 1) further validated the customized array-CGH, 2) provided additional evidence of association with ASD for 27 candidate genes, 3) described 15 CNVs never reported in the literature in association with this disorder, 4) presented evidence that around 70% of CNVs found in our cohort are not polymorphism of our population and 5) reinforced the idea of shared molecular pathways among different neurodevelopmental disorders. In addition, we described for the first time a Brazilian cohort of patients with PMS and contributed to the molecular and clinical characterization of this syndrome. We also provided additional evidence of genotype-phenotype association with regard to the presence of renal problems and speech status in patients with PMS and estimated the frequency of this syndrome among Brazilian patients with ASD and intellectual disability (syndromic or not). With these results, we hope to contribute to better understand the ASD and PMS etiology, especially in our population
O Transtorno do Espectro Autista (TEA) corresponde ao um grupo heterogêneo de alterações no neurodesenvolvimento que afeta cerca de 1% da população mundial e apresenta um forte componente genético. O TEA é caracterizado pela presença de comportamento estereotipado e interesses restritos, além de problemas de interação social e comunicação. Além disso, em 10% dos casos, o TEA ocorre como uma condição secundária somada a uma síndrome. Um exemplo é a síndrome de Phelan-McDermid (PMS), associada a uma grande variabilidade clínica. Dentre os fatores genéticos, as variações no número de cópias (CNVs) são um dos mais importantes. No entanto, o significado clínico de muitas CNVs permanece incerto, além de haver juma sub-representação de CNVs pequenas associadas ao TEA na literatura. Dentro deste contexto, este projeto teve como objetivos 1) caracterizar CNVs grandes e pequenas em pacientes brasileiros com TEA utilizando uma lâmina de array-CGH previamente customizada no Laboratório de Genética do Desenvolvimento - USP. 2) descrever clínica e geneticamente uma casuística de pacientes brasileiros com PMS, bem como determinar a frequência desta síndrome em pacientes com TEA e com outras alterações de neurodesenvolvimento. Como resultados, nós 1) validamos a lâmina customizada, 2) fornecemos evidencia adicional de associação com o TEA para 27 genes, 3) descrevemos 15 CNVs nunca reportadas em associação com o transtorno 4) apresentamos evidências de que cerca de 70% das CNVs encontradas em nossa coorte não são polimorfismo de nossa população e 5) reforçamos a ideia de vias moleculares compartilhadas entre diferentes alterações do neurodesenvolvimento. Além disso, descrevemos pela primeira vez uma casuística brasileira de pacientes com PMS e contribuímos para a síndrome. Fornecemos evidência adicional de associação genótipo-fenótipo no que diz respeito à presença de problemas renais e status de fala em pacientes com PMS e estimamos a frequência da síndrome entre pacientes brasileiros com TEA e com deficiência intelectual (sindrômica ou não). Com estes resultados, esperamos ter contribuído para o entendimento da etiologia tanto do TEA, quanto da PMS, sobretudo na nossa população

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Sandgren, Johanna. "Array-based Genomic and Epigenomic Studies in Healthy Individuals and Endocrine Tumours." Doctoral thesis, Uppsala universitet, Institutionen för kirurgiska vetenskaper, 2010. http://urn.kb.se/resolve?urn=urn:nbn:se:uu:diva-129533.

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Abstract:

The human genome is a dynamic structure, recently recognized to present with significant large-scale structural variation. DNA-copy number changes represent one common type of such variation and is found both between individuals and within the somatic cells of the same individual, especially in disease states like cancer. Apart from DNA-rearrangements, epigenomic changes are increasingly acknowledged as important events in the maintenance of genomic integrity. In this thesis, different array-based methods have been applied for global genomic and epigenomic profiling of both normal and cancer cells. In paper I, a genomic microarray was established and used to determine DNA-copy number variants (CNVs) in a cohort of 76 healthy individuals from three ethnic populations. We identified 315 CNV regions that in total encompassed ~3,5% of the genome. In paper II, the array was utilized to discover CNVs within several differentiated tissues from the same subject. Six variants were identified providing evidence for somatic mosaicism. In paper III and IV we studied pheochromocytomas and paragangliomas, rare endocrine tumours that most often present as benign and sporadic with unclear genetic/epigenetic cause. Genome-wide DNA-copy number analysis of 53 benign and malignant samples in paper III revealed numerous common and novel chromosomal regions of losses and gains. High frequencies of relatively small overlapping regions of deletions were detected on chromosome 1p arm, encompassing several candidate tumour suppressor genes. In paper IV, an epigenomic map for two histone modifications associated with silent (H3K27me3) or active (H3K4me3) gene transcription, was generated for one malignant pheochromocytoma. Integrated analysis of global histone methylation, copy number alterations and gene expression data aided in the identification of candidate tumour genes. In conclusion, the performed studies have contributed to gain knowledge of CNVs in healthy individuals, and identified regions and genes which are likely associated with the development and progression of pheochromocytoma/paraganglioma.

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