No description available.

Assessing hormone receptors (the estrogen and progesterone receptors) and the human epidermal growth factor receptor 2 (HER2) to guide clinical decision making revolutionized treatment for breast cancer patients. However, in the years since these biomarkers were first incorporated into routine clinical care, only a few others have been validated as clinically useful in guiding adjuvant chemotherapy decisions and are recommended by the American Society of Clinical Oncology (ASCO) for patients with hormone-positive breast cancer. For patients with triple-negative breast cancer (TNBC), which lacks hormone and HER2 receptors, not any of these biomarkers are recommended by ASCO due to insufficient evidence that they meaningfully improve clinical outcomes. Breast cancer is the second-leading cause of cancer-related death among women in the US, indicating an unmet need to improve treatments, which can be accomplished in part by identifying and validating novel predictive and prognostic biomarkers that yield actionable information about the clinical course of breast cancers, especially TNBCs. A major obstacle to improving outcomes for breast cancer patients is intratumor heterogeneity (ITH), which can be extensive in breast cancer and drives treatment resistance and relapse. I envision that assaying drivers of ITH can inform clinicians about which breast tumors may be intrinsically more aggressive and carry a greater risk of breast cancer-related morbidity and mortality. My research, presented here, primarily focuses on testing the impact of drivers of ITH (namely, centrosome amplification [CA], the clustering protein KIFC1, and mitotic propensity and its drivers) on clinical outcomes in breast cancer in multivariable models as well as the correlates of in vitro efficacy of centrosome declustering drugs (which can selectively eliminate cancer cells with CA). Collectively, these studies reveal gene signatures and immunohistochemical biomarkers that are independent predictors of aggressive breast cancer course and rational strategies to optimize targeted therapy to combat cancer cells exhibiting CA, thereby contributing to the literature on the development of precision medicine for breast cancer patients, including TNBC patients.

It is known that age-related changes impacting multiple organ systems occur earlier in people with Down syndrome (Ds), but the biological basis underlying this trisomy 21-associated propensity for premature aging is poorly understood. Given that the trisomic/normal cells from people with mosaic Ds (mDs) are identical with regards to environmental exposures and genes (except for chromosome 21 copy number), comparisons of these isogenic trisomic/disomic cells allow one to “unmask” the cellular consequences of trisomy 21 by removing extraneous factors. The primary aim of this study was to determine if trisomy 21 results in an increase in the acquisition of age-related somatic chromosomal changes. To meet this aim, chromosome-specific telomere lengths, senescence-associated distension of satellites (SADS), and chromosomal instability frequencies were compared between the isogenic trisomic/disomic cells of people with mDs ranging from 1 to 44 years of age. Chromosome-specific telomere lengths were quantified using a Q-FISH (pantelomeric probe) method. The average trisomic cell telomere length (3.609 mean, +/- 0.082 SE) was significantly less than the average disomic cell telomere length (3.888 +/- 0.083) (n=28; p

With 50,310 related deaths this year, colorectal cancer (CRC) has emerged as the second largest cause of cancer related deaths among Americans. While 70 million Americans are considered at-risk of developing CRC, it is highly curable if detected early. Cohesin proteins, which hold sister chromatids together during replication, have emerged as a potential biomarker in multiple cancer lines. Because of their probable role in DNA replication, DNA repair, chromatin nano-architecture, and gene expression, this paper assessed whether cohesion proteins could be used as a potential biomarker for colorectal cancer risk stratification. While cohesin protein mutations have been reported in different cancers and involved in chromosomal instability, its role in early cancer formation has yet to be observed. Using immunohistochemical and Quantitative Real Time PCR analysis, this thesis assessed the protein and RNA expression levels of cohesin proteins SA-1, NIPBL, and SMC3 from human biopsies at different stages and locations of colorectal cancer development. The results showed that SA-1, a structural cohesion subunit, was significantly (p<0.01) down regulated in cancerous compared to normal tissue. The SA-1 protein was also down regulated in the involved mucosa adjacent to CRC polyps. The cohesion loading protein, NIPBL, was also significantly (p<0.01) under expressed in cancerous versus normal tissue. The RNA expression analysis of rectal mucosa showed that SMC3 and SA-1 was over expressed two fold in patients harboring hyperplastic and adenomous polyps, giving evidence that cohesin proteins are differentially expressed throughout the field of carcinogenesis. Our results demonstrate for the first time that cohesion dysregulation is an early event in human colorectal cancer development and may serve as an important biomarker of field carcinogenesis.

No description available.

Orientador: Carmen Silvia Bertuzzo / Dissertação (mestrado) - Universidade Estadual de Campinas, Faculdade de Ciencias Medicas / Made available in DSpace on 2018-08-07T00:23:33Z (GMT). No. of bitstreams: 1 Amstalden_LucilaGobby_M.pdf: 4346952 bytes, checksum: 99d5f1ab2aae2aeb65d633016bc318a8 (MD5) Previous issue date: 2006 / Resumo: A Anemia de Fanconi (AF) é uma doença caracterizada por múltiplas anomalias congênitas, progressiva falha da medula óssea e alto risco para desenvolvimento de câncer. E denominada também de Síndrome da Instabilidade Cromossômica devido ao fato de suas células apresentarem hipersensibilidade a agentes indutores de quebras cromossômicas. A mais importante das características clínicas é a manifestação hematológica. A incidência da anemia aplástíca, Síndrome Mieloplásica e Leucemia Mielóide Aguda é a maior resposável pela morbidade e mortalidade na AF A incidência da AF em todo o mundo é de. aproximadamente, 3 por milhão. No Brasil não há dados sobre a prevalência da doença. Foram descobertos 12 grupos de complementação e descobertos, até o momento. 11 genes relacionados ao distúrbio. São eles: FANCA. B, C, Dl, D2, E. F G, I, J, L e M. O trabalho teve como objetivo geral a análise das mutações principais (IVS8+2A>G, IVS11+lOC, IVS3+1G>C e 1794J803dellO) do gene FANCG em pacientes com quadro clínico compatível com AF. Foram analisados 38 indivíduos por meio da técnica de PCR associada à digestão e triagem por SSCP e subseqüente seqüenciamento. Nós encontramos um homozigoto para a mutação IVS8+2A>G e uma variante neutra (H482H). Concluímos com nosso estudo que, há uma heterogeneidade molecular em nosso meio; o DEB teste não é 100% eficaz na detecção de indivíduos com AF; o Teste de Complementação deve ser introduzido o quanto antes em nosso país para auxiliar no direcionamento da pesquisa para um determinado gene e minimizar os casos em que não há a confirmação de diagnóstico e, por último, há a necessidade de um Registro Brasileiro para AF com o objetivo de recolher informações clinicas e genéticas de indivíduos com o distúrbio. / Abstract: Fanconi anaemia (FA) is an autosomal recessive disease characterised by congenital abnormalities, progressive bone marrow failure and high risk of developing cancer. It's called Chromosomal Instability Syndrome due to the fact of cells presents hipersensibility to DNA cross-linking agents like mitomycin C and diepoxybutane. The most important clinical feature is hematologic. The incidence of aplastic anemia, myelodysplastic syndrome and acute myeloide leukaemia is the most important cause of morbidity and mortality in FA. The incidence of FA is approximately three per million and the heterozygote frequency is estimated at 1 in 300 in Europe and United States. In Brazil there's not data about prevalence of FA. It was discovered at least 12 complementation groups and eleven gene have been cloned: FANCA, B, C, DJ, D2, E, F, G, I, J, L eM. The study had as general objective the analysis of the main mutation (IVS8-2A>G, TVSll+lOC, IVS3+1G>C e 1794J803dell0) of FANCG gene in patients with clinical features of FA. It was analysed 38 patients through the test polymerase chain reaction (PCR) associated with digestion and mutation screening by SSCP with posterior sequencing. Molecular analysis found a homozygote to rVS8+2A>G and a neutral variant (H482H). We concluded that there's a molecular heterogeneity in our region; it's necessary to introduce the use of complementary tests in Brazil, in order to address the molecular analysis and at last, it's necessary a Brazilian Fanconi Anemia Registry (BFAR) to receive clinical and genetics information of AF patients. / Mestrado / Ciencias Biomedicas / Mestre em Ciências Médicas

Orientador: Carmen Silvia Bertuzzo / Dissertação (mestrado) - Universidade Estadual de Campinas, Faculdade de Ciencias Medicas / Made available in DSpace on 2018-08-12T11:46:19Z (GMT). No. of bitstreams: 1 Goncalves_ClaudiaEstela_M.pdf: 1883480 bytes, checksum: 5c9ad07b778588f38de1f644ecd18847 (MD5) Previous issue date: 2008 / Resumo: A Anemia de Fanconi (AF) é uma doença que apresenta herança autossômica recessiva. É caracterizada por múltiplas anomalias congênitas, progressiva falha da medula óssea e alto risco para desenvolvimento de câncer. A mais importante das características clínicas é a manifestação hematológica, responsável pelo grande número de morbidade e mortalidade em portadores de AF. Também é chamada de síndrome da instabilidade cromossômica por apresentar hipersensibilidade a agentes clastogênicos como a mitomicina C e diepoxibutano. A incidência da AF em todo o mundo é de aproximadamente, três por milhão e a frequência de heterozigotos é estimada em um para 300 na Europa e Estados Unidos. No Brasil não há dados sobre a prevalência da doença. Foram descobertos até o momento 13 grupos de complementação (FANCA, B, C, D1, D2, E, F, G, I, J, L, M e N), e os 13 genes foram clonados e pelo menos 11 genes estão relacionados ao distúrbio. O presente estudo teve como objetivo a análise das principais mutações (IVS4+4A>T, Q13X, W22X, DG322, R185X, L496R, L554P, e R548X) do gene FANCC em pacientes com quadro clínico de AF. Foram analisados 121 indivíduos com clínica compatível à AF e com DEB teste positivo. Na amostra encontramos 14% de indivíduos heterozigotos e 4% de indivíduos homozigotos para as mutações mais freqüentes do gene FANCC. As mutações mais prevalentes foram: IVS4+4A>T com 6,6% dos alelos analisados, com freqüência similar à encontrada na literatura, W22X com 2.47% dos cromossomos analisados e Q13X com 1.23% dos cromossomos analisados. Na triagem de mutações pela técnica de SSCP, encontramos alterações nos éxons 1, 4 e 6. / Abstract: Fanconi anaemia (FA) is an autosomal recessive disease characterized by congenital abnormalities, progressive bone marrow failure and high risk of developing cancer. The most important of the clinic feature is hematologic, and too the most important cause of morbidity and mortality in FA. It's also called Chromosomal Instability Syndrome to the fact of cells presents hipersensibility to DNA cross-linking agents like mitomycin C and diepoxybutane. The incidence of FA is approximately three per million and the heterozygote frequency is estimated at 1 in 300 in Europe and United States. In Brazil there's no data about prevalence of FA. It was discovered at least 13 complementation groups (FANCA, B, C, D1, D2, E, F, G, I, J, L, M e N), and 13 genes have been cloned and there are at least 11 that are related to the disease. The study had as general objective the analysis of the main mutations (IVS4+4A>T, Q13X, W22X, DG322, R185X, L496R, L554P, and R548X) of FANCC gene in patients with clinic compatible of FA. We analyzed 121 patients with compatible clinic and positive DEB test. In the sample we found 14% of individuals heterozygous and homozygous individuals of 4% for the most frequent mutations of the gene FANCC. Mutations were more prevalent: IVS4 4 A> T with 6.6% of alleles tested, often similar to that found in the literature, W22X with 2.47% of chromosomes analyzed and Q13X with 1.23% of chromosomes analyzed. In screening for mutations by the technique of SSCP, we found changes in exons 1, 4 and 6. / Mestrado / Mestre em Farmacologia

A instabilidade cromossômica e a aneuploidia são características associadas às células malignas. Sabe-se que essas alterações podem ser resultantes de erros em eventos durante a mitose. Com o objetivo de gerar uma população com ganho de ploidia, utilizamos dois inibidores de fases distintas da mitose. Resultando no aumento da frequência celular em G2/M, ou seja, células com 4 vezes o número de cromossomos (4C) e de células com DNA acima de 4, hipertetraploide. Quantificamos a quantidade de núcleos e demonstramos que o tratamento, especificamente, levou a uma NCI, e não a multinucleação. Ambas linhagens celulares submetidas ao tratamento apresentaram alterações morfológicas, como protrusões de membrana, indicando alterações no citoesquelto. Nas análises de mRNA e da expressão proteica, observamos alterações na regulação da actina, coincidindo com a elevação dos mRNAs de YAP/TAZ, efetores co-transcricionais da via Hippo, regulada por alterações no citoesqueleto de actina. Desde modo, propomos, que o tratamento utilizado é um método eficiente para o estudo de células aneuploides e da NCI, que o citoesqueleto de actina é modulado por esse fenótipo e requer YAP/TAZ, provavelmente para manter a sobrevivência e favorecer a proliferação celular observada após o tratamento. / Chromosomal instability and aneuploidy are characteristics associated with malignant cells. It is known that these changes may be due to errors in events during mitosis. In order to generate a population with gain of ploidy, we used two inhibitors of distinct phases of mitosis. Resulting in increasing cell frequency in G2/M, cells with 4 times the number of chromosomes (4C) and cells with DNA above 4, hypertetraploid. We quantified the number of nuclei and demonstrated that the treatment specifically led to NCI, not multinucleation. Both cell lines submitted to treatment presented morphological alterations, such as membrane protrusions, indicating changes in the cytoskeleton. In the analysis of mRNA and protein expression, we observed alterations in actin regulation, coinciding with the elevation of YAP / TAZ mRNAs, co-transcriptional effectors of the Hippo signaling pathway, regulated by changes in the actin cytoskeleton. We propose, that the treatment used is an efficient method for the study of aneuploid cells as well NCI. Also, the actin cytoskeleton is modulated by that phenotype which requires high YAP / TAZ, probably to maintain cell survival and promote cell proliferation observed.

Le syndrome de Bloom (BS) est une maladie génétique rare caractérisée par une forte augmentation du taux d’échanges entre chromatides soeurs, des anomalies de ségrégation des chromosomes et une prédisposition au développement de tous types de cancers. Ce syndrome est la conséquence de mutations dans les deux copies du gène BLM, codant pour une 3’-5’ ADN hélicase de type RecQ. La ou les fonctions de la protéine BLM sont encore mal définies mais les données de la littérature convergent vers un rôle de BLM dans des mécanismes de surveillance et/ou maintien de l’intégrité du génome. La protéine BLM serait impliquée dans le redémarrage de fourches de réplication bloquées pendant la phase S et serait nécessaire à la résolution de ponts anaphasiques en mitose, notamment de ponts particuliers appelées « UltraFine anaphase Bridges » (UFBs). Ces UFBs, qui relient les chromatides soeurs entre elles, ne sont pas détectables par les colorants classiques et leur présence ne peut-être révélée que par la détection des protéines PICH (Plk1-Interacting Checkpoint Helicase) ou BLM. A l’état basal, ces UFBs sont essentiellement d’origine centromérique (cUFBs).Tout l’enjeu de mon projet était de déterminer si BLM était également impliquée dans la prévention de la formation de ces cUFBs et donc si BLM jouait un rôle avant l’anaphase. Nous avons montré que BLM est recrutée aux centromères de la phase G2 jusqu’en mitose. BLM, en coopération avec la protéine PICH, est nécessaire (1) à l’organisation structurale de l’ADN centromérique, (2) à la disjonction complète des centromères, indépendamment de la voie des cohésines, suggérant une implication de ces protéines dans le processus de décaténation des centromères et (3) au recrutement de la topoisomérase IIa (Topo IIa) active aux centromères.Nos résultats révèlent ainsi une nouvelle localisation et une nouvelle fonction de la protéine BLM aux centromères et montrent pour la première fois l’implication des protéines BLM et PICH dans la décaténation centromérique avant l’anaphase. Nous proposons que BLM et PICH, par leurs activités respectives hélicase et de remodelage de la chromatine, modifient la structure des centromères pendant la pré-métaphase, rendant ainsi certaines caténations accessibles à la Topo IIa avant l’anaphase. La défaillance de ce mécanisme entraînerait la persistance de caténations centromériques non résolues avant l’anaphase. Ainsi, dans les cellules BS, la fréquence élevée de cUFBs aurait deux origines différentes : une partie correspondrait à des cUFBs formés du fait d’une décaténation défaillante des centromères avant l’anaphase, et l’autre partie correspondrait à des cUFBs « physiologiques » non résolus en anaphase. Afin de distinguer l’origine des cUFBs, nous avons appelé ceux issus de caténations non résolues avant l’anaphase les UFBs centromériques surnuméraires (SC-UFBs pour Supernumerary Centromeric UFBs). / Bloom syndrome (BS) is a rare genetic disease characterized by a sharp increase in the rate of sister chromatid exchanges, chromosome segregation abnormailities and a predisposition to the development of all types of cancers. This syndrome is caused by mutations in both copies of the BLM gene, which encodes BLM, a RecQ 3'-5 DNA helicase. The specific function(s) of BLM remain unclear, but the data from the literature converge towards a role for BLM in mechanisms monitoring and / or maintaining genome integrity. The BLM protein may be involved in restarting stalled replication forks during S phase and necessary to resolve anaphase bridges in mitosis, including particular bridges called "Ultrafine Anaphase Bridges" (UFBs). These UFBs, which link sister chromatids together, are not detectable by conventional stains and their presence can only be revealed by the detection of the proteins PICH (PLK1-interacting checkpoint helicase) or BLM. In untreated cells, UFBs originate mostly from centromeres (cUFBs).The challenge of my project was to determine whether BLM was also involved in preventing the formation of cUFBs and so, if it played a role before anaphase.We showed that BLM is recruited at centromeres from G2 phase to mitosis. BLM, in cooperation with PICH, is required for (1) structural organization of centromeric DNA, (2) completion of centromere disjunction, independently of the cohesin pathway, suggesting an involvement of these proteins in centromere decatenation process, and (3) recruitment of active topoisomerase IIα (Topo IIα) to centromeres. Thus, we report a new localization and a new function of BLM at centromeres, revealing for the first time a new role for BLM and PICH in a previously unknown centromeric decatenation mechanism, crucial for complete centromere disjunction.We propose that the combined action of BLM and PICH promotes, through their helicase and chromatin remodelling activities, respectively, the organization of centromeric chromatin, thereby rendering some centromeric catenates accessible to Topo IIa before the onset of anaphase. The failure of this mechanism may lead to the persistence of some centromeric catenations not resolved before anaphase. Thus, the increase in the frequency of centromeric UFBs in BLMdeficient cells has two different origins: cUFBs arising from catenations not resolved before anaphase and physiological cUFBs not processed at anaphase onset. Two distinguish the two cUFB origins, we defined the former as supernumerary centromeric UFBs (SC-UFBs).

Over 80% of colorectal cancers (CRCs) are sporadic/randomly arising tumors. Interval CRCs represent a subset of sporadic tumors that develop within 6-36 months after a negative colonoscopy. Interval CRCs are suggested to exhibit altered biological properties that contribute to rapid growth and proliferation. We hypothesize that chromosomal instability (CIN), or aberrant chromosome numbers, contributes to the etiology of Interval CRCs. We have assembled a Manitoban cohort of Interval and sporadic (control) CRC tumor samples, and established a fluorescence in situ hybridization approach to characterize CIN by enumerating specific chromosomes. The results of this study indicate that 75% of Interval CRCs exhibit a CIN phenotype, making CIN the most prevalent contributor to genomic instability in Interval CRCs. Only once we grasp a better understanding of the tumorigenic pathways through which Interval CRCs develop, can we tailor screening strategies and treatment options to specifically identify and combat this subset of sporadic CRC.