Like with many cancers, a single ovarian tumor can display remarkable diversity in genetics, epigenetics, expression profiles, microenvironment and cell differentiation and plasticity. This so-called intratumoral heterogeneity (ITH) is thought to greatly increase mortality by enabling tumors to adapt quickly to therapy, metastasize, and recur, thus the study of ITH holds great clinical significance. Clonal evolution and cancer stem cell (CSC) theory are two models for the initiation and propagation of a tumor, which offer differing views on the way that ITH is developed and maintained. In the clonal evolution model, cancer arises from a single cell and, through genetic instability, proliferates into a diverse population of daughter cells, which develop additional mutations and undergo Darwinian selection under the influence of the tumor microenvironment. Each cell of the clonal evolution model may be capable of initiating a tumor independently. In CSC theory, cancer arises from the transformation of a stem cell that has the capacity to self-renew and differentiate into a diverse population of daughter cells. Each cell is NOT capable of tumorigenesis as most are terminally differentiated and do not harbor self-renewing capabilities. According to CSC theory, small, rare subpopulations of CSCs persist throughout chemotherapy and are responsible for repopulating the heterogeneous tumor post-treatment. The hypothesis that CSCs may play a role in ovarian cancer progression is the subject of this thesis. Many studies have detected the presence of stem cell markers and dysregulated stem cell signaling pathways in ovarian cancer, but doubts remain as to the existence of ovarian CSCs; critics have pointed out inherent flaws in experimental designs meant to identify and characterize CSCs. For example, the presence of cancer cells which express the stem cell marker CD133 has been correlated to both positive and negative impacts on prognosis. Further challenging the study of ovarian CSCs is the lack of consensus on the true cell of origin for ovarian cancer - whether it be from the fallopian tube epithelium or ovarian surface epithelium, or elsewhere in the peritoneal cavity - this will have important implications for the identification and characterization of tumorigenic ovarian CSCs. Advocates of clonal evolution theory have put forth incredible effort to reveal the extent of inter and intra-tumoral heterogeneity in ovarian cancer, and from these data there has arisen a general consensus that cancer cell populations do evolve in a step-wise fashion, accumulating additional mutations over time. The involvement of cancer stem cells in this progression and how exactly they fit in (as a cell of origin or arising from genetic mutations), as well as their significance for different cancer types, is a question worth answering. Despite the challenges facing the study of ovarian CSCs, the clinical impact of cells with stem-like properties has been repeatedly demonstrated, especially with regard to metastatic processes and chemoresistance. Moreover, new drugs which target stem cell pathways have proven effective in the treatment of ovarian cancer. The existence of a rare subset of cells that have enhanced tumor-initiating properties is apparent in ovarian cancer, and more work is needed to characterize the unique identifiers and behavior of these cells in vivo. Future experiments involving lineage tracing promise to deepen our understanding of the nature of ovarian CSCs and address whether normal stem cells might serve as the cell of origin.

Defects within the DNA damage response (DDR) pathways are common in human malignancies. This is especially true in high-grade serous ovarian cancer (HGSOC) where defects within the Homologous Recombination (HR) pathway may be present in up to 50% of tumours. Oncolytic adenovirus is a potential novel therapy for human malignancies. These viruses infect malignant cells and multiply selectively within them causing cell death and release of mature virions. Here, I have investigated the role of the DDR in determining the efficacy of the E1A-CR2 deleted adenovirus type 5 (Ad5) vector, dl922-947, in ovarian cancer. I show that infection with dl922-947 stimulates a robust DDR within the host cell, which the virus manipulates in order to ensure optimal viral replication. In a panel of HGSOC cell lines, the extent of overreplication of genomic DNA and the degree of genomic damage following infection with dl922-947 was shown to correlate closely with viral efficacy. Functional HR, however, promoted viral DNA replication and augmented overall anti-cancer efficacy. Mechanistically, both BRCA2 and RAD51 localised to viral replication centres within the infected cell nucleus. RAD51 co-localisation was also demonstrated in cells with defective HR and occurred independently of BRCA2. In addition, a direct interaction was identified between RAD51 and adenovirus E2 DNA binding protein. Using functional assays of HR competence, I show that Ad5 infection does not alter cellular ability to repair DNA double-strand break damage via HR. These data suggest that oncolytic adenoviral therapy may be most clinically relevant in tumours with intact HR function. Using a high-throughput siRNA DNA repair screen, potential novel targets have been identified that can increase the efficacy of dl922-947 (for example: NONO) and also result in increased resistance (RPA). These results highlight the complex interplay between adenovirus and host cell. Further understanding of these pathways is vital to increase efficacy, develop biomarkers and improve patient selection into clinical trials for these therapies.

Wong Wai Yin. / Thesis (M.Phil.)--Chinese University of Hong Kong, 2005. / Includes bibliographical references. / Abstracts in English and Chinese. / Acknowledgement --- p.i / Abstract --- p.iii / Abbreviation --- p.vii / Chapter CHAPTER 1 --- INTRODUCTION --- p.1-1 / Chapter 1.1 --- Classification of common epithelial ovarian tumors --- p.1-2 / Chapter 1.1.1 --- Serous tumors --- p.1-4 / Chapter 1.1.2 --- Mucinous tumors --- p.1-5 / Chapter 1.1.3 --- Endometrioid tumors --- p.1-6 / Chapter 1.1.4 --- Clear cell tumors --- p.1-6 / Chapter 1.1.5 --- Cancer staging --- p.1-7 / Chapter 1.1.6 --- Tumor grading --- p.1-8 / Chapter 1.2 --- Etiology --- p.1-10 / Chapter 1.2.1 --- Factors associated with increased risks --- p.1-10 / Chapter 1.2.2 --- Factors associated with decreased risks --- p.1-12 / Chapter 1.2.3 --- Other factors --- p.1-13 / Chapter 1.3 --- Understanding of progression of ovarian carcinoma --- p.1-13 / Chapter 1.4 --- Current screening test for ovarian cancer --- p.1-15 / Chapter 1.4.1 --- Transvaginal utrasound --- p.1-15 / Chapter 1.4.2 --- Serum tumor markers --- p.1-16 / Chapter 1.5 --- Molecular basis of ovarian cancer --- p.1-18 / Chapter 1.5.1 --- Loss of heterozygosity --- p.1-18 / Chapter 1.5.2 --- Microsatellite instability --- p.1-19 / Chapter 1.5.3 --- Oncogenes --- p.1-19 / Chapter 1.5.4 --- Tumor suppressor genes --- p.1-21 / Chapter 1.6 --- Microarray gene expression profiling analysis --- p.1-25 / Chapter 1.6.1 --- Princeple of DNA micorarray --- p.1-26 / Chapter 1.6.2 --- Types of microarray --- p.1-29 / Chapter 1.7 --- Gene expression profiling of ovarian cancer --- p.1-29 / Chapter 1.7.1 --- Up-regulated genes in ovarian cancer --- p.1-30 / Chapter 1.7.2 --- Down-regulated genes in ovarian cancer --- p.1-32 / Chapter 1.8 --- Project aims --- p.1-35 / Chapter CHPATER 2 --- MATERIALS AND METHODS --- p.2-1 / Chapter 2.1 --- Materials --- p.2-1 / Chapter 2.1.1 --- Patients --- p.2-1 / Chapter 2.1.2 --- Ovarian tissue specimen --- p.2-1 / Chapter 2.2 --- Methods --- p.2-2 / Chapter 2.2.1 --- Preparation of OCT-embedded Specimen Sections --- p.2-2 / Chapter 2.2.2 --- Microdissection of Tumor Cells from Specimen Sections --- p.2-3 / Chapter 2.2.3 --- Disruption of normal ovarian frozen tissue --- p.2-3 / Chapter 2.2.4 --- Total RNA Extraction --- p.2-3 / Chapter 2.2.4.1 --- RNA Isolation --- p.2-4 / Chapter 2.2.4.2 --- DNase I Digestion --- p.2-4 / Chapter 2.2.4.3 --- RNA Cleanup and Elution --- p.2-5 / Chapter 2.2.5 --- Oligonucleotide Microarray --- p.2-6 / Chapter 2.2.5.1 --- Two-Cycle cDNA Synthesis --- p.2-6 / Chapter 2.2.5.2 --- Synthesis of Biotin-Labeled cRNA --- p.2-9 / Chapter 2.2.5.3 --- Fragmenting the cRNA for Target Preparation --- p.2-9 / Chapter 2.2.5.4 --- Target Hybridization --- p.2-10 / Chapter 2.2.5.5 --- "Array Washing, Staining, and Scanning" --- p.2-11 / Chapter 2.2.5.6 --- Statistical Analysis of Microarray Data --- p.2-11 / Chapter 2.2.6 --- Quantitative Real-time Polymerase Chain Reaction --- p.2-13 / Chapter 2.2.6.1 --- Primer and Probe --- p.2-13 / Chapter 2.2.6.2 --- Reverse-transcription --- p.2-13 / Chapter 2.2.6.3 --- Plate Setup --- p.2-14 / Chapter 2.2.6.4 --- Fluocogenic PCR --- p.2-14 / Chapter 2.2.6.5 --- Statistical Analysis of Quantitative Real-time PCR Data --- p.2-15 / Chapter CHAPTER 3 --- RESULTS --- p.3-1 / Chapter 3.1 --- Microarray gene expression data analysis --- p.3-1 / Chapter 3.1.1 --- Unsupervised Gene Selection --- p.3-1 / Chapter 3.1.2 --- Supervised Gene Selection --- p.3-3 / Chapter 3.1.2.1 --- Gene expression profiles distinguish Serous Epithelial Ovarian Tumor from Normal Ovary and identifydifferentially expressed genes --- p.3-3 / Chapter 3.1.2.2 --- Gene expression profiles distinguish Advanced Stage Serous Epithelial Ovarian Tumor from Early Stage Serous Epithelial Ovarian Tumor and identify differentially expressed genes --- p.3-22 / Chapter 3.1.2.3 --- Gene expression profiles distinguish Metastatic Serous Epithelial Ovarian Tumor from Primary Serous Epithelial Ovarian Tumor and identify differentially expressed genes --- p.3-24 / Chapter 3.2 --- Validation of microarray data by quantitative Real-time PCR --- p.3-27 / Chapter 3.2.1 --- Fold change of candidate genes --- p.3-27 / Chapter 3.2.2 --- Correlation between microarray and quantitative real-time PCR results --- p.3-29 / Chapter 3.2.3 --- Comparison of the expression of candidates genes among the different histological types of epithelial ovarian tumors --- p.3-32 / Chapter CHAPTER 4 --- DISCUSSION --- p.4-1 / Chapter 4.1 --- Global gene expression profiling using oligonucleotide microarray --- p.4-1 / Chapter 4.1.1 --- "Sensitivity, specificity and reproducibility of the Affymetrix GeneChip® microarray" --- p.4-1 / Chapter 4.1.2 --- Microarray analysis software --- p.4-3 / Chapter 4.1.2.1 --- DNA-Chip Analyzer software --- p.4-3 / Chapter 4.1.2.2 --- Comparison of statistical methods for analysis of Affymetrix GeneChip® microarray data --- p.4-5 / Chapter 4.2 --- Validation of microarray data --- p.4-7 / Chapter 4.2.1 --- Advantages of using real-time PCR for mRNA quantification --- p.4-8 / Chapter 4.2.2 --- Comparison of mRNA gene expression by RT-PCR and DNA microarray --- p.4-9 / Chapter 4.3 --- Gene expression profiling in serous ovarian cancer compared with normal ovarian epithelium --- p.4-10 / Chapter 4.3.1 --- Potential biomarkers or therapeutic targets in ovarian cancer --- p.4-12 / Chapter 4.4 --- Gene expression profiling in advanced serous ovarian cancer compared with early ovarian cancer --- p.4-16 / Chapter 4.4.1 --- Potential prognostic markers or therapeutic targets in advanced ovarian cancer --- p.4-17 / Chapter 4.5 --- Gene expression profiling in metastatic cancer compared with primary ovarian cancer --- p.4-22 / Chapter 4.5.1 --- Potential predictive markers or therapeutic targets in metastatic cancer of ovary origin --- p.4-23 / Chapter CHAPTER 5 --- CONCLUSIONS --- p.5-1 / Chapter CHAPTER 6 --- FUTURE PROSPECT --- p.6-1 / REFERENCES --- p.R-1

Orientadores: Sophie Françoise Mauricette Derchain, Luis Otávio Zanatta Sarian / Tese (doutorado) - Universidade Estadual de Campinas, Faculdade de Ciências Médicas / Made available in DSpace on 2018-11-07T13:25:01Z (GMT). No. of bitstreams: 1 Moraes_DenisedaRochaPittaLimade_D.pdf: 1846599 bytes, checksum: 5c83c44db1103b63041e2ceb928d48d4 (MD5) Previous issue date: 2012 / Resumo: Objetivo: Avaliar a acurácia da mesotelina, CA125, HE4 e índice ROMA na diferenciação de mulheres brasileiras com tumores malignos de ovário daquelas com tumores benignos e ou mulheres saudáveis, e avaliar se os sintomas específicos relatados pelas mulheres podem ser usados em associação à expressão desses marcadores séricos, na diferenciação pré-operatória de neoplasia maligna de ovário. Sujeitos e Métodos: Neste estudo de corte transversal foram incluídas 199 mulheres com massa anexial (67 com tumores malignos e 132 com tumores benignos) e 150 mulheres saudáveis. Todas as mulheres com massa anexial, atendidas no hospital do Departamento de Obstetrícia e Ginecologia da Faculdade de Medicina da UNICAMP, foram convidadas a participar do estudo. Um grupo-controle, de mulheres saudáveis atendidas nos ambulatórios de menopausa e planejamento familiar no mesmo hospital, foi selecionado. Após uma explicação sobre os métodos e objetivo da pesquisa, todas as mulheres responderam o questionário com relação aos sintomas específicos. Foram coletados dados sobre a idade e índice de massa corpórea e sangue periférico para quantificação da mesotelina, o CA125 e a HE4. Foi usado o algoritmo de particionamento recursivo baseado no modelo de regressão linear para verificar a contribuição da idade e de cada marcador sérico no diagnóstico de tumores malignos. Foram comparadas as áreas sob as curvas (AUCs) obtidas através das curvas ROC (Receiver Operator Characteristics) de cada marcador sérico e índice ROMA, para diferenciar mulheres com tumores malignos. Foi calculada a proporção de mulheres com cada um dos 22 sintomas específicos nos grupos com tumores malignos de ovário, tumores benignos e mulheres saudáveis. O sintoma foi considerado positivo quando ocorria mais que 12 vezes ao mês e por até um ano. A proporção de sintomas foi comparada utilizando teste de qui-quadrado ou teste exato de Fischer, quando apropriado. Os 16 sintomas específicos aplicáveis a toda a coorte e para o qual a periodicidade foi verificada foram submetidos à análise pelo Método de Ward para agrupamento hierárquico. Os agrupamentos de sintomas e sintomas isolados identificados foram: abdômen (abdômen inchado e/ou aumento do volume abdominal); dor (dor pélvica, costas e/ou abdominal); pernas inchadas; digestão (estômago cheio e/ou náusea /vômito); alimentação (dificuldade para comer e/ou empachada); sente alguma massa abdominal; diversos (fadiga e/ou dificuldade para respirar); bexiga (urgência em urinar e/ou urinar frequentemente). Foi avaliada a proporção de mulheres com cada agrupamento de sintomas ou sintomas isolados em mulheres com tumores malignos, tumores benignos e saudáveis, através do teste qui-quadrado para tendências. Utilizou-se um algoritmo de particionamento recursivo para verificar a contribuição da idade da mulher, de cada agrupamento de sintomas ou sintomas isolados, estado menopausal, perda de peso e marcadores séricos no diagnóstico de tumores malignos. Resultados: O CA125 foi o marcador sérico com maior capacidade para discriminar mulheres com tumores malignos (p<0,001). Entre as mulheres com tumores benignos e CA125 positivo, a HE4 foi positiva em apenas um caso e a mesotelina foi positiva em outro. Em mulheres com CA125 negativo, a idade, a mesotelina e a HE4 não contribuíram para a diferenciação entre mulheres com tumores malignos, tumores benignos e saudáveis. Em contrapartida, em mulheres com CA125 positivo, a HE4 contribuiu significantemente para detecção de mulheres com tumores malignos (p<0,01). A AUC da mesotelina foi menor que das AUC dos outros marcadores. O ROMA e o CA125 apresentarm melhores AUCs do que o HE4. A proporção de mulheres com cada um dos agrupamentos de sintomas ou sintomas isolados foi significativamente maior em mulheres com tumores malignos, quando comparadas àquelas com tumores benignos e, destas, comparadas com as mulheres saudáveis (p tendência em todas as comparações <0,01). Após a análise multivarida, as associações mais significativas para detecção de tumores malignos de ovário foram as do agrupamento abdômen (p<0,001), expressão do CA125 (p<0,001), agrupamento dor (p=0,01) e perda de peso (p=0,03). Conclusões: Em mulheres com CA125 negativo, a mesotelina e HE4 não contribuíram para detecção do carcinoma de ovário. Entretanto, em mulheres com CA125 positivo, a HE4 contribuiu para diferenciar aquelas com tumores malignos. Em mulheres com tumores malignos de ovário, os sintomas específicos, abdômen e dor foram significantemente mais frequentes. Podem ser utilizados em associação ao CA125 na diferenciação de tumores malignos em mulheres com massa anexial / Abstract: Objective: To evaluate the accuracy of mesothelin, CA125, HE4 and ROMA index in the differentiation of Brazilian women with ovarian malignant tumors from those with benign tumors or healthy women; and to evaluate whether the prevalence of specific self-reported symptoms can be used in association to the expression of serum markers for the preoperative differentiation of ovarian malignant tumors. Study Design: For this cross sectional study, 199 women with adnexal mass (67 with malignant tumors and 132 with benign tumors) and 150 healthy women were included. All women with adnexal masses, attending the hospital of the Department of Obstetrics and Gynecology of the Unicamp School of Medicine were invited to participate in the study. A control group of healthy women attending menopause and family planning clinics at the same hospital were selected. After an explanation about the study research methods and purpose all women answered a survey regarding specific symptoms. There were also collected data on age and body mass index. Peripheral blood was collected for serum measurements of mesotelina, CA125 and HE4. A recursive partitioning algorithm, based on a linear regression model was used to confirm the contribution of age and each of the serum markers to the diagnosis of malignant tumors. Comparison of Area Under the Curve (AUC) obtained through Receiver Operator Characteristics (ROC) curves for each of the serum markers and ROMA index were used to differentiating women with malignant tumors. We next calculated the proportion of women with each of the 22 specific symptoms in the groups of women with ovarian malignant tumors, benign tumors and healthy women. We considered a symptom positive if it occurred more than 12 times per month and for less than one year. The proportions were pairwise compared using chi-square or the Fisher exact test where appropriate. The 16 specific symptoms which applied to the entire cohort and for which the periodicity had been ascertained were further subjected to the Ward's Hierarchical Clustering Method. Clusters of symptoms and isolated symptoms were: abdomen (abdominal bloating and/or increased abdomen size); pain (pelvic, back and/or abdominal pain); leg swelling; digestion (indigestion and/or nauseas /vomiting); eating (unable to eat normally and/or feeling full quickly); able to feel abdominal mass; miscellaneous (fatigue and/or difficulty breathing); bladder (urinary urgency and/or frequent urination). We evaluated the trend in proportion of women with each cluster of symptoms in the groups of women with malignant tumors, benign tumors and healthy women using the chi-squared test for trend in proportions. Another recursive partitioning algorithm was used to confirm the contribution of patient age, clusters of symptoms, menopausal status, weight loss and the serum markers to the diagnosis of malignant tumors Results: CA125 was the serum marker that had the greatest capacity to discriminate women with malignant tumors (p<0.001). Among the women with benign tumors and positive CA125, HE4 was positive in only one case and mesothelin in another case. In women with negative CA125 neither age nor mesothelin nor HE4 contributed any further to the differentiation between women with malignant, tumors benign tumors and healthy women. In contrast, for women with positive CA125, HE4 contributed significantly to the detection of women with malignant tumors (p<0.01). The AUC for mesothelin was smaller than that for all the other curves, and ROMA and CA125 had better AUC than HE4. The proportion of women with each of the clusters of symptoms and isolated symptoms decreased significantly from the group of women with malignant tumors to that with benign tumors and from this group to the healthy women (p for trends in all comparisons= <0.01). After a multivariate analysis the association that contributed the most to the detection of malignant ovarian tumors was that of the abdomen cluster (p<0.001), CA125 expression (p<0.001), pain cluster (p=0.01) and weight loss (p=0.03). Conclusion: In women with negative CA125 neither mesothelin nor HE4 contributed to detect ovarian carcinoma. HE4 was helpful to differentiate malignant tumors when CA125 is positive. Specific symptoms, abdomen and pain were significantly higher in women with malignant ovarian tumors and may be used along with the CA125 to select women with ovarian malignancy among those with adnexal masses / Doutorado / Oncologia Ginecológica e Mamária / Doutora em Ciências da Saúde

O objetivo do estudo realizado no Laboratório de Pesquisa em Suínos e Laboratório de Oncologia Veterinária da Faculdade de Medicina Veterinária e Zootecnia da Universidade de São Paulo– FMVZ. USP, São Paulo – SP, foi verificar as degenerações císticas ovarianas e suas características histopatológicas de fêmeas tratadas com gonadotrofinas exógenas e flushing. Foram utilizadas 72 marrãs pré-púberes da linhagem Pen Ar Lan NAIMA&reg;, com idade média de 157,49 &plusmn; 5,01 dias, e 96,65 &plusmn; 7,70 Kgs de peso, distribuídas em quatro tratamentos em arranjo fatorial 2x2, sendo um dos fatores o emprego ou não do flushing, e o outro a administração ou não do hormônio. A ração utilizada continha 16% de PB e 3.286,73 Kcal/Kg ME. A partir do 7&ordm; dia do primeiro estro, induzido com hormônio, as fêmeas do tratamento com flushing receberam 50% de incremento da mesma ração. No 16&ordm; dia, após o primeiro estro, as marrãs receberam aplicação de 600 UI de eCG (Novormon&reg:) e 2,5 mg de LH (Lutropin&reg;), 72 horas após, caracterizando os quatro.tratamento (com e sem homônio, com e sem flushing). Todas as marrãs foram expostas ao macho duas vezes ao dia, a partir do 10&ordm; dia e inseminadas ao segundo estro com sêmem heterospérmico. Ao abate, realizado no 5º dia após inseminação, os genitais foram colhidos para exame dos ovários (averiguação das patologias ovarianas macroscopica e histologicamente) e também os embriões pa averiguação da viabilidade embrionária. As características avaliadas foram número de ovulações, percentual de cistos ovarianos e percentuais de cistos foliculares, de corpo lúteo e luteinizados. Não foram encontradas diferenças significativas, entre os tratamentos, para os percentuais do número de ovulações, cistos ovarianos, cistos foliculares, corpo lúteo e luteinizados. Houve diferença significativa na viabilidade embrionária, constatando-se efeito adverso da combinação hormonal provocando queda da viabilidade, o mesmo não ocorrendo para o efeito flushing, o qual provocou aumento da viabilidade. Concluiu-se que tanto o emprego das gonadotrofinas (eCG e LH), quanto o uso do flushing, não influíram na ocorrência de cistos ovarianos e que nos estudos iniciais sobre a gênese da patologia, pela influência dos referidos fatores, os diferentes tipos de cistos estudados, não provocaram efeitos deletérios na dinâmica do desenvolvimento folicular, não constatando-se reflexos negativos na puberdade em marrãs / The experiment carried out in LPS (Swine Research Laboratory) and LOV (Oncology Veterinary Laboratory) of Veterinary Medicine and Animal Science Faculty of São Paulo University (FMVZ-USP), verified the cystic degeneration and their histopatological traits in gilts, treated witht the combination of gonadotophins and flushing. 72 pre-pubertal gilts of PenArLan Naïma&reg; lineage with 157,49 &plusmn; 5,01, days of age and 96,65 &plusmn; 7,70 Kg of live weight were alloted in 4 treatments in a factorial arrangement 2X2. The feed used contained 16% (CP) and 3.286,73 Kcal/Kg ME. From the seventh day on first estrus, induced with gonadotrophins, the females with flushing treatment received a increment of 50% the same feed. From the sisteenth day of the first estrus the gilts that received hormone treatment (600 UI of eCG na 2,5 mg of LH (Lutropin&reg;) 72 hours late), were exposed to a boar twice at day, from the tenth day when the gilts were inseminated t the second estrus with a heterospermic semen. At slaughter on the fifth day after insemination, the genitals were remove to exmine the ovarian patologies, and the embryos. There weren’t significances among treatments to the percentuals of the ovulation number, ovarian cystic percentage; foliculous percentage, corpus luteos and luteinazed cystics. There was significant diference in embryonic viability that indicated an adverse effects of the hormonal combinations in comparision to the flushing treatment. We concluded that both gonadotrophins (eCG and LH) and fushing have influenced on ovarian cystic ocorrence and in the beginning of the study based on the influence of the two factors (hormone and flushing) in the patological cystic genesis, haven’t brought about any deleterious effects of the differents cystic in the dinamic folicular development with no negative reflexion on puberty in gilts

O objetivo do estudo realizado no Laboratório de Pesquisa em Suínos e Laboratório de Oncologia Veterinária da Faculdade de Medicina Veterinária e Zootecnia da Universidade de São Paulo– FMVZ. USP, São Paulo – SP, foi verificar as degenerações císticas ovarianas e suas características histopatológicas de fêmeas tratadas com gonadotrofinas exógenas e flushing. Foram utilizadas 72 marrãs pré-púberes da linhagem Pen Ar Lan NAIMA&reg;, com idade média de 157,49 &plusmn; 5,01 dias, e 96,65 &plusmn; 7,70 Kgs de peso, distribuídas em quatro tratamentos em arranjo fatorial 2x2, sendo um dos fatores o emprego ou não do flushing, e o outro a administração ou não do hormônio. A ração utilizada continha 16% de PB e 3.286,73 Kcal/Kg ME. A partir do 7&ordm; dia do primeiro estro, induzido com hormônio, as fêmeas do tratamento com flushing receberam 50% de incremento da mesma ração. No 16&ordm; dia, após o primeiro estro, as marrãs receberam aplicação de 600 UI de eCG (Novormon&reg:) e 2,5 mg de LH (Lutropin&reg;), 72 horas após, caracterizando os quatro.tratamento (com e sem homônio, com e sem flushing). Todas as marrãs foram expostas ao macho duas vezes ao dia, a partir do 10&ordm; dia e inseminadas ao segundo estro com sêmem heterospérmico. Ao abate, realizado no 5º dia após inseminação, os genitais foram colhidos para exame dos ovários (averiguação das patologias ovarianas macroscopica e histologicamente) e também os embriões pa averiguação da viabilidade embrionária. As características avaliadas foram número de ovulações, percentual de cistos ovarianos e percentuais de cistos foliculares, de corpo lúteo e luteinizados. Não foram encontradas diferenças significativas, entre os tratamentos, para os percentuais do número de ovulações, cistos ovarianos, cistos foliculares, corpo lúteo e luteinizados. Houve diferença significativa na viabilidade embrionária, constatando-se efeito adverso da combinação hormonal provocando queda da viabilidade, o mesmo não ocorrendo para o efeito flushing, o qual provocou aumento da viabilidade. Concluiu-se que tanto o emprego das gonadotrofinas (eCG e LH), quanto o uso do flushing, não influíram na ocorrência de cistos ovarianos e que nos estudos iniciais sobre a gênese da patologia, pela influência dos referidos fatores, os diferentes tipos de cistos estudados, não provocaram efeitos deletérios na dinâmica do desenvolvimento folicular, não constatando-se reflexos negativos na puberdade em marrãs / The experiment carried out in LPS (Swine Research Laboratory) and LOV (Oncology Veterinary Laboratory) of Veterinary Medicine and Animal Science Faculty of São Paulo University (FMVZ-USP), verified the cystic degeneration and their histopatological traits in gilts, treated witht the combination of gonadotophins and flushing. 72 pre-pubertal gilts of PenArLan Naïma&reg; lineage with 157,49 &plusmn; 5,01, days of age and 96,65 &plusmn; 7,70 Kg of live weight were alloted in 4 treatments in a factorial arrangement 2X2. The feed used contained 16% (CP) and 3.286,73 Kcal/Kg ME. From the seventh day on first estrus, induced with gonadotrophins, the females with flushing treatment received a increment of 50% the same feed. From the sisteenth day of the first estrus the gilts that received hormone treatment (600 UI of eCG na 2,5 mg of LH (Lutropin&reg;) 72 hours late), were exposed to a boar twice at day, from the tenth day when the gilts were inseminated t the second estrus with a heterospermic semen. At slaughter on the fifth day after insemination, the genitals were remove to exmine the ovarian patologies, and the embryos. There weren’t significances among treatments to the percentuals of the ovulation number, ovarian cystic percentage; foliculous percentage, corpus luteos and luteinazed cystics. There was significant diference in embryonic viability that indicated an adverse effects of the hormonal combinations in comparision to the flushing treatment. We concluded that both gonadotrophins (eCG and LH) and fushing have influenced on ovarian cystic ocorrence and in the beginning of the study based on the influence of the two factors (hormone and flushing) in the patological cystic genesis, haven’t brought about any deleterious effects of the differents cystic in the dinamic folicular development with no negative reflexion on puberty in gilts

Ovarian cancer is the deadliest gynecological malignancy affecting women. Diagnosis often occurs late due to non-specific symptoms, but if detected early, there is excellent chance for survival. One of the most important risk factors is family history. Up to 24% of cases are due to inherited loss-of-function mutations in genes involved in the DNA damage response. The theory underlying hereditary cancers is Knudson's two-hit theory of cancer causation, where two hits are necessary for cancer to occur in an individual: one in the germline and one in the tissue. The genes, CHEK1 and CHEK2, are modulators of the DNA damage response, and could be susceptible to a first hit. There is little to no evidence about whether loss-of-function mutations in either of these two genes can lead to ovarian cancer. Using a cohort of 587 ovarian cancer cases and 557 controls, this study sought to determine if CHEK1 and CHEK2 are associated with ovarian cancer. Applying Fisher's exact test to compare mutation rates and the t test to compare age at time of diagnosis, the alternative hypothesis about an association between disease and mutations in CHEK1 and CHEK2 was rejected, but an association between younger age at diagnosis in cases and mutations in either gene was confirmed. The association between age and mutations in either of these genes suggests that there is some influence of age on disease, but a clear association between development of disease and mutations cannot yet be established. This research has implications for social change: By recognizing the need to test earlier in women with mutations in CHEK1 and/or CHEK2, they will have a higher chance of survival and better health outcomes, not only for ovarian cancer but for related cancers as well.

The tumor suppressor gene TP53 sits at the crux of response to cellular stresses. This is the most frequently inactivated gene in human tumors, being the target of somatic mutations. The protein product of TP53 is p53, and plays a crucial role in anti-proliferative signals through the induction of apoptosis, senescence, and cell-cycle arrest when activated by stresses such as genotoxic chemotherapeutic drugs. Therefore, the status of TP53 mutation in a tumor has profound implications for the tumorigenic potential as well as the response to anti-cancer therapies. Indeed, numerous studies have shown a predictive and prognostic value of TP53 mutations to the response to chemotherapy, but just as many studies show no significant contribution of TP53 mutations to chemotherapy response. This controversy is partly due to the lack of standard methods of TP53 mutation detection, but more importantly, it is due to the categorization of all TP53 mutations into one group. Certain mutations in TP53 can confer a mutant p53 with new, gained activities, not normally present in the WT p53 protein. These have been commonly called "gain of function" (GOF) p53 proteins, and some GOF p53 proteins can even confer oncogenic properties. However, not all gained functions are necessarily implicated in oncogenicity. Using stringent criteria, we have defined a select group of GOF TP53 mutations that do function as oncogenic proteins as oncomorphic TP53 mutations. In this work, we utilize data available from a large patient population through The Cancer Genome Atlas (TCGA) as well as data available from the University of Iowa Gynecologic Oncology Tumor Bank to examine the association of oncomorphic TP53 mutations with patient outcome using advanced serous ovarian cancer as a model. We demonstrate that oncomorphic TP53 mutations are associated with worse progression-free survival, chemoresistance, and higher rates of recurrence than other mutations in TP53 that have no evidence of oncomorphic abilities. We identify molecular alterations in patients with oncomorphic TP53 mutations, particularly the increased expression of β-catenin. We also observe that oncomorphic p53 proteins lose the normal protein:protein interactions with the microRNA microprocessing complex, implicating the role of dysregulated miRNAs in pathways associated with chemoresistance. The cumulative results from our studies provide human evidence for the consideration of different classes of TP53 mutations. Patients with oncomorphic TP53 mutations deserve careful follow-up therapy and may require novel treatment regimens to improve outcomes. We propose that stratification of patients should be considered based upon the individual TP53 mutation identified from their tumors.

Cancer develops in many organs and tissues in the body through genetic and environmental modifications to acquire the hallmarks of cancer. The hallmarks of cancer allow the cells to become malignant and progress to a tumorigenic state. It has previously been shown in various carcinomas that HIF-2a, a key component in hypoxia adaptation, has a role in autonomous growth, the first hallmark of cancer. Ovarian cancer is the most lethal of the gynecological malignancies and accounts for 3% of new cases in women annually but is the fifth most common cause of death due to cancer. Here, it is shown in two ovarian carcinoma cell lines that HIF-2a is involved in in vitro and in vivo growth. It is also shown that the effect of HIF-2a is due to its role in autonomous growth and not vascularization with the use of in vitro spheroids. From recent findings in the laboratory the oxygen-stimulated translation initiation complex was discovered and HIF-2a is one of its components. In the absence of HIF-2a there is a downregulation in translation in hypoxia in ovarian carcinoma. This is also seen in a HIF-2a translational target, IGF1R and its downstream signaling pathway, which may be involved in autonomous growth as well as other hallmarks of cancer. Taken together, the data in this thesis presents the importance of HIF-2a in autonomous growth and cancer progression in ovarian carcinoma, as well as verifying its role in translation.

The long-term natural history of ovarian reserve after adjuvant chemotherapy for breast cancer has been poorly described. We recruited 52 breast cancer survivors treated with adjuvant chemotherapy before 40 years of age who remained premenopausal after chemotherapy treatment. Twenty (38.5%) were more than five years out from treatment. Ovarian reserve estimates were compared with a control group. Anti-Müllerian hormone (AMH), follicle stimulating hormone and luteinizing hormone demonstrated significant differences consistent with reduced ovarian reserve in breast cancer survivors. Mean AMH was 6.65 pmol/l in survivors compared to 17.43 in controls (p < 0.001). Attained age and age at the time of treatment were correlated with AMH levels in breast cancer survivors. Conclusion: Ovarian reserve is significantly reduced in young breast cancer survivors. Age is the major predictor of AMH level in survivors. A 35 year old breast cancer survivor has an AMH level similar to a 45 year old control.