Dentre as manifestações clínicas observadas em pacientes sépticos, as disfunções neurológicas são, provavelmente, as de fisiopatologia mais obscura e pobremente explorada, o que consequentemente as torna de difícil entendimento e tratamento médico. Adicionalmente, estudos epidemiológicos indicam que a síndrome séptica é mais frequente e mais letal em pacientes idosos. Nesse contexto, este trabalho objetivou comparar os efeitos da sepse, induzida pelo modelo de ligadura e perfuração cecal, no encéfalo de ratos jovens e idosos por meio da análise da expressão gênica de larga escala (transcriptoma), a fim de averiguar como as alterações no padrão de expressão podem estar relacionadas a disfunções neurológicas. Os resultados deste estudo indicaram a diminuição da expressão dos genes Bcl-3, S100A8 e uridina fosforilase 1, bem como o aumento da expressão de Stefin A3, sendo tais efeitos característicos das manifestações comuns da sepse no sistema nervoso central, independentemente da idade dos animais; por outro lado, a diminuição da expressão do gene da haptoglobina foi observada apenas nos animais idosos com sepse. De forma geral, na comparação entre animais idosos e jovens, os resultados desta pesquisa apontaram que animais idosos apresentam uma quantidade menor de genes modificados pela sepse, o que sugere menor capacidade de ativar mecanismos neuroprotetores / Among the clinical manifestations observed in septic patients, the neurological dysfunctions are probably the most obscure and poorly explored pathophysiology, which consequently makes them difficult to understand and to treat. Additionally, epidemiological studies indicate that septic syndrome is more frequent and more lethal in elderly patients. In this context, this article is aimed at comparing the effects of sepsis, induced by the ligature model and cecal perforation, on the brain of young and elderly rats by means of the analysis of the large scale gene expression (transcriptome), in order to investigate how the changes in this expression may be related to neurological dysfunctions. The results of this study indicated decreased expression of the Bcl-3, S100A8 and uridine phosphorylase 1 genes, as well as increased expression of Stefin A3, these effects being characteristic of the common manifestations of central nervous system sepsis regardless of the age of the animals; on the other hand, decreased haptoglobin gene expression was observed only in the elderly animals with sepsis. In general, in the comparison between old and young animals, the results of this research indicated that elderly animals present a smaller amount of genes modified by sepsis, which suggests a smaller capacity to activate neuroprotective mechanisms

Introdução: Os micro RNA (miRNA) são formados a partir de RNA precursores de fita dupla que contém entre 60 a 110 nucleotídeos e formam estruturas do tipo hairpin. Imediatamente após sua transcrição pela RNA polimerase II a enzima Dicer promove a clivagem do RNA precursor em seqüências menores contendo 19 a 22 nucleotídeos. Após a clivagem, o miRNA integra-se ao complexo silenciador induzido pelo RNA (RISC) que o conduz ao seu RNA mensageiro (mRNA) homólogo recém transcrito. Esta associação promove a degradação do mRNA, ou interfere na tradução da proteína caracterizando um grande mecanismo de controle da expressão dos genes. Este mecanismo está relacionado ao desenvolvimento de órgãos e tecidos, e está envolvido no processo de carcinogênese. Nosso objetivo é identificar um perfil de expressão de miRNA que defina o adenocarcinoma de próstata de prognóstico favorável e desfavorável considerando os níveis de PSA e dados anatomopatológicos. Materiais e métodos: Foram selecionados 53 pacientes com tumores desfavoráveis (mediana do escore de Gleason igual a 8, 79,2% estadiados pT3, mediana de PSA 10,1 ng/mL e mediana do volume tumoral de 23%) e 45 considerados favoráveis (mediana do escore de Gleason igual a 5, 80% estadiados pT2, mediana de PSA de 7,8 ng/mL e mediana do volume tumoral de 11,5%). O controle foi representado por 7 pacientes com hiperplasia prostática benigna (HPB). Todos os pacientes foram submetidos a prostatectomia radical pelo mesmo cirurgião. Os espécimes cirúrgicos foram examinados na sua totalidade pelo mesmo patologista. A análise dos miRNA foi feita a partir de tecido congelado e tecido incluído em parafina usando a técnica da reação em cadeia da polimerase em tempo real quantitativa (qRT-PCR) utilizando primers e sondas Taqman® específicas. O RNU43 foi usado como controle interno. Resultados: Com exceção dos miRNA 199a, 21, 15a, 16 e 25 que se mostraram subexpressos tanto nos casos desfavoráveis como nos favoráveis, houve uma diminuição global na expressão dos miRNA com redução estatisticamente significativa na expressão dos miRNA 143, 145 e 146a, 191, 218 e Let7c em tumores desfavoráveis em relação aos tumores favoráveis. Conclusão: Demonstramos que no processo de transição entre os carcinomas favoráveis e desfavoráveis de próstata existe uma perda global na expressão de miRNA que podem ser importantes controladores de expressão de uma série de genes relacionados a progressão desta neoplasia. Dados experimentais avaliando o papel desses miRNA devem ser conduzidos para que possamos definir seu papel na evolução do câncer de próstata / Introduction: micro RNA (miRNA) are formed from double-stranded RNA precursors that contain between 60-110 nucleotides and form structures such as hairpin. Immediately after their transcription by RNA polymerase II, the enzyme Dicer promotes the cleavage of precursor RNA sequences containing minor 19-22 nucleotides. After cleavage, the miRNA is part of the RNA-induced silencing complex (RISC) that leads to its messenger RNA (mRNA) newly transcribed counterpart. This association promotes the degradation of mRNA, or interferes with the protein translation characterizing a great mechanism for controlling gene expression. This mechanism is related to the development of organs and tissues, and may be involved in the process of carcinogenesis. Our goal is to identify a miRNA expression profile that distinguishes prostate adenocarcinoma of favorable and unfavorable prognosis considering the PSA and pathological findings. Material and Methods: We studied 53 patients with tumors considered unfavorable (Median of Gleason score 8, 79.2% staged pT3, median of PSA 10.1 ng/mL and median of tumor volume of 23%) and 45 considered favorable (Median of Gleason score 5, 80% staged pT2, median of PSA 7.8 ng/mL and median of tumor volume of 11.5%). The control group was represented by seven patients with benign prostatic hyperplasia (BPH). All patients underwent radical prostatectomy by the same surgeon. The surgical specimen was examined entirely by the same pathologist. The analysis of miRNA was made from frozen and paraffin embedded tissue by quantitative real-time polymerase chain reaction (qRT-PCR) using the Taqman® specific primers and probes. The RNU43 was used as a internal control. Results: Except for miRNAs 199a, 21, 15a, 16 e 25 that were underexpressed by both favorable and unfavorable prostate cancer, there was a global decrease of all miRNAs studied, and some differences were statistically significant as miRNAs 143, 145 e 146a, 191, 218 e Let7c that were underexpressed in unfavorable carcinomas compared favorable tumor. Conclusion: We have demonstrated that in the process of transition between favorable and unfavorable prostate cancer there is a global loss of expression of miRNAs. These molecules can be important controllers of a series of genes related to cancer progression. Experimental studies are needed in order to comprehend the role of these genes in prostate carcinogenesis

Les Carcinomes Papillaires du Rein (pRCC) représentent la seconde forme histologique de cancers du rein . Ils correspondent à une entité hétérogène de tumeurs subdivisées en type I et type II sur leurs caractéristiques anatomopathologiques. Leur pronostic au stade métastatique est inferieur à celui des carcinomes à cellules claires. Les caractéristiques biologiques des pRCC sont mal connues et n’ont pas permis de développer jusqu'à ce jour de thérapeutiques spécifiques.Ce travail propose, en première partie, une synthèse des données disponibles biologiques, anatomo-pathologiques, thérapeutiques et pronostiques des pRCC. Cette synthèse a fait l’objet d’une publication. ( Albiges et al. The Oncologist 2012)Le second volet est dédié à l’analyse de la place du proto-oncogène MET au sein des pRCC de type I et II et plus particulièrement les différentes modalités d’activation de ce gène. Cette analyse (i) caractérise les anomalies quantitatives de l’ADN du gène MET (CGH array pour les pRCC de type II et CGMA pour les pRCC de type I) et leur corrélation au niveau d’expression génique; (ii) recherche l’existence de mutations activatrices du domaine tyrosine kinase par séquençage du gène MET chez les pRCC de type I; et (iii) analyse également les niveaux d’expression du ligand et des co-activateurs de ce récepteur MET. Ces résultats sont en cours de publication (Albiges et al. Clinical Cancer Research)Le troisième et dernier volet de ce travail vise à identifier des pistes biologiques propres aux pRCC par l’analyses de sous groupes distinguable en termes de profils d’expression génique et surtout par l’analyses des anomalies de l’ADN identifiées par CGH array des pRCC de type II, couplées aux données de transcriptome. / Papillary renal cell carcinomas (pRCC) are the second most common form of Renal Carcinomas and belongs to the non clear cell carcinomas family. This tumour type is an heterogeneous group of tumours usually subdivided in type I and type II according to pathological features. The prognosis of pRCC in the metastatic setting is worse to clear cell carcinoma’s prognosis. Biological characteristics of pRCC are poorly known and did not allow the development of specific targeted therapies.This work first presents a synthesis of published data regarding biology, pathology, therapeutics and prognosis of pRCC. This review has been published. (Albiges et al. The Oncologist 2012)Second part is dedicated to the analysis of MET proto-oncogene across pRCC. The main focus is to assess MET activation drivers. This analysis (i) characterises MET gene DNA copy number alterations (CGH array for type II pRCC and CGMA approach for Type I pRCC) and their correlation with gene expression profiling; (ii) assess activating mutations within the tyrosine kinase of MET gene in the type I pRCC; and (iii) investigate expression level of ligand and co-activators of MET receptor. This analysis is under publication. (Albiges et al. Clinical Cancer Research)Third and last part of this work aims at identifing new biological pathway specific to pRCC using clustering of gene expression profiling and DNA abnormalities assessed by CGHarray inthe type II pRCC subtypes with matching gene expression data.

Tao Kin Pong. / Thesis (M.Phil.)--Chinese University of Hong Kong, 2005. / Includes bibliographical references (leaves 144-152). / Abstracts in English and Chinese. / Chapter i. --- Cover page --- p.1 / Chapter ii. --- Table of contents --- p.2 / Chapter iii. --- Abstract of thesis (English version and Chinese Version) --- p.4 / Chapter iv. --- Acknowledgements --- p.8 / Chapter v. --- Abbreviations --- p.9 / Chapter 1. --- Introduction / Interstitial tissue and Leydig cells --- p.11 / Seminiferous tubules --- p.11 / "Male germ cell line and spermatogenesis (Mitotic, Meiotic and Post-meiotic)" --- p.12 / Sertoli cells --- p.14 / Specialized organizations of junction present in testis --- p.15 / Dynamic nature of Sertoli-Sertoli & Sertoli-germ cell junctions --- p.16 / Role of proteases and protease inhibitors in male gametogenesis --- p.17 / Proteases and Proteases Inhibitors expressed in testis --- p.18 / Hormonal control of spermatogenesis --- p.19 / Hypothyroidism and testis development --- p.21 / Genes to be studied: / Proteases --- p.22 / Proteases Inhibitors --- p.27 / Other spermatogenesis related genes --- p.30 / Chapter 2. --- Objectives --- p.32 / Chapter 3. --- Materials and Methods / Animal treatments and tissue collection --- p.33 / RNA preparation --- p.34 / RT-PCRs --- p.35 / Real-time PCRs --- p.35 / Data manipulations and Statistics --- p.36 / Primer sequences used in this experiment --- p.37 / Chapter 4. --- Results / "Effect of neonatal hypothyroidism on developmental profile of body weight, absolute and relative testicular weight" --- p.40 / Developmental transcription profiles of genes under normal and hypothyroidism --- p.43 / Screening Data --- p.78 / Expression of non-spermatogenic genes at neonatal age --- p.88 / Responsiveness of gene transcription after thyroxin replacement --- p.89 / Changes of gene expression under different hypothyroid regimens --- p.97 / Chapter 5. --- Discussion / Changes in testicular size under hypothyroidism --- p.107 / Five patterns of transcription profile --- p.107 / "Suggestion on the role of ""MEIOTIC"" proteases and inhibitors" --- p.111 / "Suggestion on the role of ""POST-MEIOTIC"" proteases and inhibitors" --- p.113 / "Explanations on ""SOMATIC"" genes" --- p.114 / "Explanations on ""MITOTIC"" genes" --- p.115 / Explanations on the un-clustered pattern --- p.116 / Explanations on the age down-regulated group --- p.116 / Proposed clustering of genes according to their transcription profile --- p.117 / "Expression of some ""non-spermatogenic"" genes before puberty" --- p.123 / Neonatal hypothyroidism as a model for studying reproductive physiology --- p.125 / Different components of spermatogenesis --- p.127 / Chapter I. --- Roles of nuclear and chromatin related genes in assisting meiosis --- p.128 / Chapter II. --- Roles of specific transcription regulators in assisting gene selection --- p.129 / Chapter III. --- Role of signal transduction molecules for translation and activation --- p.131 / Chapter IV. --- Role of proteases and inhibitors for matrix and junctions dynamics --- p.132 / The somatic proteases and inhibitors system in the testis --- p.133 / Spermatogenic proteases and inhibitors system --- p.134 / Chapter V. --- Role of matrix and junctional molecules in intercellular interactions --- p.137 / Chapter VI. --- Role of cytoplasmic motors in cellular movement --- p.139 / Chapter 6. --- Conclusion / Proposed story of spermatogenesis - involvement of proteases and inhibitors --- p.140 / Future Direction --- p.141 / Chapter 7. --- References --- p.144

Aplicando tecnologia de microarray, objetivamos traçar o perfil do programa de maturação das Mo-DC pulsadas com HIV autólogo inativado por AT-2, a fim de identificar marcadores específicos de ativação funcional e sugerir um perfil de expressão de genes úteis na identificação de respostas ao modelo in vitro das Mo-vacina DC. Essas informações podem ajudar a estabelecer assinaturas moleculares das funções celulares mais relevante para a melhoria das vacinas terapêuticas. O perfil transcricional foi analisado com base das vias celulares moduladas das Mo-DCs no estado imaturo, transitório e maduro. O HIV-1 inativado por AT-2 induz ativação de genes associados à apresentação de antígenos. Os conjuntos de genes do citoesqueleto podem influenciar a mudança de comportamento migratório das Mo-DCs ativadas. O aumento na expressão dos receptores celulares contribuem para o recrutamento de monócitos, DCs e macrófagos para o local da infecção. Além disso, modulam a resposta imune inata e adaptativa, incluindo a polarização das células Th e sub-regulação da resposta inflamatória, que pode interferir significativamente com a resposta imune. Coletivamente, o perfil transcricional das Mo-DCs induzido pelo HIV-1 inativado com AT-2 reflete uma significativa reprogramação imunológica e celular das células envolvidas na resposta imune do hospedeiro. Os resultados deste estudo focaram na interpretação de genes específicos dos perfis de transcrição das Mo-DCs como modelo terapêutico utilizado na vacina anti-HIV. As análises de assinaturas gene associado e sua correlação as respostas funcionais simplificam a identificação de indivíduos susceptíveis a vacina e a compreensão de eventuais falhas em ensaios clínicos. Microarray permitiu a análise quantitativa e simultânea da expressão de um elevado número de genes. Os estudos do perfil de expressão foram extremamente úteis para identificar os eventos moleculares e vias envolvidas nas funções de celular induzida por estímulos específicos. Em particular, os resultados sobre o padrão global da expressão dos genes subjacentes as modificações induzidas pelo HIV-1 inativado por AT-2, na fase inicial da administração do antígeno, pôde ser extremamente útil para a identificarmos marcadores de ativação e avaliar os efeitos biológicos que poderiam estar envolvidos para modificação e otimização de estratégias vacinação com Mo-DC / Applying microarray technology, we intend to profile the program to mature Mo-DC pulsed with autologous inactivated HIV by AT-2, in order to identify specific markers of functional activation and suggest a profile of expression of specific genes, useful identification of responders to in vitro model of Mo-DC vaccine. Such information may help to establish detailed molecular signatures of cellular functions most relevant to improving the therapeutic vaccines. The transcriptional profile was analyzed on the basis of the cellular pathways modulated in immature MoDC, transitional MoDC and mature MoDC. The AT-2-inactivated HIV-1 induction of MoDC results in the activation of genes associated with antigen presentation functions. A set of cytoskeletal genes that may potentially mediate shape change and migratory behavior of activated MoDC is also observed. The increase in the expression of immune receptors contribute to the recruitment of monocytes, DCs, and macrophages to the site of infection. Moreover, they modulate both innate and adaptive immune response, including the polarization of Th cells, and the down-regulation of the inflammatory response, which may significantly interfere with the immune response. Collectively, the transcriptional profile induced by AT-2-inactivated HIV-1 in MoDc reflects a significant cellular and immunological reprogramming of cells directly involved in the host immune response. The results of this study focused on the interpretation of specific genes of transcription profile of MoDC used in therapeutic HIV vaccine model. Supplementing the analyses with examination of associated gene signatures and their correlation to functional responses will simplify the identification of responsive vaccine individuals and the understanding of eventual failures in individuals enrolled in clinical trials. Microarray approach allows quantitative and simultaneous analysis of gene expression of a large amount of genes and the systematic studies of expression patterns are extremely useful for identify molecular events and key pathways involved in cellular functions induced by specific stimuli. In particular, data on the global pattern of gene expression underlying the modifications induced by AT-2-inactivated HIV-1 in MoDC, at early stages of antigen administration, may be extremely helpful for the identification of exclusive activation markers to trace the biological effects of modifications/optimizations of the MoDc vaccination strategy

INTRODUÇÃO: O aprimoramento dos métodos de rastreamento e a conscientização da população geral contribuíram para o diagnóstico cada vez mais precoce do câncer de mama e proporcionou, juntamente com o avanço na terapêutica, altas taxas de sobrevida. O estado do acometimento axilar por metástase é um dos principais fatores prognósticos em pacientes com câncer de mama, particularmente naquelas com diagnóstico em estádio inicial. Na última década, esforços científicos foram direcionados para simplificar a amostragem dos linfonodos axilares, diminuindo a morbidade, mas respeitando os princípios oncológicos. Nesse sentindo, a biópsia do linfonodo sentinela foi considerada o avanço mais importante. Ao se obter um método preditor do estado axilar, que apresente os benefícios da abordagem padrão dissecção axilar e biópsia de linfonodo sentinela porém sem seus efeitos colaterais e que seja facilmente reproduzível, realizaremos um grande avanço na avaliação e terapêutica do câncer de mama inicial. MÉTODOS: Foi realizado estudo transversal retrospectivo com base nos prontuários de pacientes com câncer de mama invasivo, não metastático, com qualquer idade, atendidas entre 1999 e 2007 no Setor de Mastologia da Disciplina de Ginecologia do Departamento de Obstetrícia e Ginecologia da Faculdade de Medicina da Universidade de São Paulo e Clínica Professor José Aristodemo Pinotti, cujo estudo histopatológico e imunoistoquímico foi supervisionado por um único médico patologista. Realizamos uma subdivisão imunoistoquímica dos tumores, sendo considerado luminal A os tumores com receptores hormonais positivos e HER 2 negativo; luminal B os com receptores hormonais positivos e HER 2 positivo; HER 2 as pacientes com receptores hormonais negativos e HER 2 positivo e triplo negativo aquelas com receptores hormonais e HER 2 negativos. Correlacionamos esses subtipos com as variáveis clínicas idade e tamanho tumoral para predizer a probabilidade de acometimento linfonodal axilar. RESULTADOS: Duzentos e trinta e nove casos foram analisados. No subtipo luminal A, a possibilidade de metástase foi maior quanto menor a idade da paciente e maior o tamanho do tumor. Essa foi a única associação que apresentou diferença estatisticamente significante. As pacientes que possuíam tumores triplo negativo tiveram, aproximadamente, 90% menos chance de metástase linfonodal que as pacientes com tumor luminal A. CONCLUSÕES: As pacientes com tumor luminal A apresentaram, significativamente, maior probabilidade de metástase linfonodal axilar. As pacientes com de tumores triplo negativo, com idade superior a 55 anos ou tumores menores que 2 cm, revelaram menor probabilidade de metástase axilar / INTRODUTION: The improvement of screening methods and awareness of general population contributed to the increasingly early diagnosis of breast cancer and provided, together with advances in therapy, high survival rates. The status of axillary involvement is a major prognostic factor in patients with breast cancer, particularly those with early stage. In the last decade, research efforts were directed to simplify the sampling of axillary lymph nodes, decreasing the morbidity, but respecting the oncological principles. In this sense, the sentinel lymph node biopsy was considered the most important advance. If we obtain a method to predict the axillary status, with the benefits of the standard approach - axillary dissection and sentinel lymph node biopsy - without its side effects and easily reproducible, we will hold a major advance in the assessment and treatment of early breast cancer. METHODS: We conducted a retrospective cross-sectional study based on records of patients with invasive breast cancer, non metastatic, with any age, treated between 1999 and 2007 in the breast cancer Sector of the Gynecology Discilpine of the Department of Obstetrics and Gynecology, Faculty of Medicine, University of São Paulo and Private Clinic of Professor José Aristodemo Pinotti, whose histopathological studies were supervised by a single pathologist. We performed an immunohistochemical subdivision of the tumors, and considered luminal A tumors with positive hormonal receptors and negative HER 2, luminal B with positive hormonal receptors and positive HER 2, HER 2 patients with negative hormonal receptors and positive HER 2 and the triple-negative those with negative hormonal receptors and HER 2. Those subtypes were correlated with the clinical variables age and tumor size in predicting the likelihood of axillary lymph node involvement. RESULTS: Two hundred and nine cases were analyzed. In the luminal A, the possibility of axillary metastasis was higher in the younger patients and larger tumors. That was the only combination that showed statistically significant difference. The patients who had triplenegative tumors had approximately 90% less chance of lymph node metastasis than patients with tumors luminal A. CONCLUSIONS: The patients with luminal A tumors showed a significantly association with greater likelihood of axillary lymph node metastasis. The patients with triple negative tumors, age over 55 years or tumors smaller than 2 cm showed a lower likelihood of axillary lymph node metastasis

Dentre as manifestações clínicas observadas em pacientes sépticos, as disfunções neurológicas são, provavelmente, as de fisiopatologia mais obscura e pobremente explorada, o que consequentemente as torna de difícil entendimento e tratamento médico. Adicionalmente, estudos epidemiológicos indicam que a síndrome séptica é mais frequente e mais letal em pacientes idosos. Nesse contexto, este trabalho objetivou comparar os efeitos da sepse, induzida pelo modelo de ligadura e perfuração cecal, no encéfalo de ratos jovens e idosos por meio da análise da expressão gênica de larga escala (transcriptoma), a fim de averiguar como as alterações no padrão de expressão podem estar relacionadas a disfunções neurológicas. Os resultados deste estudo indicaram a diminuição da expressão dos genes Bcl-3, S100A8 e uridina fosforilase 1, bem como o aumento da expressão de Stefin A3, sendo tais efeitos característicos das manifestações comuns da sepse no sistema nervoso central, independentemente da idade dos animais; por outro lado, a diminuição da expressão do gene da haptoglobina foi observada apenas nos animais idosos com sepse. De forma geral, na comparação entre animais idosos e jovens, os resultados desta pesquisa apontaram que animais idosos apresentam uma quantidade menor de genes modificados pela sepse, o que sugere menor capacidade de ativar mecanismos neuroprotetores / Among the clinical manifestations observed in septic patients, the neurological dysfunctions are probably the most obscure and poorly explored pathophysiology, which consequently makes them difficult to understand and to treat. Additionally, epidemiological studies indicate that septic syndrome is more frequent and more lethal in elderly patients. In this context, this article is aimed at comparing the effects of sepsis, induced by the ligature model and cecal perforation, on the brain of young and elderly rats by means of the analysis of the large scale gene expression (transcriptome), in order to investigate how the changes in this expression may be related to neurological dysfunctions. The results of this study indicated decreased expression of the Bcl-3, S100A8 and uridine phosphorylase 1 genes, as well as increased expression of Stefin A3, these effects being characteristic of the common manifestations of central nervous system sepsis regardless of the age of the animals; on the other hand, decreased haptoglobin gene expression was observed only in the elderly animals with sepsis. In general, in the comparison between old and young animals, the results of this research indicated that elderly animals present a smaller amount of genes modified by sepsis, which suggests a smaller capacity to activate neuroprotective mechanisms

INTRODUÇÃO: O aprimoramento dos métodos de rastreamento e a conscientização da população geral contribuíram para o diagnóstico cada vez mais precoce do câncer de mama e proporcionou, juntamente com o avanço na terapêutica, altas taxas de sobrevida. O estado do acometimento axilar por metástase é um dos principais fatores prognósticos em pacientes com câncer de mama, particularmente naquelas com diagnóstico em estádio inicial. Na última década, esforços científicos foram direcionados para simplificar a amostragem dos linfonodos axilares, diminuindo a morbidade, mas respeitando os princípios oncológicos. Nesse sentindo, a biópsia do linfonodo sentinela foi considerada o avanço mais importante. Ao se obter um método preditor do estado axilar, que apresente os benefícios da abordagem padrão dissecção axilar e biópsia de linfonodo sentinela porém sem seus efeitos colaterais e que seja facilmente reproduzível, realizaremos um grande avanço na avaliação e terapêutica do câncer de mama inicial. MÉTODOS: Foi realizado estudo transversal retrospectivo com base nos prontuários de pacientes com câncer de mama invasivo, não metastático, com qualquer idade, atendidas entre 1999 e 2007 no Setor de Mastologia da Disciplina de Ginecologia do Departamento de Obstetrícia e Ginecologia da Faculdade de Medicina da Universidade de São Paulo e Clínica Professor José Aristodemo Pinotti, cujo estudo histopatológico e imunoistoquímico foi supervisionado por um único médico patologista. Realizamos uma subdivisão imunoistoquímica dos tumores, sendo considerado luminal A os tumores com receptores hormonais positivos e HER 2 negativo; luminal B os com receptores hormonais positivos e HER 2 positivo; HER 2 as pacientes com receptores hormonais negativos e HER 2 positivo e triplo negativo aquelas com receptores hormonais e HER 2 negativos. Correlacionamos esses subtipos com as variáveis clínicas idade e tamanho tumoral para predizer a probabilidade de acometimento linfonodal axilar. RESULTADOS: Duzentos e trinta e nove casos foram analisados. No subtipo luminal A, a possibilidade de metástase foi maior quanto menor a idade da paciente e maior o tamanho do tumor. Essa foi a única associação que apresentou diferença estatisticamente significante. As pacientes que possuíam tumores triplo negativo tiveram, aproximadamente, 90% menos chance de metástase linfonodal que as pacientes com tumor luminal A. CONCLUSÕES: As pacientes com tumor luminal A apresentaram, significativamente, maior probabilidade de metástase linfonodal axilar. As pacientes com de tumores triplo negativo, com idade superior a 55 anos ou tumores menores que 2 cm, revelaram menor probabilidade de metástase axilar / INTRODUTION: The improvement of screening methods and awareness of general population contributed to the increasingly early diagnosis of breast cancer and provided, together with advances in therapy, high survival rates. The status of axillary involvement is a major prognostic factor in patients with breast cancer, particularly those with early stage. In the last decade, research efforts were directed to simplify the sampling of axillary lymph nodes, decreasing the morbidity, but respecting the oncological principles. In this sense, the sentinel lymph node biopsy was considered the most important advance. If we obtain a method to predict the axillary status, with the benefits of the standard approach - axillary dissection and sentinel lymph node biopsy - without its side effects and easily reproducible, we will hold a major advance in the assessment and treatment of early breast cancer. METHODS: We conducted a retrospective cross-sectional study based on records of patients with invasive breast cancer, non metastatic, with any age, treated between 1999 and 2007 in the breast cancer Sector of the Gynecology Discilpine of the Department of Obstetrics and Gynecology, Faculty of Medicine, University of São Paulo and Private Clinic of Professor José Aristodemo Pinotti, whose histopathological studies were supervised by a single pathologist. We performed an immunohistochemical subdivision of the tumors, and considered luminal A tumors with positive hormonal receptors and negative HER 2, luminal B with positive hormonal receptors and positive HER 2, HER 2 patients with negative hormonal receptors and positive HER 2 and the triple-negative those with negative hormonal receptors and HER 2. Those subtypes were correlated with the clinical variables age and tumor size in predicting the likelihood of axillary lymph node involvement. RESULTS: Two hundred and nine cases were analyzed. In the luminal A, the possibility of axillary metastasis was higher in the younger patients and larger tumors. That was the only combination that showed statistically significant difference. The patients who had triplenegative tumors had approximately 90% less chance of lymph node metastasis than patients with tumors luminal A. CONCLUSIONS: The patients with luminal A tumors showed a significantly association with greater likelihood of axillary lymph node metastasis. The patients with triple negative tumors, age over 55 years or tumors smaller than 2 cm showed a lower likelihood of axillary lymph node metastasis

This thesis is about the use of massive DNA sequencing to investigate the transcriptome. During recent decades, several studies have made it clear that the transcriptome comprises a more complex set of biochemical machinery than was previously believed. The majority of the genome can be expressed as transcripts; and overlapping and antisense transcription is widespread. New technologies for the interroga- tion of nucleic acids have made it possible to investigate such cellular phenomena in much greater detail than ever before. For each application, special requirements need to be met. The work presented in this thesis focuses on the transcrip- tome and the development of technology for its analysis. In paper I, we report our development of an automated approach for sample preparation. The procedure was benchmarked against a publicly available reference data set, and we note that our approach outperformed similar manual procedures in terms of reproducibility. In the work reported in papers II-IV, we used different massive sequencing technologies to investigate the transcriptome. In paper II we describe a concatemerization approach that increased throughput by 65% using 454 sequencing,and we identify classes of transcripts not previously described in Populus. Papers III and IV both report studies based on SOLiD sequencing. In the former, we investigated transcripts and proteins for 13% of the human gene and detected a massive overlap for the upper 50% transcriptional levels. In the work described in paper IV, we investigated transcription in non-genic regions of the genome and detected expression from a high number of previ- ously unknown loci. / QC 20100723

Microarray expression analysis was carried out to identify genes with a role in epithelial ovarian cancer (EOC). The U133A Affymetrix GeneChipRTM was used to determine the expression patterns of the 3p25.3-ptel genes represented on the microarray in 14 primary cultures of normal ovarian surface epithelial (NOSE) samples, 25 frozen malignant ovarian tumor samples and four EOC cell lines. Seven genes with differential expression patterns in the tumor samples compared to the NOSE samples were identified as candidates for further analysis, starting with ARPC4, SRGAP3 and ATP2B2. Although none of the candidates had been previously studied in ovarian cancer, several had either family or pathway members that had. Expression patterns seemed unaffected by either tumor histopathological subtype or the allelic imbalances observed with loss of heterozygosity (LOH) analysis. The absence of association with genomic context suggested that differential expression was the result of transcriptional regulation rather than direct targeting.