Os sarcomas de partes moles constituem um grupo diverso de neoplasias que podem surgir nos tecidos conjuntivos praticamente de qualquer região do corpo. A ressonância magnética (RM) é atualmente o exame de escolha para detecção, estadiamento regional e acompanhamento desses tumores. A segmentação das lesões a partir das imagens de RM da rotina permite extrair dados quantitativos, que tem potencial para adicionar informações a análise. O propósito do estudo é avaliar a reprodutibilidade da segmentação manual de sarcomas de partes moles em imagens de RM de pacientes com diagnóstico definitivo confirmado por meio da histopatologia. Como objetivo secundário foi realizada a comparação da segmentação manual e semiautomática a fim de validar a segmentação semiautomática como método alternativo para segmentação desses tumores. Foi estudada uma coorte retrospectiva de 15 pacientes consecutivos com diagnóstico confirmado de sarcoma de partes moles acompanhados em nosso serviço, no período de janeiro de 2006 até janeiro de 2016, com imagens de RM adquiridas previamente ao tratamento e disponíveis para análise no formato DICOM. Foi utilizado o software 3D Slicer para realizar as segmentações pelos métodos manual e semiautomático. Três radiologistas fizeram as segmentações de forma independente e às cegas para permitir avaliação interobservador. Os resultados obtidos demonstram haver alta reprodutibilidade intraobservador com coeficiente de similaridade de Dice entre as segmentações variando de 0,849 a 0,979 e as distâncias Hausdorff variando de 3,53 mm a 20,96 mm e uma boa reprodutibilidade interobservador com coeficientes de similaridade de Dice variando de 0,741 a 0,972 e distâncias Hausdorff variando de 5,83 a 60,84 mm. Foi encontrada uma concordância substancial entre as segmentações realizadas pelo método semiautomático quando comparadas com as segmentações realizadas pelo método manual com coeficientes de similaridade de Dice variando de 0,871 a 0,973 e distâncias Hausdorff variando de 5,43 mm a 31,75 mm. Em relação ao tempo de segmentação não houve diferença estatisticamente significativa do método semiautomático quando comparado ao método manual (p>0,05). Também foram calculados os volumes obtidos nas diferentes segmentações e houve concordância quase perfeita entre as duas segmentações manuais realizadas pelo radiologista 1, entre as segmentações realizadas pelo radiologista 1 e pelo radiologista 2, entre as segmentações realizadas pelo radiologista 1 e pelo radiologista 3, e entre a segmentação manual e semiautomática realizadas pelo radiologista 1, sendo obtidos coeficientes de correlação intraclasse (ICC) entre 0,9927 e 0,9990. Os resultados obtidos demonstram boa reprodutibilidade intra e interobservador da segmentação manual utilizando o software 3D Slicer validando dessa forma esse método como ferramenta confiável para servir de padrão de referência em futuros estudos quantitativos desses tumores. Foi encontrada concordância quase perfeita entre as segmentações realizadas pelo método semiautomático quando comparadas com as segmentações realizadas pelo método manual, mas nossos resultados não demonstraram diferença significativa de tempo de segmentação do método semiautomático em relação ao método manual. / Soft tissue sarcomas constitute a diverse group of neoplasms that can arise in the connective tissues from virtually any region of the body. Magnetic resonance imaging (MRI) is currently the examination of choice for detection, regional staging and followup of these tumors. The segmentation of the lesions from the routine MR images allows the extraction of quantitative data, which has the potential to add information to the analysis. The purpose of the study is to evaluate the reproducibility of manual segmentation of soft tissue sarcomas in MRI images of patients with definitive diagnosis confirmed by histopathology. As a secondary objective, a comparison of manual and semiautomatic segmentation was performed to validate semiautomatic segmentation as an alternative method for segmentation of these tumors. We studied a retrospective cohort of 15 consecutive patients with confirmed diagnosis of soft tissue sarcoma accompanied at our service from January 2006 to January 2016 with MR images acquired prior to treatment and available for analysis in the DICOM format. The software was used 3D Slicer to perform segmentation by manual and semiautomatic methods. Three radiologists did the segmentations independently and blindly to allow inter-observer evaluation. The results obtained show high intraobserver reproducibility with Dice similarity coefficient between the segmentations ranging from 0.849 to 0.979 and Hausdorff distances ranging from 3.53 mm to 20.96 mm and good interobserver reproducibility with Dice similarity coefficients ranging from 0.741 to 0.972 and Hausdorff distances varying from 5.83 to 60.84 mm. A substantial agreement was found between the segmentations performed by the semiautomatic method when compared to the segmentations performed by the manual method with Dice similarity coefficients ranging from 0.871 to 0.973 and Hausdorff distances ranging from 5.43 mm to 31.75 mm. Regarding the segmentation time, there was no statistically significant difference of the semiautomatic method when compared to the manual method (p> 0.05). The volumes obtained in the different segmentations were also calculated and there was almost perfect agreement between the two manual segmentations performed by the radiologist 1, between the segmentations performed by radiologist 1 and radiologist 2, between the segmentations performed by radiologist 1 and radiologist 3, and between The manual and semi-automatic segmentation performed by the radiologist 1, and intraclass correlation coefficients (ICC) between 0.9927 and 0.9990 were obtained. The results obtained demonstrate good intra and interobserver reproducibility of the manual segmentation using 3D Slicer software, thus validating this method as a reliable tool to serve as a reference standard in future quantitative studies of these tumors. Almost perfect agreement was found between the segmentations performed by the semiautomatic method when compared to the segmentations performed by the manual method, but our results did not show a significant difference in segmentation time of the semiautomatic method in relation to the manual method.

Mechanistic insights on molecular and cellular mechanisms whereby KSHV induces Kaposi?s sarcoma (KS) are key for our understanding of KS tumors and for the development of new therapies. We have previously developed an animal model for KSHV induced KS using murine bone marrow cells transfected with a KSHVBac36. We found that although these cells lacked attributes of transformed cells in vitro, they were able to cause KS-like tumors in vivo. In vivo tumorigenesis correlated with upregulation of both KSHV lytic genes and host angiogenesis suggesting that that cues provided from the microenvironment played a major role in regulating viral and host gene expression related with KSHV-induced tumorigenesis. Our goal thus, was to identify these molecular cues regulating pathogenic gene expression in KSHV infected cells in vivo. An important difference between cells kept in vitro versus in vivo is the lack of environmental extracellular matrix (ECM) signals. Therefore the mECK36 cells were cultured in vitro in matrigel, a basement membrane preparation rich in ECM proteins and its individual components, to discern the effect of host signaling by the ECM on KSHV infected cells. Investigation of gene expression through Real Time RT-PCR identified several viral and host genes associated with tumorigenesis such as KSHV vGPCR and angiogenesis associated VEGF and EGF- receptors were upregulated in response to this environment. Further analysis of the molecular activity of the cell indicated the change in transcription was due to the activation of integrin signaling, as assessed by phosphorylation of the Focal Adhesion Kinase (FAK) protein. Our results show that integrin signaling occurring in vivo through interaction with ECM serves to enhance the pathogenic viral and host gene expression of KSHV infected cells and that EGFR upregulation can be correlated with these conditions. These results points to the integrin signaling pathway or the EGF-Receptor as promising targets for therapy and prevention of KS tumors.

Tumours contain heterogeneous cell populations. A population enriched in tumour-initiating potential has been identified in soft-tissue sarcoma (STS) by the isolation of "side population" (SP) cells. In this study, we compared the gene expression profiles of SP and non-SP cells in STS and identified Hedgehog (Hh) and Notch pathways as potential candidates for the targeting of SP cells. Upon verification of the activation of these pathways in SP cells, using primary tumor xenografts in NOD-SCID mice as our experimental model, we used the Hh blocker Triparanol and the Notch blocker DAPT to demonstrate that the suppression of these pathways effectively depleted the abundance of SP cells, reduced tumour growth, and inhibited the tumour-initiating potential of the treated sarcoma cells upon secondary transplantation. The data provide additional evidence that SP cells act as tumour initiating cells and points to Hh and Notch pathways as enticing targets for developing potential cancer therapies.

Tumours contain heterogeneous cell populations. A population enriched in tumour-initiating potential has been identified in soft-tissue sarcoma (STS) by the isolation of "side population" (SP) cells. In this study, we compared the gene expression profiles of SP and non-SP cells in STS and identified Hedgehog (Hh) and Notch pathways as potential candidates for the targeting of SP cells. Upon verification of the activation of these pathways in SP cells, using primary tumor xenografts in NOD-SCID mice as our experimental model, we used the Hh blocker Triparanol and the Notch blocker DAPT to demonstrate that the suppression of these pathways effectively depleted the abundance of SP cells, reduced tumour growth, and inhibited the tumour-initiating potential of the treated sarcoma cells upon secondary transplantation. The data provide additional evidence that SP cells act as tumour initiating cells and points to Hh and Notch pathways as enticing targets for developing potential cancer therapies.

Predictors of response of AIDS-associated Kaposi-Sarcoma to standard chemotherapy Overview: Kaposi Sarcoma is the most common HIV-associated cancer. Its etiology and pathogenesis is not fully understood. Little is known about what predicts prognosis, survival and therapeutic response in HIV-KS. In South Africa given the high seroprevalence rates of HIV-l and human herpes virus 8 (HHV 8), Kaposi's sarcoma is a significant problem. The majority of patients have been treated solely with palliation due to the poor outcome associated with a diagnosis of HIV-KS, more so in the absence of highly active antiretroviral therapy (HAART). Since the national ARV rollout programme and the availability and accessibility of HAART to all patients with a diagnosis of HIV-KS, a new strategy has to be established to enable adequate patient selection for chemotherapy. There have been a few published studies addressing the predictors of response to chemotherapy in the first world. However, this is the first study of these factors in HIV-l infected African patients with Kaposi's sarcoma. Aim: To identify and assess the potential value of several parameters predictive of outcome, survival and therapeutic response in HIV- infected patients with KS. Clinical, hematological, biochemical, immunological and virological variables were evaluated. Methods: We collected data from 25 patients with AIDS-KS who were enrolled in a phase III randomized controlled trial comparing HAART alone with the combination of HAART and chemotherapy. All patients were from the combination therapy arm. The following variables were evaluated as predictors of prognosis and therapeutic response: age, gender, ethnic origin, Haemoglobin (Hb), white blood cells (WBCs), lymphocytes, neutrophils, platelets, S.albumin, ALP, GGT, CD4 count, HIV viral load. These variables were assessed in patients at baseline and month 6 of therapy. Patients were staged into good risk and poor risk according to the AIDS clinical trial group (ACTG) criteria. The outcomes assessed were response to treatment and mortality. Results: A total of 25 patients participated to the study. Of these 16(64%) were males and 9(36%) were females, with male: female ratio of 2.7:1. Median age was 34 years (24-47); all patients were of Black African origin. Of the 21 patients, 15 (71.4%) were of good prognosis and 6(28.6%) were of poor prognosis. At baseline the median values of the different variables were as follows: Hb 10.9 g/dl, WBCs 5.95x109/L, lymphocytes 1.7 x109/L, neutrophils 3 x10 9 /L, platelets 272 x10 9 /L, S.albumin 30 gil, total protein 88 gil, ALP 64 U/L, and GTT 21 U/L, CD4 count was 255 cells/mm 3 , HIV-RNA viral load was 42000( 4.610gs). At month 6, 22 patients remained alive, their median values were: Hb 12.2 g/dl, WBCs 4.65 x109/L, lymphocytes 1.5 x109/L, neutrophils 3 x10 9 /L, platelets 301 x109/L, S.albumin 36.5 gil, total protein 84.5 gil, ALP 78.5 U/L, GTT 44.5 U/L, CD4 count 288 cells/mm3 , HIV-RNA viral load was 50500( 4.6910gs). The baseline median CD4 and HIV-RNA viral load counts for the 3 patients who died before month 6 were 47 cells/mm3 and 31000(4.610gs); respectively. Response to therapy was evaluated in 21(84%) patients as 4(16%) patients were missing, of the 21 patients 3 (14.3%) had complete response and 18(85.7%) had partial response. With respect to sex 2(14.3%) males had complete response and 12(85.7%) had partial response, 1(14.3%) female had complete response and 6 (85.7%) had partial response. Non-parametric statistics were used because of the small sample size and the skewness of the data. Variables were described using medians and ranges, and compared between two independent groups using Mann-Whitney tests. Baseline and month 6 comparisons were done using Wilcoxon signed ranks tests. Receiver Operating Characteristic (ROC) curves were used to analyze cut points to optimize sensitivity and specificity of a quantitative variable for a dichotomous outcome. Discussion In the univariate analysis age and sex didn't influence prognosis and therapeutic response, the influence of ethnic origin couldn't be assessed as all patients were of the same ethnic origin. Baseline WBCs (P= 0.004) and lymphocytes (P=0.026) were significantly associated with complete response. Higher values of GGT (p=O.OOl); ALP (P=0.006) were associated with more deaths. Baseline CD4 count and HIV viral load were not of predictive value, lthough change CD4 (P=002) and VL (p=.OOO) over time was significant and most likely attributed to response to therapy. 90.9 % of patients reached undetectable HIV-l Viral loads at month 6. CONCLUSION: Neither CD4 count nor HIV viral load at baseline predicted prognosis or survival; however there was a borderline significance of CD4 (P=0.058) towards a better survival. / Thesis (M.Med.)-University of KwaZulu-Natal, Durban, 2006.

Acute oncogenic stress can activate autophagy and facilitate permanent arrest of the cell cycle through a failsafe mechanism known as oncogene-induced senescence (OIS). Kaposi's sarcoma-associated herpesvirus (KSHV) is the etiologic agent of Kaposi’s Sarcoma (KS) and has been reported to encode oncoproteins within its latency program that engage host autophagy and OIS pathways; however, the mechanisms by which KSHV oncoproteins promote KS tumorigenesis remain unclear. Here, I demonstrate that ectopic expression of the latent KSHV protein viral cyclin (v-cyclin) deregulates the cell cycle, induces DNA-damage responses (DDRs) and promotes OIS through an autophagy-dependent mechanism. During latency, v-cyclin is co-expressed from a single transcript with a viral homolog of FLICE-inhibitory protein (v-FLIP) that blocks autophagy by binding and inhibiting Atg3. Co-expression of v-FLIP has no effect on DDRs, but efficiently blocks v-cyclin-induced autophagy and senescence. Remarkably, suppression of v-FLIP function during KSHV latency, through specific inhibitory peptides, rescues host cell autophagy and induces senescence of infected cells. Together, these data reveal a coordinated viral gene-expression program that subverts autophagy, impairs senescence, and facilitates the proliferation of KSHV-infected cells.

Diss. (sammanfattning) Stockholm : Karol. inst., 2004. / Härtill 4 uppsatser.

Lic.-avh. (sammanfattning) Stockholm : Karolinska institutet, 2006. / Härtill 3 uppsatser.

Os sarcomas ósseos representam uma proporção significativa de tumores na faixa etária pediátrica, ainda apresentando um quadro desafiador devido a sua significativa taxa de morbimortalidade. Pesquisas para o desenvolvimento de novas modalidades terapêuticas e para o desenvolvimento de métodos que identifiquem características da doença que possam permitir melhor estratificação dos pacientes através de dados clinicamente relevantes para individualizar as condutas clínicas são necessárias. Dentro deste contexto surge o conceito de radiômica, que visa extrair dados clinicamente relevantes a partir de imagens médicas. Entretanto, para colocar a radiômica em prática, é necessário selecionar, nas imagens médicas, as áreas de interesse referentes às patologias estudadas, e este processo se denomina segmentação. O objetivo primário deste estudo foi avaliar a reprodutibilidade intra e inter-observador da segmentação manual de sarcomas ósseos em imagens de ressonância magnética (RM). Como objetivo secundário, foi avaliada a capacidade da segmentação semiautomática em reduzir o tempo necessário para segmentação, mantendo similaridade com a segmentação manual. O estudo foi realizado de forma retrospectiva com inclusão de pacientes com diagnóstico de osteossarcoma ou sarcoma de Ewing confirmado por estudo histopatológico e que tivessem imagens de RM realizadas no Hospital Universitário de nossa Instituição realizadas previamente a qualquer intervenção terapêutica. Três médicos radiologistas, de forma independente e às cegas em relação as demais segmentações e em relação ao resultado histopatológico, realizaram a segmentação manual dos contornos destes tumores utilizando o software 3DSlicer, permitindo que fosse realizada avaliação da reprodutibilidade interobservador. Um dos radiologistas realizou uma segunda segmentação manual dos mesmos casos, possibilitando a avaliação da reprodutibilidade intraobservador, e, ainda, uma terceira segmentação foi realizada, utilizando metodologia semiautomática, disponível no software mencionado. Para a análise estatística, foi utilizado o coeficiente de similaridade de Dice (DICE), a distância Hausdorff (DH), comparações de volumes e análises dos intervalos de tempo necessários para realização das segmentações. Os parâmetros avaliados demonstraram haver boa reprodutibilidade intraobservador, com DICE variando entre 0,83 a 0,97; e distância Hausdorff variando entre 3,37 a 28,73 mm. Também foi demonstrada boa reprodutibilidade interobservador com DICE variando entre 0,73 a 0,97; e distância Hausdorff variando entre 3,93 a 33,40 mm. A segmentação semiautomática demonstrou boa similaridade em relação à segmentação manual (DICE variando entre 0,71 a 0,96 e DH variando entre 5,38 a 31,54 mm), havendo redução significativa do tempo necessário para segmentação. Entre todas as situações comparadas, os volumes não apresentaram diferenças estatisticamente significativas (p-valor>0,05). / Bone sarcomas represent a significant proportion of tumors in the pediatric age group and they still are a challenge due to their significant morbidity and mortality rates. Reseaches are important for the development of new therapeutic modalities and for the development of methods that identify features that allow better stratification of the patients with theses diseases for individualization of their treatments. In this context emerges the concept of radiomics, which is the process of extraction of clinically relevant data from medical images. It is important to segment the areas of interest im medical images for the pratice of this process. The primary objective of this study was to evaluate the intra- and interobserver reproducibility of manual segmentation of bone sarcomas on magnetic resonance imaging (MRI). As a secondary objective, it was evaluated if the semiautomatic segmentation could be similar to manual segmentation and if the semiautomatic method could reduce the time required for segmentation. The study was performed retrospectively with the inclusion of patients with osteosarcoma or Ewing sarcoma confirmed by histopathological study and who had MRI performed at the University Hospital of our Institution prior to any therapeutic intervention. Three radiologists, independently and blindly in relation to the other segmentations and in relation to the histopathological results, performed the manual segmentation of the contours of these tumors using 3DSlicer software, allowing an interobserver reproducibility evaluation. One of the radiologists performed a second manual segmentation of the same cases, allowing the evaluation of intraobserver reproducibility. A third segmentation was performed, using semi-automatic methodology, available in the mentioned software. For the statistical analysis, Dice similarity coefficient (DICE), Hausdorff distance (DH), comparisons between volumes and time intervals for segmentations were used. The parameters evaluated demonstrated a good intraobserver reproducibility, with DICE ranging from 0.83 to 0.97 and Hausdorff distance ranging from 3.37 to 28.73 mm. Good interobserver reproducibility was also demonstrated with DICE ranging from 0.73 to 0.97 and Hausdorff distance ranging from 3.93 to 33.40 mm. Semiautomatic segmentation demonstrated good similarity to manual segmentation (DICE ranging from 0.71 to 0.96 and HD ranging from 5.38 to 31.54mm), and there was significant reduction in the time required for segmentation. Among all the situations compared, the volumes did not present significant statistical differences (p-value> 0.05).

Orientador: Silvia Regina Brandalise / Dissertação (mestrado) - Universidade Estadual de Campinas, Faculdade de Ciencias Medicas / Made available in DSpace on 2018-08-09T02:02:46Z (GMT). No. of bitstreams: 1 Deneno_BiancaPratelezzi_M.pdf: 2701966 bytes, checksum: 77adce634cd407570541182cd2a6232b (MD5) Previous issue date: 2007 / Resumo: Os tumores ósseos malignos perfazem 7% dos tumores em pacientes menores de 20 anos de idade, sendo 10 a 15% deles, localizados em pelve. Com o advento da poliquimioterapia, radioterapia e novas técnicas cirúrgicas, a taxa de sobrevida livre de doença, em cinco anos, das crianças e adolescentes acometidos por tumor ósseo não metastático, está ao redor de 70%. Atualmente, pode ser observado o impacto dos efeitos estéticos e funcionais decorrentes do tratamento, e sua interferência na qualidade de vida desses indivíduos. O objetivo do presente estudo foi avaliar retrospectivamente, os resultados funcionais em crianças e adolescentes portadores de tumores pélvicos, submetidos à hemipelvectomia interna tipo II, com ou sem reconstrução do anel pélvico. Foram avaliados 31 pacientes portadores de tumor pélvico, tratados no Centro Boldrini e no Hospital A.C. Camargo, entre 1994 e 2005. O resultado funcional foi baseado no Sistema de Avaliação Funcional, padronizado por Enneking et al (1993). Os critérios analisados foram: dor, função, aceitação emocional, necessidade de suporte, capacidade de deambulação e marcha. Dos 31 casos seguidos, 12 (38,7%) fizeram a reconstrução do anel pélvico com enxerto de fíbula e 19 (61,3%) não a fizeram. A média de idade dos pacientes foi de 11,4 anos (4-17,8 anos). A média de seguimento para os 31 casos foi de 41 meses. O resultado funcional final obtido aos 12 meses do pós-operatório, foi excelente em 17,4 % dos pacientes (todos com reconstrução do anel pélvico), bom em 60,9% dos pacientes, regular em 17,4% e ruim em 4,3%. A média do escore foi melhor no grupo de pacientes com a reconstrução com autoenxerto de fíbula, comparativamente ao grupo de pacientes que não tiveram esta reconstrução, sendo significativa a diferença entre os grupos (p=0,008) / Abstract: Seven percent of all malignant tumors in patients less than 20 years old are bone tumors, being 10 to 15% of them localized in pelvis. The overall survival of these non metastatic patients, with polichemoterapy, radiation and new surgery approach is 70% in 5 years. Nowadays, the impact of esthetical and functional effects due to treatment interfere in their quality of life. The objective of this study was to assess, retrospectively, the functional results in children and adolescents with pelvic tumors, who underwent type II internal hemipelvectomy, with or without reconstruction of the pelvic ring. Thirty-one patients with pelvic tumor were treated at Boldrini¿s Hospital and A.C. Camargo Hospital during the period of 1994 and 2005. The functional result was evaluated based on the System for the Functional Evaluation, standardized by Enneking et al (1993). The analyzed criteria were: pain, function, emotional acceptance, need to support, walking ability and gait. Out of the 31 cases analyzed, 12 (38,7%) underwent reconstruction of the pelvic ring with fibular graft and 19 (61,3%) did not have reconstruction. The mean age of the patients was 11. 4 years (4 - 17,8 years). The mean of follow-up for the 31 cases was 41 months. The functional result, obtained 12 months after the surgery, was excellent in 17,4 % of the patients (all of them with pelvic ring reconstruction), good in 60,9% of the patients, regular in 17,4% and poor in 4,3%. The mean of score was better in the group of patients who underwent reconstruction with fibular autograft, comparatively to the group of patients who did not have pelvic ring reconstruction. The global functional result showed significant difference among the groups with and without reconstruction of the pelvic ring (p = 0,008) / Mestrado / Saude da Criança e do Adolescente / Mestre em Saude da Criança e do Adolescente