The nucleus of epidermal keratinocytes (KCs) is a complex and highly compartmentalized organelle, whose structure is markedly changed during terminal differentiation and transition of the genome from a transcriptionally active state seen in the basal and spinous epidermal cells to a fully inactive state in the keratinized cells of the cornified layer. Here, using multicolor confocal microscopy, followed by computational image analysis and mathematical modeling, we demonstrate that in normal mouse footpad epidermis, transition of KCs from basal epidermal layer to the granular layer is accompanied by marked differences in nuclear architecture and microenvironment including the following: (i) decrease in the nuclear volume; (ii) decrease in expression of the markers of transcriptionally active chromatin; (iii) internalization and decrease in the number of nucleoli; (iv) increase in the number of pericentromeric heterochromatic clusters; and (v) increase in the frequency of associations between the pericentromeric clusters, chromosomal territory 3, and nucleoli. These data suggest a role for nucleoli and pericentromeric heterochromatin clusters as organizers of nuclear microenvironment required for proper execution of gene expression programs in differentiating KCs, and provide important background information for further analyses of alterations in the topological genome organization seen in pathological skin conditions, including disorders of epidermal differentiation and epidermal tumors.

Poor drug penetration through tumor tissue has emerged as a fundamental obstacle to cancer therapy. The solid tumor microenvironment presents several physiological abnormalities which reduce the uptake of intravenously administered therapeutics, including leaky, irregularly spaced blood vessels, and a pressure gradient which resists transport of therapeutics from the bloodstream into the tumor. Because of these factors, a systemically administered anti-cancer agent is unlikely to reach 100% of cancer cells at therapeutic dosages, which is the efficacy required for curative treatment. The goal of this project is to use high-intensity focused ultrasound (HIFU) to enhance drug delivery via phenomena associated with acoustic cavitation. ‘Cavitation’ is the formation, oscillation, and collapse of bubbles in a sound field, and can be broadly divided into two types: ‘inertial’ and ‘stable’. Inertial cavitation involves violent bubble collapse and is associated with phenomena such as heating, fluid jetting, and broadband noise emission. Stable cavitation occurs at lower pressure amplitudes, and can generate liquid microstreaming in the bubble vicinity. It is the combination of fluid jetting and microstreaming which it is attempted to explore, control, and apply to the drug delivery problem in solid tumors. First, the potential for cavitation to enhance the convective transport of a model therapeutic into obstructed vasculature in a cell-free in vitro tumor model is evaluated. Transport is quantified using post-treatment image analysis of the distribution of a dye-labeled macromolecule, while cavitation activity is quantified by analyzing passively recorded acoustic emissions. The introduction of exogenous cavitation nuclei into the acoustic field is found to dramatically enhance both cavitation activity and convective transport. The strong correlation between inertial cavitation activity and drug delivery in this study suggested both a mechanism of action and the clinical potential for non-invasive treatment monitoring. Next, a flexible and efficient method to simulate numerically the microstreaming fields instigated by cavitating microbubbles is developed. The technique is applied to the problem of quantifying convective transport of a scalar quantity in the vicinity of acoustically cavitating microbubbles of various initial radii subject to a range of sonication parameters, yielding insight regarding treatment parameter choice. Finally, in vitro and in vivo models are used to explore the effect of HIFU on delivery and expression of a biologically active adenovirus. The role of cavitation in improving the distribution of adenovirus in porous media is established, as well as the critical role of certain sonication parameters in sustaining cavitation activity in vivo. It is shown that following intratumoral or intravenous co-injection of ultrasound contrast agents and adenovirus, both the distribution and expression of viral transgenes are enhanced in the presence of inertial cavitation. This ultrasound-based drug delivery system has the potential to be applied in conjunction with a broad range of macromolecular therapeutics to augment their bioavailability for cancer treatment. In order to reach this objective, further developmental work is recommended, directed towards improving therapeutic transducer design, using transducer arrays for treatment monitoring and mapping, and continuing the development of functionalized monodisperse cavitation nuclei.

La sénescence cellulaire, ou l’arrêt irréversible de la prolifération, influence des processus physiologiques et pathologiques, comme le cancer. Parmi les caractéristiques de la sénescence, se retrouve le PSAS ou phénotype sécrétoire associé à la sénescence. Le PSAS est composé d’une variété de cytokines, facteurs de croissance et protéases. Ses actions pro- et anti-tumorale sont connues, mais l’on ignore laquelle prédomine. Mes travaux s’attardent aux effets du PSAS sur l’induction de la sénescence dans les cellules environnantes et à sa régulation. Nous avons démontré que le PSAS ne synergise pas avec la dysfonction télomérique chronique ou aigue, afin de causer un arrêt de croissance. Également, l’étude du mécanisme responsable de l’induction de la sénescence par stress à la chromatine, suggère que la kinase c-Abl n’est pas requise pour cette voie, contrairement à des publications antérieures. Mes travaux éclairent les mécanismes d’action et la régulation du PSAS dans la sénescence induite par dysfonction télomérique et par stress à la chromatine. / Cellular senescence, or irreversible proliferation arrest, is known for its influence on physiological and pathological processes, such as cancer. Among the features found in the senescent phenotype is the inflammatory secretome, also known as the senescence associated secretory phenotype (SASP). The SASP consists of a variety of factors such as cytokines, growth factors and proteases. It is widely recognized that SASP can have either a pro- or anti-tumor effect, but it is not clear which one predominates. My work focused on the SASP effects on the induction of senescence in surrounding cells and its regulation mechanisms. We demonstrated that the SASP does not synergize with chronic or induced telomere dysfunction to cause cellular proliferation arrest. Also, study of chromatin stress-induced senescence mechanism suggests that kinase c-Abl is not required for this pathway, contrary to what had been previously published. My work helps understand the regulatory and working mechanisms of the SASP in chromatin stress-induced and telomere dysfunction-induced senescence models.

Melanomas are one of the most aggressive types of tumours, with increasing incidence, high mortality and high potential to metastasize to a variety of diverse locations. The aim of this thesis was to study the tumour as a complex structure consisting not only of tumour cells but also of tumour stroma. Stromal cells play a major role in cancer biology. This is well documented for example in squamous cell epithelium tumours of the head and neck. Similar mechanisms can be expected to occur in melanomas. In the first experiment, we simulated the conditions in vivo during the metastatic process and studied the influence of non-adhesive environment both with and without the influence of stromal fibroblasts. The presented data demonstrates a change of tumour cells' phenotype leading to increased plasticity of the melanoma cells in these conditions. It also indicates the crucial role of stromal fibroblasts in interactions with melanoma cells. Cancer cell lines show variability in their behaviour, which is in accordance with well-known melanoma heterogeneity in clinical practice. The previous experiments in our laboratory indicate that cancer associated fibroblasts are able to influence the phenotype of a tumour cell line and this effect is based on a tumour type-unspecific mechanism. In the second part of...

La morbi-mortalité élevée des carcinomes épidermoïdes de la cavité orale (CECO), qui peuvent se développer à partir de lésions orales à potentiel malin (LOPM), rend indispensable le développement de nouvelles stratégies thérapeutiques. Le décryptage de l’hétérogénéité moléculaire aux différentes étapes de la carcinogénèse orale pourrait permettre de personnaliser les stratégies thérapeutiques de prévention et de traitement de ces cancers. Notre objectif était de caractériser l’hétérogénéité transcriptomique des LOPM et des CECO.Nous avons d’abord défini des signatures transcriptomiques associées aux changements histologiques de la carcinogénèse orale observés dans le modèle murin induit par le 4-NQO, montrant la pertinence de l’analyse de la dynamique temporelle du transcriptome pour améliorer la prévention des CECO. Cependant, ce modèle ne représentant qu’un sous-groupe particulier des CECO, nous avons ensuite étudié l’hétérogénéité inter-lésionnelle des LOPM en identifiant deux sous-types transcriptomiques principaux nommés « classical » et « immunological », qui sont caractérisés par différents biomarqueurs de risque de CECO.Au stade invasif, nous avons également étudié l’hétérogénéité transcriptomique des CECO HPV-négatifs entre les patients non-fumeurs non-buveurs (NFNB) et les patients fumeurs buveurs (FB). Le microenvironnement immunitaire était la principale différence biologique entre NFNB et FB, suggérant un bénéfice accru des immunothérapies chez les NFNB. Le profil transcriptomique de réponse antivirale observé dans les CECO des NFNB pourrait être en faveur de leur origine virale. En conclusion, l’hétérogénéité transcriptomique des LOPM et CECO suggère de personnaliser les stratégies thérapeutiques des patients porteurs de ces lésions / Oral squamous cell carcinomas (OSCC), which may develop from oral premalignant lesions (OPL), are associated with a substantial morbidity and mortality. A better understanding of the molecular heterogeneity at different steps of oral carcinogenesis may help to refine prevention and treatment strategies of patients suffering from OPL and OSCC. Our goal was to decipher transcriptomic hetereogeneity of OPL as well as OSCC. Using the 4-NQO murine model of oral carcinogenesis, we first identified transcriptomic signatures that characterized the dynamics of gene expression changes through different stages of disease progression, and that could be relevant for refining prevention strategies. Because this model represents only a subgroup of patients suffering from OSCC, we then investigated inter-OPL molecular heterogeneity. We identified two distinct gene expression subtypes, which were named classical and immunological and were characterized by different biomarkers of cancer risk. At invasive steps, we investigated transcriptomic heterogeneity between HPV-negative OSCC from never-smoker never-drinker (NSND) and smoker drinker (SD) patients. The immune microenvironment was the main biological difference between OSCC from NSND and SD, suggesting higher clinical benefit of immunotherapies in OSCC from NSND. The antiviral gene expression profile of OSCC from NSND could suggest a viral origin.In conclusion, we investigated transcriptomic heterogeneity of OPL as well as OSCC, that could help to refine their prevention and treatment strategies

Chez les patients, la tumeur développe une immunosuppression qui compromet les capacités des effecteurs de l’immunité à réagir. Longtemps décriée, l’immunothérapie est considérée aujourd’hui comme une thérapie à part entière avec la chirurgie, la chimiothérapie et la radiothérapie. Parmi les premières immunothérapies développées dans les années 1990, l’IL-2 forte dose fut approuvée dans le traitement du mélanome et du cancer du rein métastatiques. Cependant, sa forte toxicité et la faible proportion de patients répondeurs ont limité son usage clinique. Une autre cytokine, l’IL-15, apparaît prometteuse dans le traitement des cancers pour son activité similaire à l’IL-2 sans ses facteurs limitants. Cependant, seule, son activité anti-tumorale n’est pas optimale du fait de sa courte demi-vie ou encore de l’affinité intermédiaire de liaison à son récepteur βγ. Différentes stratégies ont été mises au point pour améliorer l’efficacité de l’IL-15, notamment, sa stabilisation à travers l’association avec sa chaine de haute affinité IL-15Rα. Une molécule de fusion associant de manière covalente la partie sushi+ de l’IL-15Rα avec l’IL-15 humaine a été développée : le RLI. Au cours de cette thèse, nous avons étudié chez la souris, le potentiel thérapeutique de cette molécule dans le traitement des cancers solides. Nous avons ainsi mis en évidence une activité anti-tumorale du RLI et une association synergique avec un anticorps monoclonal anti-PD-1. / In cancer patients the immune system is compromised by the tumor and its microenvironment. In recent decades the role of the immune system in tumor control has been controversial, though today cancer treatments that modulate immunity are a reality. Immunotherapy is a unique therapy adding to pre-existing methods of cancer control: surgery, chemotherapy and radiotherapy. In the 1990’s, high dosing of IL-2 was the first immunotherapy approved by the FDA to treat metastatic melanoma and renal carcinoma, however high toxicities and low responder rates have limited its clinical use. IL-15 is another promising cytokine therapy. The similar properties with IL-2 and differences in terms of toxicities and Treg induction make IL-15 an attractive potential therapy. Nonetheless, the short half-life and intermediate affinity for its receptor βγ compromise its efficacy. Different strategies have been developed to facilitate IL-15 therapeutic bioactivity. IL-15Rα as a chaperon molecule allows stability of IL-15 in vivo. RLI is a fusion molecule that covalently links sushi+IL-15Rα, the binding domain of its high affinity receptor and a recombinant human IL-15. We have studied the therapeutic potential of RLI in mouse tumor models. Our results show anti-tumor activities of RLI and synergistic combination with anti-PD-1, a monoclonal antibody.

Les phénomènes cicatriciels et de carcinogenèse partagent des points communs et peuvent être définis comme des processus complexes et adaptatif régulés par les interactions entre l'hôte et le microenvironnement tissulaire. Après la chirurgie, la radiothérapie est la seconde modalité la plus efficace dans le traitement du cancer et une approche thérapeutique multimodale impliquant chirurgie, radiothérapie et chimiothérapie est aujourd’hui classiquement utilisée. Des résultats récents suggèrent que, en plus des effets létaux sur les cellules tumorales, la radiothérapie modifie le microenvironnement tissulaire. Ces modifications affectent le phénotype cellulaire, le métabolisme des tissus, et les événements de signalisation entre les cellules.Les interactions complexes entre les cellules stromales et les cellules cancéreuses suscitent beaucoup d’intérêt et dans la première partie de ma thèse, j’ai exploré les interactions entre stroma et cellules de carcinome en réponse à la radiothérapie par modulation génétique du stroma après irradiation. J’ai constaté que les fibroblastes, indépendamment de leur statut RhoB, ne modulaient pas la radiosensibilité intrinsèque des TC- 1, mais produisaient des facteurs diffusibles capables de modifier le devenir des cellules tumorales. Ensuite, j’ai constaté que fibroblastes sauvages et RhoB déficients stimulaient la migration des TC-1 par des mécanismes distincts, impliquant respectivement TGF- β1 et MMP. J’ai également constaté que la co-irradiation, des fibroblastes et des TC- 1, abrogait le phénotype pro-migratoire des TC-1 par répression de la sécrétion du TGF- β et des MMP. Alors que le protocole de co-irradiation utilisé mime la situation clinique, mes résultats sont en désaccord avec les publications récentes et suggèrent que le stroma irradié ne renforce pas la migration des cellules tumorales mais au contraire pourrait être manipulé pour promouvoir une réponse immunitaire anti-tumorale.Deuxièmement, mes expériences in vivo, semblent confirmer les données obtenues in vitro et montrent que l’irradiation préalable du lit tumoral ne stimule ni croissance de la tumeur et ni sa dissemination. Nos résultats semblent montrer que l’irradiation du stroma ne favorise pas la migration des cellules de carcinome et ceci indépendamment de leur génotype. La Troisième Partie De Mon Projet, a été consacrée à l’étude des cellules tumorale circulantes (CTC) après la radiothérapie. En accord avec les résultats rapportés après la chirurgie, le nombre de CTC augmente dans la circulation sanguine après radiothérapie probablement à cause des lésions vasculaires radio-induites ou/et par induction d’EMT dans les cellules tumorales. Néanmoins ces CTC semblent être piégées dans la cavité cardiaque. La signification de la présence de ces CTC pour le développement métastatique n’est pas élucidée mais on peut suspecter un effet promoteur de métastase. Ainsi le microenvironnement pourrait avoir des effets antagonistes promoteurs ou inhibiteurs de malignité. / Wound healing and carcinogenesis are defined as complex, adaptive processes which are controlled by intricate communications between the host and the tissue microenvironment. A number of phenotypic similarities are shared by wounds and cancers in cellular signaling and gene expression. Radiotherapy is the second most effective modality of cancer treatment after surgery and can be used, either alone or in combination with chemotherapy. Recent findings suggest that radiotherapy apart from tumor cell death also rapidly and persistently modifies the tissue microenvironment. These modifications affect cell phenotype, tissue metabolism, bidirectional exchanges and signaling events between cells. The complex interactions between stromal cells and cancer cells are of immense interest and in The First Part of My Thesis, I tried to explore the crosstalk between stromal and carcinoma cells in response to radiotherapy by genetic modulation of the stroma and irradiation. We found that fibroblasts, irrespective of their RhoB status, do not modulate intrinsic radiosensitivity of TC-1 but produce diffusible factors able to modify tumor cell fate. Then we found that Wt and RhoB deficient fibroblasts stimulated TC-1 migration through distinct mechanisms respectively, TGF-β1 and MMP-mediated. We also found that co-irradiation of fibroblasts and TC-1 abrogated the pro-migratory phenotype by repression of TGF-β and MMP secretion. This result is highly relevant to the clinical situation and suggests that conversely to, the current view; irradiated stroma would not enhance carcinoma migration and could be manipulated to promote anti-tumor immune response. Secondly, our in vivo experiments, tends to confirm the in vitro data showing that irradiated tumor bed does not stimulate tumor growth and escape. Our results also challenges the view that irradiated stroma would promote migration of carcinoma cells as we show that independently from their genotype co-irradiation of fibroblasts and carcinoma cells repressed carcinoma cell migration and confirmations studies are currently performed in vivo. The Third Part of My Project, was dedicated to investigate the effect on CTC release after radiotherapy. Consistently with the results reported after surgery , the number of CTC increases in the blood stream after radiotherapy probably due to radiation-induced vascular injury induced or/and by EMT induction in tumor cells but these cells seemed to be entrapped into the cardiac cavity. The significance of these CTC to metastatic development is still under investigation but there is evidence for a metastasis-promoting effect of RT from animal studies.Thus the microenvironment can exert antagonist stimulatory or inhibitory effects on malignant cells.

Submitted by Fabíola Silva (fabiola.silva@famerp.br) on 2016-09-29T20:17:35Z No. of bitstreams: 1 josehenriquemferreira_dissert.pdf: 1670640 bytes, checksum: 3707d57cc4f4e98d8edb6594786c1b7e (MD5) / Made available in DSpace on 2016-09-29T20:17:35Z (GMT). No. of bitstreams: 1 josehenriquemferreira_dissert.pdf: 1670640 bytes, checksum: 3707d57cc4f4e98d8edb6594786c1b7e (MD5) Previous issue date: 2014-10-20 / Introduction: Mammary neoplasms are the most common type of tumor in dogs. Some proteins play an important role in tumor progression, thus are candidate prognosis markers. During tumor growth, the transcription factor induced by hypoxia (HIF-1α) activates the expression of vascular endothelial growth factor (VEGF), promoting angiogenesis. Interleukin-8 (IL-8) is a pro-inflammatory and pro-angiogenic cytokine and has been associated with tumor progression. Objectives: To evaluate the prognostic value of HIF-1α, VEGF and IL-8 proteins in tumor tissue in dogs with mammary tumors, correlating them with clinicopathological parameters, clinical outcome and survival. Material and Methods: The concentrations of HIF-1α, VEGF and IL-8 proteins were evaluated by ELISA (Enzyme-linked immunosorbent) in macerated tumor of 25 bitches with mammary tumors and control samples and compared statistically. Results: The levels of HIF-1α, VEGF and IL-8 were higher in dogs over the age of 10 years and that had died (p <0.05). Moreover, HIF-1α concentrations were elevated in the tumors of dogs who developed metastases (p = 0.04), while VEGF levels were highest in tumors with clinical stages III and IV (p = 0.03) and IL-8 tumors in tumor with development greater than six months (p = 0.03). Still, high levels of HIF-1α, VEGF and IL-8 were also related to shorter overall survival (p <0.05). Conclusions: High levels of HIF-1α, VEGF and IL-8 are associated with features of poor prognosis, suggesting that the assessment of these proteins in tumor macerated has important prognostic value. / Introdução: As neoplasias mamárias são o tipo mais comum de tumor na espécie canina. Algumas proteínas exercem importante papel na progressão tumoral e portanto, são candidatos marcadores de prognóstico. Durante o crescimento tumoral, o fator de transcrição induzido por hipóxia (HIF-1α) ativa a expressão do fator de crescimento endotelial vascular (VEGF), promovendo a angiogênese. A interleucina-8 (IL-8) é uma citocina pró-inflamatória e pró-angiogênica e tem sido associada à progressão tumoral. Objetivos: Avaliar o valor prognóstico das proteínas HIF-1α, VEGF e IL-8 no tecido tumoral em cadelas com neoplasia mamária, relacionando-os com os parâmetros clínico-patológicos, evolução clínica e sobrevida. Material e Métodos: As concentrações das proteínas HIF-1α, VEGF e IL-8 foram avaliadas pelo método de ELISA (Enzyme-linked immunosorbent Assay) no macerado tumoral de 25 cadelas com neoplasia mamária e amostras controle e comparadas estatisticamente. Resultados: Os níveis de HIF-1α, VEGF e IL-8 foram maiores em cadelas com idade superior a 10 anos e que vieram a óbito (p < 0,05). Além disso, concentrações de HIF-1α foram elevadas nos tumores de cadelas que desenvolveram metástase (p = 0,04), enquanto os níveis de VEGF foram maiores em tumores com estadiamento clínico III e IV (p = 0,03), e de IL-8 em tumores com evolução tumoral maior que seis meses (p = 0,03). Ainda, níveis elevados de HIF-1α, VEGF e IL-8 também foram relacionados com menor tempo de sobrevida global (p < 0,05). Conclusões: Altas concentrações de HIF-1α, VEGF e IL-8 estão associadas com características de pior prognóstico, sugerindo que a avaliação dessas proteínas no macerado tumoral possui importante valor prognóstico.

Introdução: Os cânceres de mama subtipos Luminal A e B (HER2 negativo) podem apresentar prognóstico variável, a depender do índice de proliferação, avaliado pelo Ki67. As células malignas e as células estromais adjacentes (fibroblastos e células de resposta imune ) podem interagir tanto pelo contato célula a célula como por fatores secretados por elas, ambas influenciando no comportamento tumoral. Já foi demonstrado que as células estromais podem aumentar a proliferação das células do câncer da mama. Objetivo: Nosso objetivo foi avaliar o perfil de expressão gênica de células do estroma em câncer de mama luminal A e luminal B e analisar se este se correlaciona com o prognóstico da doença. Pacientes e Métodos/ Resultados: Amostras de tumores de 11 pacientes na pós menopausa foram analisadas, todas elas HER2 negativas. A expressão de Ki67 foi <= 10 % em 5 pacientes (luminal A) e >= 30 % em outras 6 amostras(Luminal B ). Células estromais foram microdissecadas para a extração de RNA, que posteriormente foi hibridizado na plataforma de microarray Agilent G485 -1A GE 8x60K. Após a normalização, 50 % dos genes com a maior variância foram selecionados para análise por SAM duas classes desemparelhado (software TMEV ) e aceitando FDR 14.1%, 35 sequências foram identificadas como diferencialmente expressas, incluindo 16 genes conhecidos, entre as células estromais das amostras de Luminal A versos Luminal B, todos mais expressos nas amostras B. Dentre as funções biológicas enriquecidas em genes diferencialmente expressos encontram-se regulação positiva do sistema imune, incluindo genes como ZAP70 (proteína quinase 70kDa associada a cadeia zeta (TCR)), CD38 (molécula CD38); UBASH3A (ubiquitina associada e SH3 domínio que contém A); PLA2G7 (fosfolipase A2, grupo VII (fator acetil ativador de plaquetas no plasma)); NCR3 (citotoxicidade natural, provocando receptor 3). Nosso próximo passo foi avaliar se a expressão de alguns genes selecionados estava associada com prognóstico de tumores luminais. Para tal selecionamos amostras de outro grupo de 89 pacientes com seguimento de pelo menos 5 anos, cujos tumores eram ER(+), HER2(-), para análise de expressão proteica em Tissue microarray. Caracterizamos os fibroblastos destas amostras com 3 marcadores de fibroblastos: actina de músculo liso (AML), S100A4 e caveolina-1 (CAV1) e analisamos a marcação da proteína ZAP70. Correlacionamos a expressão proteica de todos os marcadores com as características anatomopatológicas da amostra. Observamos que fibroblastos de todas as amostras de tumor de mama expressam AML, S100A4 e CAV1, em diferentes proporções, entretanto não detectamos diferença entre os tumores luminais A e B. Também não obsevamos diferença de expressão de AML, S100A4 e CAV1 em relação a grau histológico, comprometimento linfonodal e estadiamento clínico. Nestas amostras não detectamos expressão proteica de ZAP70 em fibroblastos tumorais. Conclusão: Houve expressão diferencial de 16 genes relacionados a processos imunes, todos eles mais expressos em células estromais de tumores Luminal B em relação a luminal A / Introduction: Luminal breast cancer subtypes A and B (HER2 negative) may present a variable prognosis, depending on tumor proliferation index, evaluated by Ki67 expression. Malignant cells and adjacent stromal cells (fibroblasts and immune response cells) may interact by both cell contact and secreted factors and influence tumor behavior. It was shown that stromal cells may enhance breast cancer cells proliferation. Objective: Our aim was to evaluate stromal cells gene expression profile in luminal A and luminal B tumors and to evaluate whether selected transcripts expressed in stromal cells may be associated with prognosis in breast cancer. Material/ Methods and Results: Hormone receptor positive tumor samples from 11 post menopausal patients were analyzed, all of them Her2 negative. Ki67 expression <= 10% (luminal A) was observed in five and Ki67 >= 30% (luminal B) in six samples. Stromal cells were microdissected for RNA extraction, which was hybridized in Agilent G485-1A GE 8x60K microarray platform. After normalization, 50% of the genes with the highest variance were selected for further analysis by two class unpaired SAM (TMEV software) and accepting FDR 14,1%, 35 sequences, including 16 known genes, were found differentially expressed between stromal cells from luminal A vs luminal B breast cancer samples, all of them more expressed in luminal B. Among biological functions enriched in genes found differentially expressed were positive regulation of immune system process, including genes as: ZAP70 (zeta-chain (TCR) associated protein kinase 70kDa); CD38 (CD38 molecule); UBASH3A (ubiquitin associated and SH3 domain containing A); PLA2G7 (phospholipase A2, group VII (platelet-activating factor acetylhydrolase, plasma); NCR3 (natural cytotoxicity triggering receptor 3). Our next step was evaluate whether expression of selected genes was associated with prognosis in another group of patients. Tumor samples from 89 patients with at least 5 years of follow up, all of them estrogen receptor positive and HER2 negative, were selected. Tissue microarray was prepared with stromal tumor compartment from paraffin embedded tumor samples. Fibroblasts were characterized for the expression of 3 fibroblasts markers (alfa-SMA, alpha smooth muscel actin; S100A4 and CAV1, caveolin 1), and ZAP70. Correlation of expression of these markers with prognostic variables was determined. Expression of alfa-SMA, S100A4 and CAV1 was detected in fibroblasts from all tumor samples in different proportions, however no differential expression was observed between luminal A and B tumors. Neither difference was detected on the expression of these proteins in relation with histological grade, lymph node involvement and clinical stage. Conclusion: A differential expression of 16 genes involved in immune process was found, all of them more expressed in fibroblasts from luminal B as compared with luminal A tumors

No description available.