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Historiquement, SOCS1 a été décrite comme une protéine capable de réguler négativement la signalisation induite par les cytokines. Dès lors, un vif engouement s’est porté sur cette protéine quant à un probable rôle de suppresseur de tumeur, notamment grâce à ses fonctions anti-inflammatoires. De surcroit, son expression est fréquemment perdue dans de nombreux cancers des suites de phénomènes épigénétiques. Récemment, il a même été démontré que SOCS1 pouvait rentrer en complexe protéique avec le suppresseur de tumeur P53, et était requis pour son activité transcriptionnelle. Finalement, notre laboratoire a participé à démontrer que SOCS1 inhibait la signalisation et les fonctions du récepteur MET dans les hépatocytes. Étant donné l’influence bien connue de l’inflammation, de l’épigénétique, de P53 et de MET dans le cancer colorectal (CCR), nous avions émis l’hypothèse que SOCS1 agirait en tant que suppresseur de tumeur dans le CCR. Dans la présente étude, nous avons modulé l’expression de SOCS1 dans un modèle cellulaire de progression tumorale colique, soit les cellules CT36 et CT26. Puis, les caractéristiques biologiques dites cancéreuses de ces lignées ont été évaluées in-cellulo et in-vivo. Contre toute attente, nos résultats révèlent une action pro-tumorale menée par SOSC1 dans le CCR. La surexpression de SOCS1 dans les cellules CT26 provoque une augmentation de la croissance cellulaire, de la survie sans ancrage à la matrice extracellulaire et de la capacité à former des tumeurs in vivo. Et à l’inverse, sa diminution, par ARN interférent, provoque chez les cellules CT36 une baisse de ces mêmes fonctions cancéreuses. Au niveau moléculaire, nous avons validé que SOCS1 conservait bien son aptitude à inhiber les voies de signalisation des cytokines et du récepteur MET ainsi qu’à promouvoir l’activité de p53. Nous avons aussi identifié que SOCS1 diminue l’expression des protéines STAT1, P21cip/kip, et de la protéine adaptatrice P66SHC. En conclusion, ce travail de recherche révèle, pour la première fois, que l’expression de SOCS1 favoriserait le développement de certains CCR. Les protéines STAT1, P21 cip/kip et P66SHC étant connues pour être impliquées dans des processus antiprolifératifs et pro-apoptotiques, leurs régulations négatives par SOCS1 pourraient en être les raisons.

The transmembrane isoform of mucin 1 (MUC1/TM) is a well recognized tumor antigen, contributing to tumorigenesis and immune evasion. While MUC1/TM has been correlated with malignancy, it appears that a secreted splice variant of MUC1 (MUC1/sec) has antitumor properties and prevents tumor development. It was discovered that MUC1/sec expressing tumor cells (DA-3/sec) have a significant reduction in expression of urokinase plasminogen activator (uPA) relative to the parental tumor line, and tumor cells expressing MUC1/TM (DA-3/TM). The serine protease uPA, has been found to be involved in growth promoting signaling, angiogenesis, and induction of matrix remodeling leading to metastasis. Furthermore, the tumor suppressive and interferon responsive Stat1 transcription factor is dramatically upregulated in DA-3/sec cells. In addition, treatment of various murine and human cell lines with conditioned media containing MUC1/sec results in up-regulation of Stat1. DA-3/sec tumor cells are also sensitized to the anti-proliferative effects of IFN-g. Furthermore, transfection of the Stat1 gene into DA-3 tumor cells leads to a downregulation of uPA, and delays tumor progression. Since myeloid-derived suppressor cells (MDSC) play a critical role in tumor-induced immunosuppression, we investigated their recruitment by DA-3/sec and DA-3/TM cells. DA-3/sec tumor cells recruit dramatically lower levels of MDSC, relative to DA-3/TM cells. Since MUC1/sec down-regulates tumor expression of uPA, its potential role in MDSC recruitment was investigated. Tumor-derived uPA is capable of recruiting MDSC, and correlates with tumor development. In addition to diminishing recruitment of MDSC, the effect of MUC1/sec on MDSC suppressive mechanisms was investigated. MUC1/sec, or its unique immunoenhancing peptide (IEP), is capable of blocking expression of arginase 1 and production of reactive oxygen species (ROS) in MDSC, implicated in the suppression of T cells. These findings demonstrate a new mechanism of MDSC recruitment, and provide evidence that MUC1/sec has antitumor properties affecting both tumor cells and MDSC. Furthermore, it was discovered that MDSC home to the liver in addition to the tumor, bone marrow, blood, and spleen of tumor bearers, as previously described. The liver is thus an organ where MDSC accumulate and can contribute to immunosuppression directly and indirectly, via interactions with a variety of immune cells.

Enterohemorrhagic Escherichia coli (EHEC) serotype O157:H7 is a foodborne pathogen that causes significant morbidity and mortality in developing and industrialized nations. EHEC infection of host epithelial cells is capable of inhibiting the interferon gamma (IFNγ) pro-inflammatory pathway through the inhibition of Stat-1 phosphorylation, which is important for host defense against microbial pathogens. The aim of this thesis was to determine the bacterial factors involved in the inhibition of Stat-1 tyrosine phosphorylation. Human HEp-2 and Caco-2 epithelial cells were challenged directly with either EHEC or bacterial culture supernatants, stimulated with IFNγ, and then protein extracts were analyzed by immunoblotting. The data showed that IFNγ-mediated Stat-1 tyrosine phosphorylation was inhibited by EHEC secreted proteins. Using 2D-Difference Gel Electrophoresis, EHEC Shiga toxins were identified as candidate inhibitory factors. EHEC Shiga toxin mutants were then generated, complemented in trans, and mutant culture supernatant was supplemented with purified Stx to confirm their ability to subvert IFNγ-mediated cell activation. I conclude that E. coli-derived Shiga toxins represent a novel mechanism by which EHEC evades the host immune system.

Enterohemorrhagic Escherichia coli (EHEC) serotype O157:H7 is a foodborne pathogen that causes significant morbidity and mortality in developing and industrialized nations. EHEC infection of host epithelial cells is capable of inhibiting the interferon gamma (IFNγ) pro-inflammatory pathway through the inhibition of Stat-1 phosphorylation, which is important for host defense against microbial pathogens. The aim of this thesis was to determine the bacterial factors involved in the inhibition of Stat-1 tyrosine phosphorylation. Human HEp-2 and Caco-2 epithelial cells were challenged directly with either EHEC or bacterial culture supernatants, stimulated with IFNγ, and then protein extracts were analyzed by immunoblotting. The data showed that IFNγ-mediated Stat-1 tyrosine phosphorylation was inhibited by EHEC secreted proteins. Using 2D-Difference Gel Electrophoresis, EHEC Shiga toxins were identified as candidate inhibitory factors. EHEC Shiga toxin mutants were then generated, complemented in trans, and mutant culture supernatant was supplemented with purified Stx to confirm their ability to subvert IFNγ-mediated cell activation. I conclude that E. coli-derived Shiga toxins represent a novel mechanism by which EHEC evades the host immune system.

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No description available.

No description available.

How CD4 T cells protect against influenza A virus is poorly understood. Here, we address two central questions to better understand how CD4 T cells contribute to immunity during primary and secondary infection. First, we investigate the CD4 T cell-intrinsic requirements for three major transcription factors associated with an antiviral T cell phenotype (termed ‘Th1'): STAT1, STAT4, and T-bet, in directing CD4 T cell responses. We show that STAT4-deficiency does not affect the phenotype or function of wildtype or T-bet-/- CD4 T cells while STAT1-/- cells are virtually undetectable in infected host mice. Depleting NK cells rescues the STAT1-/- cells that phenocopy the compromised Th1 identity of T-bet-/- cells. Finally, we show that cytokine-mediated STAT4 activation enhances infection-induced Th1-polarization and that engaging STAT1 and STAT4 during priming dramatically improves CD4 T cell antiviral capacity. These results are relevant to T cell-based vaccine strategies aiming to promote the most efficient anti-viral T cell responses. Second, we asked the extent to which the recall of influenza-specific lung-resident memory CD4 T cells (TRM) impact the generation of new primary anti-viral T cells. TRM rapidly induce local inflammatory responses that control infection before protective T cells activated in secondary lymphoid organs reach sites of infection. Whether antigen-sensing by TRM can impact T cell priming in secondary lymphoid organs is unclear. We show that activation of influenza-primed lung TRM by antigen delivered into the airways enhances the number and activation status of antigen-bearing dendritic cells in draining lymph nodes. This accelerates the priming of naïve T cells and enhances their recruitment to the lung. Importantly, this TRM-dependent circuit enables productive T cell responses even against levels of airways antigen too low to otherwise activate naïve T cells. This adjuvant-like impact of lung TRM highlights a novel integration of local and regional T cell immunity.

Le cancer du sein est le cancer le plus fréquent chez la femme. Malgré la chimiothérapie, certaines patientes ont une réponse incomplète et récidivent quelques années plus tard. Le but de ce travail était d’étudier les mécanismes moléculaires mis en place par les cellules tumorales en réponse aux traitements et leur rôle éventuel dans la récidive. La modélisation de ces événements a d’abord été réalisée grâce à des modèles expérimentaux appelés PDX (pour patient derived xenograft) impliquant la greffe de tumeurs de patientes chez la souris immunodéficiente. Une analyse transcriptomique a permis de mettre en évidence, au sein des PDXs qui répondent à la chimiothérapie, la surexpression précoce de gènes cibles de la voie interféron (IFN)/STAT1, suggérant que cette signature moléculaire pourrait être utilisée comme biomarqueur prédictif de la réponse initiale au traitement. Cette signature persiste néanmoins dans les cellules tumorales résiduelles, suggérant qu’elle pourrait également avoir un rôle dans la récidive observée pour l’ensemble des PDX étudiées. L’analyse fonctionnelle de cette signature IFN/STAT1 a été réalisée in vitro, en traitant diverses lignées de cancer du sein par du mafosfamide, principe actif d’une chimiothérapie classique induisant des dommages à l’ADN. Nos résultats montrent que, sous chimiothérapie, certaines lignées (dont les cellules MFC7) sont capables d’exprimer des IFNs de type I conduisant à l’activation autocrine/paracrine de la voie IFN/STAT1. Une étude mécanistique a mis en évidence l’implication du détecteur de l’ADN STING (stimulator of IFN genes) en amont de la production d’IFN, comme cela a été bien décrit dans des cellules immunitaires en présence de pathogènes. Nous avons montré que l’inhibition individuelle de l’expression de différents acteurs de la voie STING/IFN/STAT1 potentialise l’effet de la chimiothérapie, impliquant cette cascade dans la résistance au traitement. En résumé, notre travail suggère que l’activation de la voie STING/IFN/STAT1 dans les cellules tumorales mammaires traitées par chimiothérapie pourrait jouer un double rôle : biomarqueur de la réponse dans un premier temps, puis acteur de résistance dans un second temps. Ce travail ouvre de nouvelles perspectives pronostiques et thérapeutiques pour la prise en charge du cancer du sein. / Breast cancer is the most frequent cancer in women. Despite chemotherapy, tumor response is often incomplete, and relapse is frequently observed. The aim of this work was to analyze the molecular mechanisms triggered in breast cancer cells in response to chemotherapy. These events were modeled using breast cancer patient-derived xenografts (PDXs), i.e. samples of human tumors engrafted into immunodeficient mice. Transcriptomic analysis highlighted precocious induction of the interferon (IFN)/STAT1 pathway in response to chemotherapy only in tumors responding to treatment, suggesting that this molecular signature could be used as a biomarker of the initial response. The activation of this pathway persisted in residual tumor cells, suggesting that it could also play a role in cancer recurrence observed in all PDX models that we investigated. Functional deciphering of the IFN/STAT1 pathway was performed in vitro by stimulating breast cancer cell lines with mafosfamide, the active principle of a classical chemotherapy inducing DNA damage. In some cell lines, e.g. MCF-7 cells, this treatment triggered the upregulation of type I IFN expression leading to cell-autonomous activation of the IFN/STAT1 signaling pathway. A mechanistic study revealed the involvement of the DNA sensor STING (stimulator of IFN genes) upstream of the IFN production, mimicking what happens in immune cells facing pathogen infection. Individual silencing of various actors of the STING/IFN/STAT1 pathway potentiated genotoxic treatment efficacy, indicating that this cascade may be involved in tumor resistance to treatment. In summary, our study suggests that cell-intrinsic activation of STING/IFN/STAT1 pathway in response to chemotherapy could play a dual role: first, it may be used as a predictive biomarker of initial response; second, it may act as a resistance mechanism to treatment. This work opens new prognostic and therapeutic perspectives for the clinical management of breast cancer.