Colorectal cancer (CRC) is a major problem in affluent countries worldwide. In Australia it is the second most commonly diagnosed malignancy with approximately 13,000 new cases diagnosed each year. This disease is also the leading cause of cancer related death in Australia with approximately 4,500 fatalities each year. Epidemiological studies have shown geographical variation in the incidence of disease, with diet considered to be a key contributing factor to CRC risk. In particular, diets high in fibre and low in fat have been demonstrated to reduce the risk of developing CRC. Fibre is heterogeneous in nature and can be categorised into different subtypes. Resistant starch is a component of fibre which remains largely intact throughout the gastrointestinal tract until it reaches the colon. Here it undergoes bacterial fermentation to produce the short chain fatty acids (SCFAs) acetate, propionate and butyrate (BuA). Each of the SCFAs are bioactive in the colon, with the most active being BuA. The beneficial effects of fibre have been linked to BuA’s ability to induce colon cancer cell differentiation, reduce proliferation and initiate apoptosis. Interestingly, in normal cells BuA is utilised as the preferential energy source and has been shown to promote proliferation. With an apparent “paradoxical effect” on normal and cancerous cells BuA has been the subject of much investigation as a potential anticancer agent. Despite numerous studies investigating BuA actions, the exact biological mechanisms remain largely undefined. This thesis explored a possible mechanism for BuA-induced apoptosis and inhibition of proliferation. In 2003, two publications provided evidence that SCFAs, including BuA, were ligands to two members of a previously orphan family of G-protein coupled receptors (GPCRs); GPCR41 and 43. Of the two receptors BuA had the strongest effect on GPCR43. Consequently this thesis investigated the possibility that BuA acts to decrease CRC proliferation and induce apoptosis by binding to and activating GPCR43 on CRC cells. It was hypothesised that GPCR43 acted as a “BuA sensor” on the surface of the cell to mediate the effects of BuA. This experimental work utilised PCR, Q-PCR, measures of apoptosis, proliferation and differentiation and RNAi knockdown. The key areas of investigation included: (1) Determining if GPCR43 was present on a range of CRC cell lines with a cell line to represent adenocarcinoma, carcinoma and metastatic stage of disease. (2) Investigating the expression of GPCR43 with manipulated nutrient media and different levels of cell confluence. (3) Exploring GPCR43 expression in normal and malignant human patient biopsies. (4) Determining if the inhibition of G-protein function using inhibitors influenced BuAinduced changes to apoptosis and proliferation. (5) Using RNAi, investigating the effect that GPCR43 knockdown would have on BuA-induced changes to proliferation and apoptosis. The key findings from this work included: (1) Presence of GPCR43 on some but not all CRC cell lines. (2) Modulation of GPCR43 expression with exposure to BuA and altered glucose concentrations in the media. (3) An influence of G-protein inhibition on BuA-induced apoptosis but not proliferation in some cell lines. (4) GPCR43 knockdown using RNAi indicated that GPCR43 is not exclusively required for BuA to regulate apoptosis and proliferation. The results from this work indicate that GPCR43 is not likely to exclusively mediate BuA’s effects, but opens up new areas of research into the exact role of GPCR43 on CRC cells. / Thesis (Ph.D.) - University of Adelaide, School of Molecular and Biomedical Science, 2008

Thesis (Ph.D.)--Indiana University, 2009. / Title from screen (viewed on August 28, 2009). Department of Biochemistry and Molecular Biology, Indiana University-Purdue University Indianapolis (IUPUI). Advisor(s): William Bosron. Includes vita. Includes bibliographical references (leaves 102-111).

Σκοπός της παρούσας εργασίας ήταν η μελέτη της έκφρασης των γονιδίων NFY-C και RORA σε αδενοκαρκινώματα παχέος εντέρου και η σχέση της έκφρασης του NFY-C με την κατάσταση του γονιδίου p53. Η NFY-C πρωτεΐνη είναι μια από τις τρεις υπομονάδες του μεταγραφικού παράγοντα NFY, ο οποίος προσδένεται στους υποκινητές πολλών ευκαρυωτικών γονιδίων, περιλαμβανομένων γονιδίων που σχετίζονται με τον κυτταρικό κύκλο, επάγοντας τη μεταγραφή τους. Η ογκοκατασταλτική πρωτεΐνη p53 συνδέεται με τον NFY για τη μεταγραφή γονιδίων. Η πρωτεΐνη RORA είναι ένας υποδοχέας στεροειδών ορμονών, ο οποίος εμπλέκεται σε μια σειρά κυτταρικών διαδικασιών. Στην παρούσα μελέτη προσδιορίστηκαν τα επίπεδα mRNA του NFY-C και του RORA με ποσοτική RT-PCR σε 81 δείγματα καρκινικού ιστού παχέος εντέρου και 53 δείγματα φυσιολογικού ιστού παχέος εντέρου από ασθενείς με αδενοκαρκίνωμα παχέος εντέρου και διερευνήθηκε η σχέση τους με τα κλινικοπαθολογικά χαρακτηριστικά των ασθενών. Η πρωτεϊνική έκφραση του NFY-C προσδιορίστηκε με ανοσοϊστοχημεία σε 92 δείγματα νεοπλασματικού ιστού και 36 δείγματα φυσιολογικού ιστού παχέος εντέρου. Η κατάσταση των εξωνίων 5, 6, 7 και 8 του γονιδίου p53 προσδιορίστηκε με τη μέθοδο PCR-SSCP. Τα επίπεδα mRNA και πρωτεΐνης του NFY-C ήταν αυξημένα στους καρκινικούς ιστούς σε σχέση με τους φυσιολογικούς και διέφεραν ανάμεσα σε όγκους διαφορετικής εντόπισης. Τα υψηλά επίπεδα mRNA του NFY-C σχετίστηκαν με μεγαλύτερο ελεύθερο νόσου διάστημα (TTP) σε ασθενείς σταδίου Β. Η NFY-C πρωτεΐνη εντοπίστηκε μόνο στο κυτταρόπλασμα του κυττάρου και όχι στον πυρήνα και η έκφρασή της διέφερε ανάμεσα σε όγκους διαφορετικού σταδίου. Τα επίπεδα mRNA του RORA ήταν μειωμένα στους καρκινικούς ιστούς σε σχέση με τους φυσιολογικούς και διέφεραν ανάμεσα σε όγκους διαφορετικής εντόπισης. Η πρωτεϊνική έκφραση του NFY-C σχετίζεται με την κατάσταση του p53 ως προς την επιβίωση των ασθενών και το TTP. Συμπερασματικά, οι διαφορές έκφρασης των μορίων NFY-C και RORA υποδηλώνουν έναν πιθανό ρόλο στην καρκινογένεση του παχέος εντέρου. / The aim of the current study was to evaluate NFY-C and RORA expression and investigate the correlation of NFY-C expression with p53 status in patients with colorectal carcinoma. NFY-C protein is one of the three subunits of nuclear factor Y, which binds to the promoters of many eucaryotic genes, including cell cycle – related genes, inducing their transcription. p53 tumor supressor protein binds to NFY and induce gene expression. RORA protein is a steroid hormone receptor, which is implicated in many cellular processes. In the current study NFY-C and RORA mRNA levels were evaluated by quantitative RT-PCR in 81 neoplastic colorectal tissue specimens and 53 normal tissue specimens from patients with colorectal adenocarcinoma and were analysed in relation to clinicopathological parameters of the patients. Protein expression of NFY-C was assessed by immunochemistry in 92 malignant and 36 normal samples from patients with colorectal adenocarcinoma. p53 status of the exons 5, 6, 7 and 8 was detected using PCR-SSCP method. NFY-C mRNA and protein levels were elevated in malignant tissues compared to normal tissues and were related to the primary site of the tumor. Elevated mRNA levels of NFYC of stage B patients were significantly correlated with time to disease progression. NFY-C protein was detected only in the cytoplasm and not in the nucleus and its expression was correlated with the stage of the disease. RORA mRNA levels were decreased in normal tissues compared to malignant tissues and were related to the primary site. Protein expression of NFY-C in combination with p53 status is associated with overall survival and time to disease progression of the patients. In conclusion, the differential expression of NFY-C and RORA indicates a possible role for these molecules in colon carcinogenesis.

The Human Protein Atlas project is a Swedish research initiative that uses antibody-based proteomics for large scale protein profiling in human tissues and cells. Affinity-purified antibodies are produced within the project and used for immunohistochemical staining on tissue micro arrays (TMAs) in order to map the human proteome and publish the result in a protein atlas (www.proteinatlas.org). In this thesis, TMAs were used for analysis of protein expression patterns in order to identify and explore potential biomarkers of clinical relevance. In Paper I, protein expression of SATB2 was studied in colorectal cancer. The results show that SATB2 is a sensitive and specific biomarker for colorectal cancer, staining 85% of all investigated tumors. Moreover, SATB2 in combination with CK20 showed positivity in 97% of all colorectal carcinomas and is therefore suitable as a complementary tool in clinical differential diagnostics of cancer. In Paper II, ANLN was explored as a prognostic biomarker for breast cancer. A high nuclear fraction of ANLN in breast cancer was significantly correlated to large tumor size, high histological grade, hormone receptor negative tumors, high proliferation rate and poor prognosis. Furthermore, ANLN depletion in breast cancer cell lines resulted in cell cycle arrest and cellular senescence with altered cell morphology. In Paper III, young age at breast cancer diagnosis was investigated as an independent risk factor for poor prognosis. TMAs were produced from a selection of patients from a previously defined register-based cohort. The analysis shows that young women with luminal B tumors have a 2.2-fold higher risk of dying of breast cancer compared to older women. In Paper IV, vascular expression of CD93 was explored by image analysis of the tissue-based breast cancer cohort produced in Paper III. The analysis shows that young women with breast cancer display a significantly higher CD93-positive vessel area in their tumors. High CD93-positive vessel area was significantly associated with hormone receptor negative tumors, grade, Ki-67, EGFR and a poor prognosis. In conclusion, this thesis shows that protein expression profiling using TMAs is an important tool for identifying and exploring potential novel biomarkers for cancer.

The intestinal epithelium represents one of the most actively renewing tissues in the body, and is widely used as a model system to study epithelial cell biology. ASPP2, a member of the ASPP (apoptosis stimulating protein of p53) protein family, has been shown to act as a regulator of epithelial cell polarity and tumour suppressor. This study investigated whether the dual function of ASPP2 is involved in the regulation of intestinal homeostasis and tumourigenesis, with a particular interest in the distinction between epithelial cell autonomous and non-autonomous mechanisms. Germline and intestinal epithelial cell-specific ASPP2 conditional knockout mice were employed in this study. Deficiency of ASPP2 in the intestinal epithelium resulted in delayed recovery from dextran sulfate sodium (DSS)-induced acute colitis, concurrent with a reduction in the expression of proinflammatory cytokines such as interleukin (IL)-1β and IL-6. Moreover, ASPP2-deficient mice showed increased susceptibility to Azoxymethane/DSS-induced colorectal tumourigenesis. While wild-type and ASPP2-deficient crypts showed similar incidence of tumour formation, the local immune microenvironment of ASPP2-deficient mice favoured tumour progression. The intestinal organoid culture was established to supplement in vivo experiments. The feasibility of the system was demonstrated with small intestinal organoids, in the context of proliferation, differentiation, and cell death. Using the established workflow, a colonic organoid-based tissue regeneration model was developed. The intrinsic susceptibility of organoids to DSS-induced cell death was not affected by the loss of ASPP2. However, ASPP2-deficient colonic organoids were less responsive to the pro-proliferative effects of IL-6, but were more sensitive to tumour necrosis factor-α-induced cell death in the presence of IL-22. In conclusion, this project undertook parallel examinations of animal models and organoids, demonstrating that a deficiency of ASPP2 in the intestinal epithelium results in dysregulated epithelial-immune cell interactions. This may partially explain the pathological conditions observed in ASPP2-deficient mice. Importantly, this study highlights the possibility of using organoids to investigate epithelial cell non-autonomous factors implicated in intestinal pathogenesis.

Avec 42 000 nouveaux cas diagnostiqués en 2012, le cancer colorectal (CCR) représente en France le troisième cancer en termes d’incidence. Les métastases, qui surviennent principalement au niveau du foie et des poumons, en constituent la principale cause de décès. Malgré les progrès récents de la chimiothérapie et des agents ciblés, le taux de survie à 5 ans des patients présentant un CCR métastasé reste faible. Aujourd’hui, la résection chirurgicale est le seul traitement curatif, cependant moins de 20% des patients porteurs de métastases sont opérables. Il existe donc un grand nombre de patients présentant un CCR métastasé pour lequel aucun traitement curatif ne peut être proposé. La formation des métastases à partir d’une tumeur primaire résulte d’une longue série d’étapes séquentielles liées les unes aux autres. L’issue de ce processus dépend à la fois des propriétés intrinsèques des cellules tumorales et de la réponse de l’hôte. Il a récemment été montré que les couples chimiokines/récepteurs interviennent dans le contrôle des différentes étapes de la progression tumorale.Le projet de recherche développé au cours de mon travail de thèse avait pour objectif d’utiliser les chimiokines et leurs récepteurs dans de nouvelles stratégies thérapeutiques pour bloquer et/ou éradiquer les métastases hépatiques et pulmonaires des CCRs. Le travail s’est articulé selon deux axes dans lesquels nous avons montré d’une part que, le blocage du récepteur de chimiokines CXCR7 permet de limiter les métastases pulmonaires de CCRs et d’autre part que, le transfert de gène codant CX3CL1 au niveau du foie entraîne une réponse anti-tumorale efficace dans les métastases hépatiques de CCRs. / With 42 000 newly-diagnosed patients in 2012, the colorectal cancer (CRC) represents the third type of cancer in terms of incidence in France. The leading cause of death from CRC is the development of metastases and these metastases will occur mostly within the liver (50% of the patients) and within the lungs (15%). Despite recent progress, notably in the chemotherapies now used and the targeted agents, the rate of 5-years survival for late stage CRC remains low. Nowadays, the surgical resection is the only curative treatment proposed to patients with metastatic CRC, however less than 20% of them have an operable tumour. There is therefore a high number of patients for whom no cure is currently available. A primary tumour’s dissemination to a second organ is the result of a long process made of numerous cross-linked steps. The final outcome of this process depends on the intrinsic properties of tumour cells as well as the host response. Recently, it has been shown that the chemokine-chemokine receptor pairs (initially described as regulating the leukocyte migration) play crucial roles in the various stages involved in tumour progression. The aim of the research project developed during my PhD was to assess the use of the chemokines and their receptors in new therapeutic strategies to block and/or eradicate the hepatic and pulmonary metastases of CRC. Our work has been organized along two main lines of approach. We have shown that the blockage of the CXCR7 chemokine receptor enables the limitation of the CRC metastases within the lungs and that the CX3CL1 gene transfer into the hepatocytes leads to an efficient anti-tumor response in the CRC metastases within the liver.

Colorectal cancer (CRC), which includes cancer of the large bowel and rectum is the third most common cancer in men and the second in women and there is a poorer survival rate in less developed regions of the world such as West Africa mainly due to the ‘out of reach’ costs of chemotherapy. Evidence suggests that aspirin, a non-steroidal anti-inflammatory drug (NSAID) has the potential to decrease incidence of, or mortality from, a number of cancers including CRC through several mechanisms of action. However, this evidence is dampened by aspirin’s gastrointestinal (GI) toxicity, which have been found to be mostly age-dependent. The search for potential aspirin-related compounds with the same or better cytotoxic effects against cancer cells accompanied by a safer toxicity profile has been ongoing over the years and led to us to synthesise a number of novel aspirin analogues. One of the mechanisms of action suggested for the anticancer property of aspirin is the COX-dependent pathway. In this thesis SW480 cell line, a CRC cell line that is COX-2 negative and mismatch repair (MMR) proficient was used to study the possible COX-independent mechanism of action for aspirin, its analogues and diflunisal at 0.5 mM. Diflunisal was included in this study because it is also a salicylate with reports of having cytotoxic effects. OE33 and FLO1 oesophageal cancer cells were also employed in the epidermal growth factor receptor (EGFR) and synergy experiments to show effects were not just specific to SW480 cells alone. These aspirin analogues were synthesised, identified using nuclear magnetic resonance (NMR) and infra-red (IR) spectroscopy, and tested for purity using thin layer chromatography (TLC) and melting point. The findings of this study suggest that these compounds breakdown into salicylates and perturb epidermal growth factor (EGF) internalization with PN517 (fumaryldiaspirin) and PN590 (ortho-thioaspirin) also driving EGF co-localization with early-endosome antigen-1 (EEA1). The perturbation of the internalization of EGF by aspirin and PN517 was also observed by a time-lapse assay using live confocal imaging. These compounds also had specific effects on different tyrosine phosphorylation sites of the EGFR, with none but PN590 inhibiting 4 phosphorylation at Y1068, and all but PN502 (ortho-aspirin), PN548 (meta-aspirin) and PN549 (para-aspirin) inhibiting phosphorylation at Y1045 and Y1173. Given that the EGF internalization assay involved the cells being treated with compounds for 2 h, cells were also treated for this same time period and probed with pEGFR 1045, which resulted in the compounds having no significant effect on phosphorylation at that site which is responsible for the ubiquitination of the EGFR. Most of these compounds were apoptotic with some showing a combination of apoptosis and necrosis. Aspirin and its isomers drove apoptotic cell death in SW480 cells via the BCL2-BAX pathway while the thioaspirins appear to follow the p21 pathway by decreasing the expression of the protein. In addition, it was shown that PN502 (aspirin), PN517 and PN590 had synergistic effects when used in combination with oxaliplatin at ED50, ED75 and ED90 in SW480 CRC cells. The cytotoxicity of these compounds individually or in combination was determined using MTT assay followed by the use of the CompuSyn and CalcuSyn software to calculate combination index (CI), which indicated whether a drug combination was synergistic, antagonistic or additive. PN517 and PN524 were synergistic when used in combination with cisplatin in OE33 oesophageal cancer cells. Effect of these compounds on the EGFR indicates a delay or disruption of the signalling pathway involved in the proliferation of cancer cells, thus, translating into protection against tumour formation or progression while the synergistic effects of these compounds when used in combination with platinum compounds can provide patients with less toxic chemotherapeutic regimen especially in patients with CRC tumours that harbour mutant TP53 gene and normally resistant to oxaliplatin. It is therefore proposed that the perturbation of EGF internalization is a novel mechanism of action for aspirin and its analogues in cancer therapy. These positive findings shed light on the understanding of the possible mechanism of action for aspirins and gives hope for a more affordable, less toxic therapy for the prevention, treatment and management of cancer.

Introdução: A expectativa global para 2030 é uma incidência de 26,4 milhões de casos e 17 milhões de mortes causadas pelo câncer. O câncer colorretal (CCR) já é terceiro tumor mais incidente no mundo. No Brasil, o CCR é o terceiro tumor mais incidente em homens e o segundo em mulheres na região sul. Neste cenário pessimista, é importante avaliar a relação entre o custo e o benefício de tecnologias para o tratamento quimioterápico, sobretudo na gestão dos gastos na saúde pública. Objetivo: O presente trabalho objetivou avaliar as alternativas para o CCR, XELOX e mFOLFOX6, sob o aspecto econômico. Metodologia: As informações sobre a efetividade foram obtidas através de uma revisão narrativa da literatura. Realizou-se também uma revisão narrativa de estudos farmacoeconômicos e por fim uma análise de custo minimização (ACM) sob a perspectiva de um hospital de caráter público. O microcusteio foi utilizado como método para estimar o custo de cada componente que incluiu medicamentos, materiais, exames laboratorial e de imagem, atendimentos ambulatoriais, diárias de internação e recursos humanos e administrativos, permitindo identificar o custo individual por paciente, com cada alternativa. O sistema informatizado do hospital foi utilizado para a coleta dos dados. Resultados: Foram encontrados 14 estudos farmacoeconômicos, dentre os quais apenas 2 estudos nacionais, ambos em cenário metastático da doença. A ACM revelou um custo por paciente de R$ 9.925,98 (adjuvância) e R$ 8.036,95 (paliativo) para mFOLFOX6, e R$ 8.407,13 (adjuvância) e R$ 6.946,47 (paliativo) para tratamento com XELOX. Os custos de materiais e medicamentos representam cerca de 85% do custo total de XELOX; para mFOLFOX6 esse custo é menor, em torno de 36%. Por outro lado, os custos com internação e colocação de cateter ocorrem exclusivamente para mFOLFOX6, que também apresenta maior custo com recursos humanos. Conclusão: O número de pacientes e a falta de dados no sistema informatizado da instituição reforçam a necessidade de mais estudos para se afirmar que XELOX é menos oneroso que mFOLFOX6 no sistema público. O estudo é inédito por se tratar de uma ACM utilizando o método de microcusteio para comparar as alternativas em um hospital público e universitário do país, especialmente na adjuvância do CCR. / Introduction: The global expectation for 2030 is an incidence of 26.4 million cases and 17 million deaths caused by cancer. Colorectal cancer (CRC) is already the third most incident tumor in the world. In Brazil, CRC is the third most incident tumor in men and the second in women in the southern region. In this pessimistic scenario, it is important to evaluate the relationship between the cost and the benefit of technologies for chemotherapy treatment, especially in the management of public health expenditures. Objective: The present study aimed to evaluate the alternatives for CRC, XELOX and mFOLFOX6, under the economic aspect. Methodology: Information on effectiveness was obtained through a narrative review of the literature. A narrative review of pharmacoeconomic studies was also conducted, and finally, a cost minimization analysis (CMA) from a public hospital perspective was carried out. The micro-costing was used as a method to estimate the cost of each component that included medicines, materials, laboratory and imaging exams, outpatient visits, hospital stay and human and administrative resources, allowing the individual cost per patient to be identified for each alternative. The computerized system of the hospital was used to collect the data. Results: We found 14 pharmacoeconomic studies, of which only 2 national studies, both in the metastatic setting of the disease. The CMA revealed a cost per patient of BRL$ 9,925.98 (adjuvant) and BRL$ 8,036.95 (palliative) for mFOLFOX6, and BRL$ 8,407.13 (adjuvant) and BRL$ 6,946.47 (palliative) for treatment with XELOX. Material and drug costs account for about 85% of the total cost of XELOX; For mFOLFOX6 this cost is lower, around 36%. On the other hand, costs with hospitalization and catheter placement occur exclusively for mFOLFOX6, which also presents higher cost with human resources. Conclusion: The number of patients and the lack of data in the computerized system of the institution reinforce the need for further studies to assert that XELOX is less costly than mFOLFOX6 in the public system. The study is notorious because it is a CMA using the micro-costing method to compare the alternatives in a public and university hospital in the country, especially in the adjuvant treatment of CRC.

Colorectal cancer is one of the most common type of malignity. Despite of the existence of numerous studies focused on this carcinoma, there are still many unknown features regarding its diagnosis, treatment or prognosis. In the thesis we focused on the identification of novel prognostics markers that could be useful for the stratification of patients based on the disease outcomes. In the first study we immunohistochemically assessed expression of two proteins associated with cancer stem cells in the samples of primary colorectal cancer and matched liver metastasis. Goal of the study was to evaluate relation among expression of CD44 and CD133 and overall survival and disease free interval in our set of patients. We observed that increased ratio of CD133 positive compared to CD133 negative tumor glands resulted in longer disease free interval, finding which is opposite to the general view on the CD133 role in the cancer development. Our hypothesis is that we analyzed confined group of patients and followed a bit different goal, where we measured ratio between positive and negative glands in the view-field and not the intensity of staining as the previous studies did. Our second study was focused on the transcriptional analysis of the selected set of twelve genes using frozen samples from colorectal...

Le cancer colorectal métastatique est l’une des premières causes de décès par cancer dans les pays occidentaux, malgré le développement récent de nouveaux traitements ciblés. L’amélioration de la survie des patients passe par une meilleure compréhension des mécanismes moléculaires impliqués dans la progression tumorale et la formation des métastases. Compte tenu de l’importance du rôle des axes mTOR/HIF1α et CXCL12/CXCR4/CXCR7 dans le processus métastatique, nos objectifs ont été : i) d’analyser de façon extensive le statut de CXCL12 dans une collection de polypes et de tumeurs coliques de tous stades et phénotypes en comparaison avec la muqueuse saine, puis de comprendre les mécanismes régulant l’expression de la chimiokine dans les cellules tumorales ; ii) d’étudier in vitro le rôle de l’hypoxie dans la régulation de la signalisation induite par CXCL12 via CXCR4 et CXCR7 et l’intérêt d’une inhibition des axes mTOR/HIF-1α et CXCL12/CXCR4/CXCR7 en particulier sur les capacités migratoires des cellules tumorales.Nous avons montré que l’extinction du gène CXCL12 est un évènement précoce et systématique au cours de la cancérogenèse colique et que cette perte d’expression pourrait être régulée par un mécanisme d’acétylation au niveau des histones. Une expression différentielle de CXCR4 et CXCR7 au sein des tumeurs du colon a été mise en évidence. L’augmentation d’expression de CXCR7 dans les métastases par rapport aux stades précoces montre l’importance de cet axe dans le processus métastatique. Le maintien de l’expression à la surface cellulaire de CXCR4 en normoxie pendant au moins 24h après un bref passage en hypoxie n’avait pas encore été décrit. Ceci pourrait expliquer le « homing » des cellules tumorales circulantes dans les niches métastatiques selon un gradient de CXCL12. L’utilisation combinée d’irinotécan et de chalcone permettant d’inhiber la migration des cellules tumorales est une approche originale in vitro. Enfin, nous avons initié le développement de modèles métastatiques de cancer du colon par greffe orthotopique sur le caecum de souris NUDE dans l’objectif de tester de nouvelles approches thérapeutiques ciblées in vivo. / Despite the recent development of new targeted chemotherapies, metastatic colorectal cancer is still one of the leading causes of cancer related deaths in western countries. A better understanding of metastatic process would improve survival. Since the role of mTOR/HIF1α and CXCL12/CXCR4/CXCR7 axes in metastasis formation, our objectives were: i) to analyze extensively CXCL12 status in a collection of polyps and colon tumors whatever stages and phenotypes; ii) to study the role of hypoxia in CXCL12/CXCR4/CXCR7 signaling pathway in vitro and on tumor cells migration. We have shown that the CXCL12 extinction is a systematic early event during colorectal carcinogenesis. CXCL12 loss expression may be regulated by histone acetylation mechanism. There is a differential CXCR4 and CXCR7 expression in colon tumors. Increased CXCR7 expression in metastasis compared to early stages underlines the importance of this axis in metastatic process. We have shown for the first time that CXCR4 expression remained stabilized at the cell membrane 24 hours after a transient passage in hypoxia. It could explain circulating cells are attracted in metastatic niches under CXCL12 gradient. Drug combinations with chalcone and irinotecan are an original approach for inhibiting cell migration in vitro. Finally, we have initiated the development of a metastatic model of colon cancer with orthotopic colon human tumors xenograft in NUDE mice to test new therapeutic approaches in vivo.