La cornée est le tissu le plus densément innervé du corps humain. Cette innervation joue un rôle dans la régulation de la sécrétion du film lacrymal et exerce un rôle trophique direct sur l'épithélium cornéen. Les axones cornéens expriment différents types de récepteurs et répondent à des fonctions de mécano-nocicepteurs, de récepteurs au froid ou de récepteurs polymodaux. Nous avons pu identifier de nouvelles lignées de souris transgéniques et les utiliser pour caractériser ces différentes populations axonales. L'innervation cornéenne débute à E12.5 chez la souris et est régulée par les molécules de Slits et de Sémaphorines et leurs récepteurs Robo et Plexines/Neuropilines respectivement. Nous avons pu étudier le rôle de ces deux familles dans le développement de l'innervation cornéenne. Les mutants Slits et Robos présentent une réduction du nombre et de la taille des terminaisons axonales épithéliales cornéennes. A l'âge adulte, les mutants Robos présentent une dégénérescence précoce des terminaisons épithéliales. Les mutants plexine-A4 et Neuropiline-1 présentent à l'inverse une augmentation du nombre de divisions des troncs stromaux cornéens. A l'âge adulte les mutants Plexine-A4 retrouvent une organisation classique de l'innervation alors que les mutants Neuropiline-1 conservent la désorganisation de l'innervation cornéenne. Nous avons également étudié la régénération de l'innervation après lésions de grattage de l'épithélium cornéen. Nos résultats préliminaires semblent en faveur d'une augmentation de la régénération de l'innervation cornéenne chez les mutants Neuropiline-1. / The cornea is the most densely innervated tissue in the entire body. Corneal innervation plays a role in regulating the secretion of lacrimal film and exerts a direct trophic role on the corneal epithelium. Corneal axons express different types of sensory receptors ranging between mechano-, thermo-, and polymodal nociceptors. We identified transgenic mouse lines to characterize these different axonal populations. Corneal innervation begins at E12.5 in mice and is regulated by a range of axon guidance cues such as Slits and Semaphorins, which respond to their receptors Robo and Plexin/Neuropillin respectively. We studied the role of these two families in the development of corneal innervation. The Slits and Robos mutants show a reduction in the number and size of the corneal epithelial nerves endings. In adult, Robos mutants exhibit early degeneration of the epithelial nerves endings. Plexin-A4 and Neuropilin-1 mutants, on the other hand, show an increase in the number of divisions of the corneal stromal nerve trunks. In adult, Plexin-A4 mutants regain a classical organization of innervation whereas Neuropilin-1 mutants retain the disorganization of corneal innervation. Following a lesion, corneal innervation is able to regenerate, however the axons never regain their initial morphology or complexity. Due to the increased corneal innervation observed in the Neuropilin-1 mutants, we wondered whether the regeneration of innervation after scrapping lesions of the corneal epithelium could be enhanced in Neuropilin-1 loss of function. Our preliminary results support an increase in corneal innervation regeneration in Neuropilin-1 mutants.

The role of vascular endothelial growth factor (VEGF) signaling in cancer is well-known in the context of angiogenesis but is also important in the functional regulation of tumor cells themselves. Notably, autocrine VEGF signaling mediated by its co-receptors called neuropilins (NRPs) appears be essential for sustaining the proliferation and survival of cancer stem cells (CSCs), which are implicated in mediating tumor growth, progression and drug resistance. Therefore, the first half of this thesis focuses on the mechanism of VEGF-NRP-mediated support of CSCs. Aberrant activity of the Hippo pathway effector YAP and TAZ are associated with breast CSCs and have been shown to confer stem cell-like properties. I found that VEGF-NRP2 signaling contributes to the activation of YAP/TAZ in various breast cancer cells, which mediates a positive feedback loop that promotes mammosphere formation. VEGF-NRP2 signaling activated the GTPase Rac1, which inhibited the Hippo kinase LATS, which enabled the activity of YAP/TAZ. In complex with the transcription factor TEAD, TAZ then bound and repressed the promoter of the gene encoding the Rac GTPase-activating protein (Rac GAP) β2-chimaerin. By activating GTP hydrolysis, Rac GAPs effectively turn off Rac signaling; hence, YAP/TAZ-mediated repression of β2-chimaerin sustained Rac1 activity in CSCs. Depletion of β2-chimaerin in non-CSCs increased Rac1 activity, YAP/TAZ activation and mammosphere formation. Analysis of breast cancer patients revealed an inverse correlation between β2-chimaerin and TAZ expression in tumors. These findings highlight an unexpected role for β2-chimaerin in a feedforward loop of YAP/TAZ activation and the acquisition of CSC properties. Given that CSCs have been implicated in therapy resistance and are enriched in triple negative breast cancer (TNBC), which exhibits VEGF-NRP2 signaling, the second half of this thesis focuses on understanding the mechanism by which VEGF-NRP2 contributes to the chemoresistance of TNBC. I discovered that VEGF-NRP2 promote homologous recombination (HR) in BRCA1 wild-type TNBC cells by contributing to the expression and function of Rad51, an essential enzyme in the HR pathway that mediates efficient DNA double strand break repair. Mechanistically, I found that VEGF-NRP2 stimulates YAP/TAZ-dependent Rad51 expression and that Rad51 is a direct YAP/TAZ-TEAD transcriptional target. I also discovered that VEGF-NRP2-YAP/TAZ signaling contributes to the resistance of TNBC cells to chemotherapy and that Rad51 rescues the defects in DNA repair upon inhibition of either VEGF-NRP2 or YAP/TAZ in response to cisplatin. These findings reveal novel roles for VEGF-NRP2 and YAP/TAZ in DNA repair and they indicate a unified mechanism involving VEGF-NRP2, YAP/TAZ and Rad51 that contributes to resistance to platinum chemotherapy. In summary, this thesis provides novel insight into the roles of autocrine VEGF-NRP2 signaling in breast CSC function and therapy resistance and provides rationale in inhibiting NRP2 for platinum-resistant tumors that are dependent on YAP/TAZ activation.

De récents progrès dans la compréhension de la régulation de l'interaction moléculaire entre les lymphocytes T CD8 (LT CD8) et les cellules tumorales ont donné lieu à de nouvelles immunothérapies contre le cancer. En effet, les anticorps monoclonaux anti-CTLA-4 et anti-PD-1 (mAb), ciblant des récepteurs inhibiteurs à la surface des LT CD8, ont démontré des bénéfices de survie dans de nombreux cancers. Malheureusement, seule une partie des patients répond à ces traitements. Il est donc crucial de caractériser l'influence d'autres facteurs moléculaires régulant la migration et les activités fonctionnelles des LT CD8 afin d'améliorer les traitements actuels du cancer. Un candidat potentiel impliqué dans cette immunosuppression est le récepteur Neuropiline-1 (Nrp-1), qui a été initialement identifié dans le système nerveux. Nous avons identifié une large population de LT CD8 infiltrant le mélanome murin (lignée tumorale B16 greffée à des souris C57BL/6), qui exprime des niveaux élevés de Nrp-1. La présence de cette population augmente pendant la croissance tumorale, mais reste indétectable dans les organes lymphoïdes (rate et ganglion lymphatique drainant la tumeur). De plus, les LT CD8 infiltrant la tumeur exprimant Nrp-1 montre des caractéristiques d’épuisement. En parallèle, l’inhibition de la Nrp-1 avec un anticorps neutralisant augmente la capacité des LT CD8 à migrer, et à tuer les cellules tumorales autologues ex vivo. D’autre part, l’inhibition de la Nrp-1 in vivo agit en synergie avec la neutralisation du récepteur PD-1 pour le contrôle de la croissance tumorale et l’amélioration des fonctions effectrices des LT CD8. Nos résultats montrent donc comment le microenvironnement tumoral induit l'expression de la Nrp-1 sur les LT CD8 et comment ce récepteur neuronal peut participer à la régulation des fonctions effectrices des LT CD8 antitumoraux, avec de potentielles implications comme biomarqueur/cible pour une immunothérapie. / Recent advances in understanding the regulation of molecular interaction between CD8 T cell and tumor cells gave rise to novel cancer immunotherapies. Indeed, monoclonal antibodies against CTLA-4 and PD-1 (mAb), targeting inhibitory receptors on the surface of CD8 T cells, have demonstrated survival benefits in many cancers. Unfortunately, only a fraction of patients responds to these treatments. Therefore, it is crucial to characterize the influence of additional molecular factors regulating the migration and the functional activities of CD8 T cells, in order to improve current cancer treatments. A potential candidate involved in this immunosuppression function is the Neuropilin-1 (Nrp-1), which was originally identified in the nervous system. We identified a large subset of CD8 T cells infiltrating B16 melanoma (mouse model), which expresses high levels of Nrp-1. The frequency of Nrp-1+ CD8 T cells increased during tumor growth, but was undetectable in lymphoid organs (spleen and tumor-draining lymph node). Moreover, Nrp-1+ CD8 tumor-infiltrating lymphocytes (TIL) displayed features of T cell exhaustion. Importantly, blocking of Nrp-1 with a neutralizing mAb, increases the capacity of CD8 T cells to migrate and to kill autologous tumor cells. In addition, the blockade of Nrp-1 in vivo acts synergistically with the neutralization of the inhibitory receptor PD-1 to control tumor growth and restore TIL function. Our findings show how the tumor microenvironment induces Nrp-1 expression on CD8 T cells and how this neuronal receptor may participate in regulating antitumor CD8 T cell response, with potential implications as a biomarker/target for immunotherapeutic intervention.

Edema or tissue swelling is exacerbated during inflammation due to increased leukocyte infiltration and vascular permeability, after which resolution returns the tissue to homeostasis. In acute inflammatory reactions, upregulated levels of vascular endothelial growth factor (VEGF) is shown to increase vascular permeability. Vascular endothelial cells (EC) form a selective barrier regulating the degree of microvascular exchange and permeability in normal physiological and pathological settings. Vascular EC express pro-permeability VEGF receptors and neuropilin co-receptors that can mediate both stimulatory and inhibitory signals. Secreted class 3 semaphorin-3F (SEMA3F) is a high affinity ligand for the NRP2 receptor and has been shown to be anti-angiogenic through its ability to inhibit cell migration and attachment. Importantly, SEMA3F has been shown to compete for binding with VEGF to the NRP2 receptor. However, the role, if any, of SEMA3F in inflammation has yet to be fully elucidated. We hypothesize that SEMA3F reduces edema by inhibiting vascular permeability thereby promoting a quickened resolution of inflammation. To generate inflammatory lesions, delayed-type hypersensitivity cutaneous reactions were induced on the ear skin of C57BL/6 mice through topical applications of oxazolone. Total ear thickness as a readout of tissue swelling was compared to baseline (Day 0). To determine the effects of depleting SEMA3F during inflammation, ear thickness was measured after SEMA3F antibody or control IgG intraperitoneal injection into Nrp2+/- mice. To assess the effects of increased systemic SEMA3F on edema, ear thickness was measured after intravenous delivery of SEMA3F adenovirus (Ad-3F) or control adenovirus into wild-type mice. We report that SEMA3F depletion via SEMA3F antibodies led to significantly prolonged edema compared to controls. Ad-3F treated mice exhibit lower levels of inflammatory edema compared to control. We demonstrate that the SEMA3F signaling cascade is a key mediator of fluid homeostasis in inflammation. Likely, SEMA3F serves as an anti-inflammatory mechanism preventing excessive edema. / 2020-02-20T00:00:00Z

京都大学 / 0048 / 新制・課程博士 / 博士(医学) / 甲第20806号 / 医博第4306号 / 新制||医||1025(附属図書館) / 京都大学大学院医学研究科医学専攻 / (主査)教授 井上 治久, 教授 髙橋 良輔, 教授 江藤 浩之 / 学位規則第4条第1項該当 / Doctor of Medical Science / Kyoto University / DFAM

The neuropilin (Nrp) family consists of multifunctional cell surface receptors with critical roles in a number of different cell and tissue types. A core aspect of Nrp function is ligand-dependent cellular adhesion and migration, where it controls the multistep process of cellular motility through integration of ligand binding, receptor coupling and signaling via the coordinated action of its extracellular and intracellular domains. While Nrp regulates cellular adhesion and motility in the cardiovascular and nervous systems under physiological conditions, the emerging pathological role of Nrp in tumor cell migration and metastasis has been identified and provides motivation for continued efforts toward developing Nrp inhibitors. At the molecular level, the role of Nrp in adhesion and migration is intimately connected to the control of adhesive interactions and cytoskeletal reorganization. The adhesive “interactome” for Nrp draws much attention because of its lack of enzymatic activity and inability to transduce signals on its own. It is an active area of research and is still expanding dramatically. Nrp has been well defined as a co-receptor for vascular endothelial growth factor receptor (VEGFR)/vascular endothelial growth factor (VEGF) signaling through enhancing receptor-ligand interaction in angiogenesis. Here, we contribute to this concept through characterization in more biochemical detail about Nrp-1/VEGF physical interactions. VEGF has been shown to compete with Sema3 for binding to Nrp-1 b1 ligand binding pocket. This competition fine-tunes VEGF-induced angiogenesis. Our data provides a molecular mechanism for high affinity Sema3F binding to Nrp-1 in the b1 domain. As to the VEGFR-independent function, Nrp/integrin association has been demonstrated. The functional integration has been shown for Nrp/integrin in angiogenic sprouting. Both proteins are highly expressed in endothelial tip cells to mediate endothelial cell migration during angiogenesis and knockdown of either one in mice leads to embryonic lethality due to similar defects in vascular development. To identify the structure and function correlation, we characterized in more detail about Nrp-1/integrin physical interactions with biochemical and cell-based assays. Through an integrated approach of biochemical, molecular and cellular methods, we defined the direct physical interactions between Nrp-1 and integrins. We have also extended this work to demonstrate the functional importance and contribution of the interactions in integrin-mediated cell adhesion on extracellular matrix (ECM) in angiogenesis and platelet function during wound healing and provide a molecular basis for the integration of Nrps/integrins in cell migration, adhesion to ECM, breast cancer initiation and breast cancer stem cell fate determination.

Pour améliorer la sélectivité cellulaire tumorale et/ou l'efficacité anti-tumorale de la thérapie photodynamique (PDT), des molécules actives (peptides, pseudopeptides, agent alkylant) ont été conjuguées à la molécule photo-activable. Dans une 1ère partie, l'effet vasculaire de la PDT a été favorisé par le ciblage de la vascularisation tumorale de phénotype angiogénique. Un photosensibilisateur de type chlorine a été couplé à un peptide ligand de neuropiline-1 (NRP-1) et a montré son efficacité anti-tumorale in vitro et in vivo en PDT. Cependant, la partie peptidique s'est avérée instable in vivo. Des pseudopeptides modifiés au niveau du site de clivage du peptide initial ont été synthétisés et testés in vitro, conjugués ou non à la chlorine. L'incorporation cellulaire de la chlorine a été améliorée après son couplage aux pseudopeptides. Une technique d'ARN interférence, visant à éteindre l'expression de NRP-1 dans les cellules MDA-MB-231, surexprimant NRP-1, a mis en évidence l'incorporation cellulaire récepteur-dépendante des conjugués et leur sélectivité cellulaire. Les pseudopeptides et conjugués se sont révélés affins pour NRP-1 et plus stables in vivo. Le ruthénium (Ru) s'affiche comme une molécule d'intérêt thérapeutique en oncologie. Dans une 2ème partie, nous avons appréhendé l'intérêt in vivo de coupler cet agent alkylant à des photosensibilisateurs de type porphyrine et chlorine. Les études d'efficacité in vivo sur un modèle de souris nude xénogreffées en ectopique avec des cellules de carcinome oral humain KB, ont été menées par une méthodologie de plans d'expérience. Deux conjugués [Ru(_6-p-PriC6H4Me)(5-(3-pyridyl)-10,15,20-triphenylporphyrin)Cl2, ou Rut1, et Ru4(_6-p-PriC6H4Me)4(5,10,15,20-tetra(3-pyridyl)porphyrin)Cl8, ou Rut4] efficaces in vitro ont été comparés in vivo. Le composé Rut4 est apparu comme le plus efficace. Des études de biodistribution par spectrofluorimétrie fibrée et de pharmacocinétique classique ont montré un intervalle drogue-lumière optimal de 24h. Néanmoins, malgré l'optimisation des modalités thérapeutiques, le couplage du Ru avec une molécule photo-activable (porphyrine ou chlorine) n'a pas conduit à une efficacité anti-tumorale significative in vivo / To improve tumor cells selectivity and/or photodynamic therapy (PDT) anti-tumor efficiency, active molecules (peptides, pseudopeptides, alkylating agents) have been conjugated to the photo-activable molecule. In a first part, vascular effect of PDT has been promoted by targeting the tumor vasculature with angiogenic phenotype. A chlorine-type photosensitizer have been coupled to a neuropilin-1 (NRP-1)-targeting peptide and has shown its in vitro and in vivo anti-tumor efficiency for PDT. However, the peptidic sequence has been degraded in vivo. Clivage site-modified pseudopeptides have been synthesized and tested in vitro, conjugated to TPC or not. Chlorine cellular uptake has been improved after coupling with pseudopeptides. A technique of RNA interference-mediated silencing of NRP-1 in MDA-MB-231, over-expressing NRP-1, has highlighted the receptor-dependent uptake of the conjugates and their cellular selectivity. Pseudopeptides and conjugates have shown their NRP-1 affinity and their in vivo stability. Ruthenium (Ru) is considered as an interest therapeutic molecule in oncology. In a second part, we have studied the in vivo interest of coupling an alkylating agent with porphyrine and chlorine-type photosensitizers. The in vivo efficiency studies on nude mice xenografted ectopically with KB cells have been carried using experimental design approach. Two in vitro efficient conjugates [Ru(_6-p-PriC6H4Me)(5-(3-pyridyl)-10,15,20-triphenylporphyrin)Cl2, or Rut1, and Ru4(_6-p-PriC6H4Me)4(5,10,15,20-tetra(3-pyridyl)porphyrin)Cl8, or Rut4] have been compared in vivo. Rut4 conjugate has appeared to be more efficient than Rut1. Biodistribution studies using fiber spectrofluorimeter and classic pharmacokinetic studies have shown an optimal drug-light interval of 24h. However, after PDT parameters optimization, the Ru coupling with a photo-activable molecule (porphyrin or chlorine) hasn?t shown any significant in vivo anti-tumor efficiency

Le médulloblastome (MB) est la plus fréquente des tumeurs cérébrales malignes pédiatriques qui représentent la première cause de mortalité par cancer chez l’enfant. Malgré les avancées des nouveaux traitements, les risques de récidive, séquelles et décès après traitement restent importants. Le récepteur de neuropiline-1 (NRP-1) a été récemment impliqué dans la progression tumorale des MBs et semble jouer un rôle important dans le phénotype des cellules souches cancéreuses (CSCs). Le ciblage de cette molécule pourrait ainsi présenter un intérêt thérapeutique dans le traitement des MBs. Nous avons sélectionné des cellules souches de MB capables de former des médullosphères (MS) à partir de 3 lignées cellulaires (DAOY, D283-Med et D341-Med). Ces modèles ont été caractérisés par l’expression de neuropilines (NRP-1 and NRP-2) et de marqueurs phénotypiques (CD133，CD15 et NF-M). Les résultats ont montré une augmentation significative de l’expression de NRP-1 par les cellules cultivées en médullosphères confortant notre stratégie de ciblage. L’impact du traitement de ces cellules par un composé innovant ciblant spécifiquement NRP-1, le MR438, a été ensuite évalué in vitro seul et association avec la radiothérapie notamment sur l’étude de la capacité d’auto-renouvellement des CSCs de MBs. Nous avons mis en évidence une diminution de la capacité d’autorenouvellement des cellules souches de MBs après exposition au MR438 avec une radiosensibilité augmentée pour les 3 modèles cellulaires. In vivo, le composé MR438 a été evalué sur des modèles de xénogreffes hétérotopiques chez la souris nude et montre un effet radiopotentialisant significatif pour les tumeurs issues de la lignée Daoy avec une tendance à la diminution de la progression tumorale pour les 2 autres lignées. De façon intéressante, le composé MR438 induit une diminution significative du nombre de cellules souches pour l’ensemble de nos modèles. Par conséquent, le composé semblerait induire les cellules souches vers un phénotype différencié au moins pour la lignée DAOY, même si les mécanismes n’ont pas pu être clairement élucidé. En conclusion, l’inhibition de NRP-1 via MR438 semble stimuler la différenciation des cellules souches cancéreuses pouvant à terme réduire la progression du MB et apporter un bénéfice en association avec la radiothérapie. L’evaluation du composé sur des modèles orthotopiques de MB permettrait d’obtenir des informations quant à son efficacité sur des modèles plus proche de la physiopathologie tenant compte de sa distribution au niveau cérébral / Medulloblastoma (MB) is the most common malignant pediatric brain tumors which is the leading cause of cancer death in children. Despite the progress of new treatments, the risk of recurrence, morbidity, and death after treatment remain important. The neuropilin-1 receptor (NRP-1) has recently been implicated in tumor progression of MBs, which seems to play an important role in the phenotype of cancer stem cells (CSCs). Targeting this molecule could thus present an interesting therapeutic value in the treatment of MB. We have selected cancer stem like cells of MBs in the form of medullospheres (MSs) from 3 cell lines (DAOY, D283-Med and Med-D341). These models were characterized by expression of neuropilins (NRP-1 and NRP-2) and phenotypic markers (CD133, CD15 and NF-M). Results showed a significant increase of the expression of NRP-1 by our CSCs models cultured in MSs that confirms our targeting strategy. The impact of the treatment of these cells with an innovative compound specifically targeting NRP-1, MR438, was then evaluated in vitro alone and in association with radiotherapy, especially on the study of the capacity for self-renewal. A decrease of self-renewal capacity for MB stem cells after exposition of MR438 with an increase of radiosensitivity for the 3 cell models in vitro was demonstrated. In vivo, MR438 was evaluated on heterotopic xenograft models in nude mice and showed a significant augmentation of radiosensitivity for DAOY tumors with a tendency to decrease tumor progression for the other 2 cell lines. Interestingly, the compound MR438 induced a significant decrease in the number of stem cells for all of our models. The compound appeared to induce CSCs to a differentiated phenotype at least for the DAOY cells, although mechanisms could not be clearly elucidated. In conclusion, inhibition of NRP-1 via MR438 seems to stimulate the differentiation of CSCs that may eventually reduce the progression of MB and bring a benefit in association with radiotherapy. Evaluation of this compound on orthotopic models of MB would provide information on its effectiveness on models closer to the physiopathology taking into account its distribution at the cerebral level

Neuropilin (Nrp) is an essential cell surface receptor with dual functionality in the cardiovascular and nervous systems. The first identified Nrp-ligand family was the Semaphorin-3 (Sema3) family of axon repulsion molecules. Subsequently, Nrp was found to serve as a receptor for the vascular endothelial growth factor (VEGF) family of pro-angiogenic cytokines. In addition to its physiological role, VEGF signaling via Nrp directly contributes to cancer stemness, growth, and metastasis. Thus, the Nrp/VEGF signaling axis is a promising anti-cancer therapeutic target. Interestingly, it has recently been shown that Sema3 and VEGF are functionally opposed to one another, with Sema3 possessing potent endogenous anti-angiogenic activity and VEGF serving as an attractive cue for neuronal axons. We hypothesized that direct competition for an overlapping binding site within the Nrp extracellular domain may explain the observed functional competition between VEGF and Sema3. To test this hypothesis we have separately investigated the mechanisms of VEGF and Sema3 binding to Nrp. Utilizing structural biology coupled with biophysics and biochemistry we have identified both distinct and common mechanisms that facilitate the interaction between Nrp and these two ligand families. Specifically, we have identified an Nrp binding pocket to which these ligands competitively bind. The Sema3 family uniquely requires proteolytic activation in order to engage this overlapping binding site. These findings provide critical mechanistic insight into VEGF and Sema3 mediated physiology. Additionally, these data have informed the development of small molecules, peptides, and soluble receptor fragments that function as potent and selective inhibitors of VEGF/Nrp binding with exciting therapeutic potential for treating cancer.

Ces travaux de thèse portent sur la conception, la synthèse et l'étude biologique de vecteurs peptidiques pour des applications en diagnostic et/ou thérapie du cancer.Nous avons utilisé des châssis cyclodécapeptidiques fonctionnalisés de façon chimiosélective par des éléments de ciblage et des effecteurs. Ces châssis, dotés de quatre ligands c[RGDfK], possèdent une forte affinité pour les récepteurs intégrine avB3 qui sont surexprimés dans de nombreux cancers et par les cellules endothéliales de l'environnement tumoral. Ils sont en revanche peu exprimés par les tissus sain. La présentation multivalente des ligands -RGD- permet également au châssis d'être internalisé par les cellules tumorales.Nous avons donc mis au point des molécules composées du châssis peptidique, de quatre ligands -RGD- pour le ciblage tumoral, et de différents effecteurs pour plusieurs applications. A travers de multiples collaborations, nous avons relié ce vecteur à un agent hautement toxique (cryptophycine), un photosensibilisateur (DHP), un peptide pro-apoptotique (BAX), un complexe de Gallium 68 (pour une étude clinique de phase I pour une application en imagerie TEP). Nous avons également greffé ces châssis présentant quatre motifs -RGD- à des polymères ou à des nanoparticules de silice, tous deux fluorescents.Le projet principal de cette thèse était la conception de vecteurs ciblant deux récepteurs tumoraux de manière simultanée. En plus de cibler l'intégrine avB3, nous avons ciblé le récepteur NRP1 qui est lui aussi surexprimé lors de l'angiogenèse tumorale. Nous avons exploité divers réactions chimiosélectives (oxime, cycloaddition de Huisgen, amidation) pour concevoir des vecteurs fluorescents ciblant l'un des deux récepteurs ou les deux à la fois. Des tests biologiques in vitro et in vivo ont été réalisés. Il s'avère que les composés à double ciblage permettent une très bonne détection de la tumeur, mais non supérieure à des composés à mono ciblage. En revanche, la réponse cellulaire déclenchée par les composés à double ciblage est unique, et totalement différente d'une co-injection. Nous avons plusieurs éléments qui tendent à prouver qu'un complexe NRP1-vecteur-Intégrine se formerait et resterait ancré au niveau de la membrane cellulaire, bloquant son internalisation. / This thesis work is about conception, synthesis and biological activities of peptide vectors for diagnostic and/or therapeutic applications against cancer.We used cyclodecapeptidic scaffolds chemoselectively handled with targeting elements and effectors. Those scaffolds presenting four c[RGDfK] ligands have a strong affinity for integrin avB3 receptors, wich are overexpressed in various cancers and by endothelial cells from the tumor surrounding. They are poorly expressed in healthy tissues. The multivalent presentation of -RGD- motifs higly increases the internalisation of the scaffold by tumor cells. Thus we developed molecules composed by four -RGD- motifs for tumor targeting, and different effectors for various applications. Thanks to multiple collaborations, we linked the vector to a highly cytotoxic compound (cryptophycine), a photosensitiser (DHP), a pro-apoptotic peptide (BAX), a DOTA complex (for 68-Ga complexation, for PET applications). We also grafted the cyclodecapeptide bearing four -RGD- motifs to polymers or silica nanoparticles, both fluorescent.The main project of this thesis was the conception of dual targeting vectors. Our objective was to simultaneously target two receptors overexpressed in the tumor periphery. Beside the targeting of avB3, we decided to target the NRP1 receptor, which is also overexpressed during tumor angiogenesis. We exploited various chemoselective reactions (oxime, huisgen cycloaddition, peptide coupling) to synthesise fluorescent vectors targeting one of the two receptors, or both. In vitro and in vivo biological experiments were realised. We discovered that dual targeting compounds allow a really good tumor detection, but inferior to mono targeting ones. Nevertheless, the cellular answer triggered by dual targeting compounds is totally different from those obtained with other compounds, including co-injection. We found different elements that tend to show that a NRP1-vector-integrin could be formed, and would be blocked inside the cellular membrane, resulting in its internalisation's blocking.