Réponses immunitaires et cancer

Publications de l’équipe

Année de publication : 2009

Ana Ceballos, Federico Remes Lenicov, Juan Sabatté, Christian Rodríguez Rodrígues, Mercedes Cabrini, Carolina Jancic, Silvina Raiden, Mónica Donaldson, Rodolfo Agustín Pasqualini, Clara Marin-Briggiler, Mónica Vazquez-Levin, Francisco Capani, Sebastián Amigorena, Jorge Geffner (2009 Oct 28)

Spermatozoa capture HIV-1 through heparan sulfate and efficiently transmit the virus to dendritic cells.

The Journal of experimental medicine : 2717-33 : DOI : 10.1084/jem.20091579 En savoir plus
Résumé

Semen is the main vector for HIV-1 dissemination worldwide. It contains three major sources of infectious virus: free virions, infected leukocytes, and spermatozoa-associated virions. We focused on the interaction of HIV-1 with human spermatozoa and dendritic cells (DCs). We report that heparan sulfate is expressed in spermatozoa and plays an important role in the capture of HIV-1. Spermatozoa-attached virus is efficiently transmitted to DCs, macrophages, and T cells. Interaction of spermatozoa with DCs not only leads to the transmission of HIV-1 and the internalization of the spermatozoa but also results in the phenotypic maturation of DCs and the production of IL-10 but not IL-12p70. At low values of extracellular pH (approximately 6.5 pH units), similar to those found in the vaginal mucosa after sexual intercourse, the binding of HIV-1 to the spermatozoa and the consequent transmission of HIV-1 to DCs were strongly enhanced. Our observations support the notion that far from being a passive carrier, spermatozoa acting in concert with DCs might affect the early course of sexual transmission of HIV-1 infection.

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Laurence Bougnères, Julie Helft, Sangeeta Tiwari, Pablo Vargas, Benny Hung-Junn Chang, Lawrence Chan, Laura Campisi, Gregoire Lauvau, Stephanie Hugues, Pradeep Kumar, Alice O Kamphorst, Ana-Maria Lennon Dumenil, Michel Nussenzweig, John D MacMicking, Sebastian Amigorena, Pierre Guermonprez (2009 Aug 25)

A role for lipid bodies in the cross-presentation of phagocytosed antigens by MHC class I in dendritic cells.

Immunity : 232-44 : DOI : 10.1016/j.immuni.2009.06.022 En savoir plus
Résumé

Dendritic cells (DCs) have the striking ability to cross-present exogenous antigens in association with major histocompatibility complex (MHC) class I to CD8(+) T cells. However, the intracellular pathways underlying cross-presentation remain ill defined. Current models involve cytosolic proteolysis of antigens by the proteasome and peptide import into endoplasmic reticulum (ER) or phagosomal lumen by the transporters associated with antigen processing (TAP1 and TAP2). Here, we show that DCs expressed an ER-resident 47 kDa immune-related GTPase, Igtp (Irgm3). Igtp resides on ER and lipid body (LB) membranes where it binds the LB coat component ADFP. Inactivation of genes encoding for either Igtp or ADFP led to defects in LB formation in DCs and severely impaired cross-presentation of phagocytosed antigens to CD8(+) T cells but not antigen presentation to CD4(+) T cells. We thus define a new role for LB organelles in regulating cross-presentation of exogenous antigens to CD8(+) T lymphocytes in DCs.

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Matias Ostrowski, Nuno B Carmo, Sophie Krumeich, Isabelle Fanget, Graça Raposo, Ariel Savina, Catarina F Moita, Kristine Schauer, Alistair N Hume, Rui P Freitas, Bruno Goud, Philippe Benaroch, Nir Hacohen, Mitsunori Fukuda, Claire Desnos, Miguel C Seabra, François Darchen, Sebastian Amigorena, Luis F Moita, Clotilde Thery (2009 Aug 14)

Rab27a and Rab27b control different steps of the exosome secretion pathway.

Nature cell biology : 19-30; sup pp 1-13 : DOI : 10.1038/ncb2000 En savoir plus
Résumé

Exosomes are secreted membrane vesicles that share structural and biochemical characteristics with intraluminal vesicles of multivesicular endosomes (MVEs). Exosomes could be involved in intercellular communication and in the pathogenesis of infectious and degenerative diseases. The molecular mechanisms of exosome biogenesis and secretion are, however, poorly understood. Using an RNA interference (RNAi) screen, we identified five Rab GTPases that promote exosome secretion in HeLa cells. Among these, Rab27a and Rab27b were found to function in MVE docking at the plasma membrane. The size of MVEs was strongly increased by Rab27a silencing, whereas MVEs were redistributed towards the perinuclear region upon Rab27b silencing. Thus, the two Rab27 isoforms have different roles in the exosomal pathway. In addition, silencing two known Rab27 effectors, Slp4 (also known as SYTL4, synaptotagmin-like 4) and Slac2b (also known as EXPH5, exophilin 5), inhibited exosome secretion and phenocopied silencing of Rab27a and Rab27b, respectively. Our results therefore strengthen the link between MVEs and exosomes, and introduce ways of manipulating exosome secretion in vivo.

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Alexandre Boissonnas, Alix Scholer-Dahire, Luc Fetler, Sebastian Amigorena (2009 Jun 13)

Multiphoton imaging of cytotoxic T lymphocyte-mediated antitumor immune responses.

Current topics in microbiology and immunology : 265-87 : DOI : 10.1007/978-3-540-93864-4_11 En savoir plus
Résumé

The actual contribution of T lymphocytes to protection against tumors is still unclear. In vitro imaging experiments show that tumor specific cytotoxic T lymphocytes (CTLs) are competent to kill target cells by conventional cytotoxic pathways. The emergence of multiphoton imaging in the past decade now allows real time in vivo imaging of CTLs. New insights are available on the behavior of antitumor T cells during the priming phase, during their traffic within the tumor tissue, and on their interactions with tumor cells during the effector phase. Recent reports suggest that direct killing of tumor cells by CTLs is a slow process, suggesting that the ratio of effector to target cells is determinant, or that additional cytotoxic contribution by other cell types is required to induce efficient tumor rejection. This review will focus on the publications that have imaged antitumor immune responses dynamically and discuss how this new information contributes to understand the implication of CTLs in tumor rejection.

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Ariel Savina, Audrey Peres, Ignacio Cebrian, Nuno Carmo, Catarina Moita, Nir Hacohen, Luis F Moita, Sebastian Amigorena (2009 Mar 31)

The small GTPase Rac2 controls phagosomal alkalinization and antigen crosspresentation selectively in CD8(+) dendritic cells.

Immunity : 544-55 : DOI : 10.1016/j.immuni.2009.01.013 En savoir plus
Résumé

A unique subpopulation of spleen dendritic cells (DCs) that express the CD8 surface marker efficiently present phagocytosed antigens to CD8(+) T lymphocytes in a process called « crosspresentation, » which initiates cytotoxic immune responses. We now show that the small GTPase Rac2 plays a critical role in antigen crosspresentation selectively in this DC subpopulation. In CD8(+) DCs, Rac2 determines the subcellular assembly of the NADPH oxidase complex (NOX2) to phagosomes, whereas in CD8(-) DCs, Rac1 mediates the assembly of NOX2 at the plasma membrane. In the absence of Rac2, the production of reactive oxygen species (ROS) in DC-phagosomes was abolished, the phagosomal pH dropped, and the efficiency of antigen crosspresentation was reduced. We conclude that the activity of Rac1 and 2 control crosspresentation in DC subpopulations through the regulation of phagosomal oxidation and pH.

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Florence Faure, Adriana Mantegazza, Charlotte Sadaka, Christine Sedlik, Francine Jotereau, Sebastian Amigorena (2009 Jan 9)

Long-lasting cross-presentation of tumor antigen in human DC.

European journal of immunology : 380-90 : DOI : 10.1002/eji.200838669 En savoir plus
Résumé

DC cross-present exogenous antigens on MHC class I molecules, a process required for the onset of anti-tumor immune responses. In order to study the cross-presentation of tumor antigens by human DC, we compared the pathways of cross-presentation of long peptides requiring internalization and intracellular processing with the direct presentation of short peptides, which does not require intracellular processing. We found that, after brief incubations with DC, short peptides were presented to CD8(+) T cells with higher efficiencies than long peptides. After longer times of chase in the absence of peptide, however, the efficiency of presentation of the two types of peptides was reversed. After 2-3 days, DC pulsed with long peptides still activated T cells efficiently, while DC pulsed with short peptides failed to do so. Long-lasting presentation of the long peptides was, at least in part, due to a stored persistent pool of antigen, which was still available for loading on MHC class I molecules after several days of chase. These results show that the use of long synthetic peptides allows the efficient, long-lasting, presentation of tumor antigens, suggesting that long peptides represent an interesting approach for active anti-tumor vaccination.

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Année de publication : 2008

María Mittelbrunn, Gloria Martínez del Hoyo, María López-Bravo, Noa B Martín-Cofreces, Alix Scholer, Stéphanie Hugues, Luc Fetler, Sebastián Amigorena, Carlos Ardavín, Francisco Sánchez-Madrid (2008 Sep 27)

Imaging of plasmacytoid dendritic cell interactions with T cells.

Blood : 75-84 : DOI : 10.1182/blood-2008-02-139865 En savoir plus
Résumé

Plasmacytoid dendritic cells (pDCs) efficiently produce type I interferon and participate in adaptive immune responses, although the molecular interactions between pDCs and antigen-specific T cells remain unknown. This study examines immune synapse (IS) formation between murine pDCs and CD4(+) T cells. Mature pDCs formed canonical ISs, involving relocation to the contact site of the microtubule-organizing center, F-actin, protein kinase C-, and pVav, and activation of early signaling molecules in T cells. However, immature pDCs were less efficient at forming conjugates with T cells and inducing IS formation, microtubule-organizing center translocation, and T-cell signaling and activation. Time-lapse videomicroscopy and 2-photon in vivo imaging of pDC-T-cell interactions revealed that immature pDCs preferentially mediated transient interactions, whereas mature pDCs promoted more stable contacts. Our data indicate that, under steady-state conditions, pDCs preferentially establish transient contacts with naive T cells and show a very modest immunogenic capability, whereas on maturation, pDCs are able to form long-lived contacts with T cells and significantly enhance their capacity to activate these lymphocytes.

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Adriana R Mantegazza, Ariel Savina, Mónica Vermeulen, Laura Pérez, Jorge Geffner, Olivier Hermine, Sergio D Rosenzweig, Florence Faure, Sebastián Amigorena (2008 Aug 7)

NADPH oxidase controls phagosomal pH and antigen cross-presentation in human dendritic cells.

Blood : 4712-22 : DOI : 10.1182/blood-2008-01-134791 En savoir plus
Résumé

The phagocyte NADPH oxidase (NOX2) is critical for the bactericidal activity of phagocytic cells and plays a major role in innate immunity. We showed recently that NOX2 activity in mouse dendritic cells (DCs) prevents acidification of phagosomes, promoting antigen cross-presentation. In order to investigate the role of NOX2 in the regulation of the phagosomal pH in human DCs, we analyzed the production of reactive oxygen species (ROS) and the phagosomal/endosomal pH in monocyte-derived DCs and macrophages (M(diameter)s) from healthy donors or patients with chronic granulomatous disease (CGD). As expected, we found that human M(diameter)s acidify their phagosomes more efficiently than human DCs. Accordingly, the expression of the vacuolar proton ATPase (V-H(+)-ATPase) was higher in M(diameter)s than in DCs. Phagosomal ROS production, however, was also higher in M(diameter)s than in DCs, due to higher levels of gp91phox expression and recruitment to phagosomes. In contrast, in the absence of active NOX2, the phagosomal and endosomal pH decreased. Both in the presence of a NOX2 inhibitor and in DCs derived from patients with CGD, the cross-presentation of 2 model tumor antigens was impaired. We conclude that NOX2 activity participates in the regulation of the phagosomal and endosomal pH in human DCs, and is required for efficient antigen cross-presentation.

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Elisa Tagliani, Pierre Guermonprez, Jorge Sepúlveda, María López-Bravo, Carlos Ardavín, Sebastian Amigorena, Federica Benvenuti, Oscar R Burrone (2008 Feb 23)

Selection of an antibody library identifies a pathway to induce immunity by targeting CD36 on steady-state CD8 alpha+ dendritic cells.

Journal of immunology (Baltimore, Md. : 1950) : 3201-9 En savoir plus
Résumé

Improvement of the strategy to target tumor Ags to dendritic cells (DCs) for immunotherapy requires the identification of the most appropriate ligand/receptor pairing. We screened a library of Ab fragments on mouse DCs to isolate new potential Abs capable of inducing protective immune responses. The screening identified a high-affinity Ab against CD36, a multi-ligand scavenger receptor primarily expressed by the CD8alpha+ subset of conventional DCs. The Ab variable regions were genetically linked to the model Ag OVA and tested in Ag presentation assays in vitro and in vivo. Anti-CD36-OVA was capable of delivering exogenous Ags to the MHC class I and MHC class II processing pathways. In vivo, immunization with anti-CD36-OVA induced robust activation of naive CD4+ and CD8+ Ag-specific T lymphocytes and the differentiation of primed CD8+ T cells into long-term effector CTLs. Vaccination with anti-CD36-OVA elicited humoral and cell-mediated protection from the growth of an Ag-specific tumor. Notably, the relative efficacy of targeting CD11c/CD8alpha+ via CD36 or DEC205 was qualitatively different. Anti-DEC205-OVA was more efficient than anti-CD36-OVA in inducing early events of naive CD8+ T cell activation. In contrast, long-term persistence of effector CTLs was stronger following immunization with anti-CD36-OVA and did not require the addition of exogenous maturation stimuli. The results identify CD36 as a novel potential target for immunotherapy and indicate that the outcome of the immune responses vary by targeting different receptors on CD8alpha+ DCs.

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Ingrid S Zeelenberg, Matias Ostrowski, Sophie Krumeich, Angélique Bobrie, Carolina Jancic, Alexandre Boissonnas, Alain Delcayre, Jean-Bernard Le Pecq, Béhazine Combadière, Sebastian Amigorena, Clotilde Théry (2008 Feb 19)

Targeting tumor antigens to secreted membrane vesicles in vivo induces efficient antitumor immune responses.

Cancer research : 1228-35 : DOI : 10.1158/0008-5472.CAN-07-3163 En savoir plus
Résumé

Expression of non-self antigens by tumors can induce activation of T cells in vivo, although this activation can lead to either immunity or tolerance. CD8+ T-cell activation can be direct (if the tumor expresses MHC class I molecules) or indirect (after the capture and cross-presentation of tumor antigens by dendritic cells). The modes of tumor antigen capture by dendritic cells in vivo remain unclear. Here we examine the immunogenicity of the same model antigen secreted by live tumors either in association with membrane vesicles (exosomes) or as a soluble protein. We have artificially addressed the antigen to secreted vesicles by coupling it to the factor VIII-like C1C2 domain of milk fat globule epidermal growth factor-factor VIII (MFG-E8)/lactadherin. We show that murine fibrosarcoma tumor cells that secrete vesicle-bound antigen grow slower than tumors that secrete soluble antigen in immunocompetent, but not in immunodeficient, host mice. This growth difference is due to the induction of a more potent antigen-specific antitumor immune response in vivo by the vesicle-bound than by the soluble antigen. Finally, in vivo secretion of the vesicle-bound antigen either by tumors or by vaccination with naked DNA protects against soluble antigen-secreting tumors. We conclude that the mode of secretion can determine the immunogenicity of tumor antigens and that manipulation of the mode of antigen secretion may be used to optimize antitumor vaccination protocols.

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Alix Scholer, Stéphanie Hugues, Alexandre Boissonnas, Luc Fetler, Sebastian Amigorena (2008 Feb 16)

Intercellular adhesion molecule-1-dependent stable interactions between T cells and dendritic cells determine CD8+ T cell memory.

Immunity : 258-70 : DOI : 10.1016/j.immuni.2007.12.016 En savoir plus
Résumé

The initiation of cytotoxic immune responses requires the direct interaction between naive CD8+ T lymphocytes and dendritic cells (DCs). Multiphoton imaging in intact lymph nodes (LNs) showed that during priming, naive T cells and DCs establish sequentially brief (i.e., minutes) and long (hours) antigen-specific contacts. We show here that the expression of the Intercellular Adhesion Molecule-1 (ICAM-1) by mature DCs is critical for long-lasting contacts with CD8+ T cells but dispensable for short-lived antigen-specific interactions. Serial brief DC-T cell contacts induced early CD8+ T cell activation, proliferation, and differentiation into effector cytotoxic T lymphocytes in the first few days after immunization. ICAM-1-deficient mature DCs, however, failed to induce fully effective priming, because CD8+ T cells produced reduced amounts of interferon gamma and were clonally depleted after 2 weeks. In addition, Icam1(-/-) mice failed to respond to rechallenge. We conclude that ICAM-1-dependent long-lasting interactions between mature DCs and naive CD8+ T cells determine the survival of activated CD8+ T cells and the establishment of effective memory.

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Clotilde Théry, Sebastian Amigorena, Graça Raposo, Aled Clayton (2008 Jan 30)

Isolation and characterization of exosomes from cell culture supernatants and biological fluids.

Current protocols in cell biology / editorial board, Juan S. Bonifacino ... [et al.] : Unit 3.22 : DOI : 10.1002/0471143030.cb0322s30 En savoir plus
Résumé

Exosomes are small membrane vesicles found in cell culture supernatants and in different biological fluids. Exosomes form in a particular population of endosomes, called multivesicular bodies (MVBs), by inward budding into the lumen of the compartment. Upon fusion of MVBs with the plasma membrane, these internal vesicles are secreted. Exosomes possess a defined set of membrane and cytosolic proteins. The physiological function of exosomes is still a matter of debate, but increasing results in various experimental systems suggest their involvement in multiple biological processes. Because both cell-culture supernatants and biological fluids contain different types of lipid membranes, it is critical to perform high-quality exosome purification. This unit describes different approaches for exosome purification from various sources, and discusses methods to evaluate the purity and homogeneity of the purified exosome preparations.

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Cinzia Nobile, Marianne Lind, Francesc Miro, Karine Chemin, Marie Tourret, Giovanni Occhipinti, Stéphanie Dogniaux, Sebastian Amigorena, Claire Hivroz (2008 Jan 22)

Cognate CD4+ T-cell-dendritic cell interactions induce migration of immature dendritic cells through dissolution of their podosomes.

Blood : 3579-90 : DOI : 10.1182/blood-2007-08-107755 En savoir plus
Résumé

Dendritic cells (DCs) control T cell-based immunity. To do so they need to mature and migrate to sites of T-cell priming. We have previously shown that cognate interactions of human CD4+ T cells with DCs induce DC maturation. We show here that CC chemokines produced during antigen-specific T-DC interactions also induce strong morphologic modifications and migration of immature DCs. These modifications are required for efficient T-cell activation. Moreover, we show that CC chemokines produced during antigen-specific DC-T-cell interactions induce the dissolution of structures involved in cell motility and present on immature DCs (ie, podosomes). We thus propose a model in which chemokines secreted during Ag-specific contact between T cells and DCs induce disassembly of interacting and neighboring immature DC podosomes, leading to recruitment of more immature DCs toward sites of antigenic stimulation and to amplification of T-cell responses.

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Année de publication : 2007

Ariel Savina, Sebastian Amigorena (2007 Sep 14)

Phagocytosis and antigen presentation in dendritic cells.

Immunological reviews : 143-56 En savoir plus
Résumé

Like macrophages and neutrophils, dendritic cells (DCs) are considered professional phagocytes. Even if the three cell types phagocytose parasites, bacteria, cell debris, or even intact cells very efficiently, the functional outcomes of the phagocytic event are quite different. Macrophages and neutrophils scavenge and destroy phagocytosed particles, a critical step in innate immunity. DCs, in contrast, have developed means to ‘preserve’ useful information from the ingested particles that serve to initiate adaptive immune responses. Thus, both phagosomal degradation and acidification are much lower in DCs than in macrophages or neutrophils. Reduced degradation results in the conservation of antigenic peptides and in their increased presentation on major histocompatibility complex class I and II molecules. In this article, we review the mechanisms that control this delicate equilibrium between phagosomal degradation/cytotoxicity and antigen presentation in the different families of phagocytes.

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Lionel Apetoh, François Ghiringhelli, Antoine Tesniere, Michel Obeid, Carla Ortiz, Alfredo Criollo, Grégoire Mignot, M Chiara Maiuri, Evelyn Ullrich, Patrick Saulnier, Huan Yang, Sebastian Amigorena, Bernard Ryffel, Franck J Barrat, Paul Saftig, Francis Levi, Rosette Lidereau, Catherine Nogues, Jean-Paul Mira, Agnès Chompret, Virginie Joulin, Françoise Clavel-Chapelon, Jean Bourhis, Fabrice André, Suzette Delaloge, Thomas Tursz, Guido Kroemer, Laurence Zitvogel (2007 Aug 21)

Toll-like receptor 4-dependent contribution of the immune system to anticancer chemotherapy and radiotherapy.

Nature medicine : 1050-9 En savoir plus
Résumé

Conventional cancer treatments rely on radiotherapy and chemotherapy. Such treatments supposedly mediate their effects via the direct elimination of tumor cells. Here we show that the success of some protocols for anticancer therapy depends on innate and adaptive antitumor immune responses. We describe in both mice and humans a previously unrecognized pathway for the activation of tumor antigen-specific T-cell immunity that involves secretion of the high-mobility-group box 1 (HMGB1) alarmin protein by dying tumor cells and the action of HMGB1 on Toll-like receptor 4 (TLR4) expressed by dendritic cells (DCs). During chemotherapy or radiotherapy, DCs require signaling through TLR4 and its adaptor MyD88 for efficient processing and cross-presentation of antigen from dying tumor cells. Patients with breast cancer who carry a TLR4 loss-of-function allele relapse more quickly after radiotherapy and chemotherapy than those carrying the normal TLR4 allele. These results delineate a clinically relevant immunoadjuvant pathway triggered by tumor cell death.

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