Arribillaga, L. (Laura)
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- Induction of potent and long-lasting CD4 and CD8 T-cell responses against hepatitis C virus by immunization with viral antigens plus poly(I:C) and anti-CD40(Elsevier masson, 2007) Dotor, J. (Javier); Zabaleta, A. (Aintzane); Borras-Cuesta, F. (Francisco); Quer, J. (Josep); Esteban, J.I. (Juan Ignacio); Prieto, J. (Jesús); Llopiz, D. (Diana); Vayreda, F. (Francesc); Sarobe, P. (Pablo); Arribillaga, L. (Laura); Lasarte, J.J. (Juan José)Development of vaccination strategies against hepatitis C virus (HCV) is of paramount importance. With this aim, we tested the ability of dendritic cell-activating reagents polyinosinic-polycytidylic acid (poly(I:C)) and anti-CD40, as adjuvants to induce T-cell responses against HCV. Immunization of mice with these adjuvants induced dendritic cell maturation in vivo. Also, joint administration of poly(I:C) and anti-CD40 plus HCV antigens had a synergistic effect on the induction of anti-HCV T-cell responses. CD4 responses displayed a Th1 cytokine profile, and CD8 responses could be induced by immunization with a minimal CD8 epitope. Addition of a low amount of NS3 protein (as a source of Th epitopes) to the immunization mixture enhanced CD8 responses, whereas immunization with higher doses of NS3 induced both CD4 and CD8 responses. Surprisingly, immunization with NS3 protein but not with CD8 epitopes was able to induce CD8 responses and able to recognize cells expressing HCV antigens endogenously. Moreover, immunization with these adjuvants activated NK cells, which in turn helped to induce Th1 responses. Finally, this combined immunization protocol afforded long-lasting T-cell responses, suggesting that this strategy may prove to be useful in vaccination and/or treatment of HCV infection.
- Combination of a TLR4 ligand and anaphylatoxin C5a for the induction of antigen-specific cytotoxic T cell responses(Elsevier, 2012) Leclerc, C. (Claude); Fayolle, C. (Catherine); Pio, R. (Rubén); Lozano-Moreda, T. (Teresa); Rudilla, F. (Francesc); Durantez, M. (Maika); Casares, N. (Noelia); Prieto, J. (Jesús); Sarobe, P. (Pablo); Villanueva, L. (Lorea); Arribillaga, L. (Laura); Lasarte, J.J. (Juan José)The complement system and Toll-like receptors (TLR) are key innate defense systems which might interact synergistically on dendritic cells (DC) to reinforce adaptive immunity. In a previous work, we found that the extra domain A from fibronectin EDA (an endogenous ligand for TLR4) can favour antigen delivery to DC and induce their maturation. Given the potential of anaphylatoxins to cause inflammation and activation of myeloid cells, we hypothesized that a fusion protein between EDA, and anaphylatoxins C3a, C4a or C5a together with an antigen might improve the immunogenicity of the antigen. Naked DNA immunization with a construct expressing the fusion protein between C5a, EDA and the cytotoxic T cell epitope SIINFEKL from ovalbumin, induced strong antigen specific T cell responses. The purified recombinant fusion protein EDA-SIINFEKL-C5a induced activation of dendritic cells, the production of proinflammatory cytokines/chemokines and stimulated antigen presenting cell migration and NK cell activation. As compared to EDA-SIINFEKL, the fusion protein EDA-SIINFEKL-C5a did not induce the production of the immunosuppressive molecules IL-10, CCL17, CCL1, CXCL12 or XCL1 by DC. Moreover, EDA-SIINFEKL-C5a induced strong specific T cell responses in vivo and protected mice against E.G7-OVA tumor growth more efficiently than EDA-SIINFEKL or SIINFEKL-C5a recombinant proteins. Our results suggest that fusion proteins containing EDA, the anaphylatoxin C5a and the antigen may serve as a suitable strategy for the development of anti-tumor or anti-viral vaccines.
- Vaccination against hepatitis C virus with dendritic cells transduced with an adenovirus encoding NS3 protein(Nature publishing group, 2008) Riezu-Boj, J.I. (José Ignacio); Zabaleta, A. (Aintzane); Silva, L. (Leyre); Borras-Cuesta, F. (Francisco); Prieto, J. (Jesús); Llopiz, D. (Diana); Sarobe, P. (Pablo); Arribillaga, L. (Laura); Lasarte, J.J. (Juan José)Chronic infection by hepatitis C virus (HCV) is characterized by the absence of efficient antiviral T-cell responses. Thus, vaccination strategies to induce strong anti-HCV T-cell responses are of paramount importance for prophylactic and therapeutic purposes. Dendritic cells (DCs) are the most potent antigen presenting cells; therefore, immunization with these cells loaded with viral antigens offers a new approach for induction of antiviral immunity. Here we show that immunization with DCs transfected with an adenovirus encoding non-structural 3 protein, from HCV (AdNS3), induced multiepitopic CD4 T helper cell 1 (Th1) and CD8 T-cell responses in different mouse strains. These responses prevented the growth of a tumorexpressing HCV proteins, in short- and long-term experiments. Moreover, immunization with AdNS3-transfected DCs did not induce anti-adenoviral antibodies, as compared to direct immunization with AdNS3, but elicited T-cell responses even in the presence of pre-existing anti-adenoviral antibodies. Finally, responses induced by this protocol down-regulated the expression of HCV RNA in the liver. In conclusion, DCs transfected with AdNS3 may prove to be an efficient anti-HCV vaccine.
- Therapeutic effect of a peptide inhibitor of TGF-β on pulmonary fibrosis(Elsevier, 2011) Riezu-Boj, J.I. (José Ignacio); Dotor, J. (Javier); Borras-Cuesta, F. (Francisco); Basagoiti, M. (María); Sarobe, P. (Pablo); Arribillaga, L. (Laura); Cornet, M.E. (María Eugenia); Feijoo, E. (Esperanza); Lasarte, J.J. (Juan José)Pulmonary fibrosis encompasses several respiratory diseases characterized by epithelial cell injury, inflammation and fibrosis. Transforming growth factor (TGF)-β1 is one of the main profibrogenic cytokines involved in the pathogenesis of lung fibrosis. It induces fibroblast differentiation into myofibroblasts, which produce high levels of collagen and concomitantly loss of lung elasticity and reduction of the respiratory function. In the present study, we have investigated the effects of P17 (a TGF-β inhibitor peptide) on IMR-90 lung fibroblast differentiation in vitro, as well as on the inhibition of the development of bleomycin-induced pulmonary fibrosis in mice. It was found that in IMR-90 cells, P17 inhibited TGF-β1-induced expression of connective tissue growth factor and α-smooth muscle actin. In vivo, treatment of mice with P17 2days after bleomycin administration decreased lung fibrosis, areas of myofibroblast-like cells and lymphocyte infiltrate. P17 also reduced mRNA expression of collagen type I, fibronectin and the fibronectin splice isoform EDA in the lung, and increased the expression of IFN-γ mRNA. Finally, therapeutic treatment with P17 in mice with already established fibrosis was able to significantly attenuate the progression of lung fibrosis. These results suggest that P17 may be useful in the treatment of pulmonary fibrosis.
- Hepatitis C virus structural proteins impair dendritic cell maturation and inhibit in vivo induction of cellular immune responses(American society for microbiology, 2003) Zabaleta, A. (Aintzane); Borras-Cuesta, F. (Francisco); Melero, I. (Ignacio); Prieto, J. (Jesús); Sarobe, P. (Pablo); Arribillaga, L. (Laura); Arina, A. (Ainhoa); Lasarte, J.J. (Juan José)Hepatitis C virus (HCV) chronic infection is characterized by low or undetectable cellular immune responses against HCV antigens. Some studies have suggested that HCV proteins manipulate the immune system by suppressing the specific antiviral T-cell immunity. We have previously reported that the expression of HCV core and E1 proteins (CE1) in dendritic cells (DC) impairs their ability to prime T cells in vitro. We show here that immunization of mice with immature DC transduced with an adenovirus encoding HCV core and E1 antigens (AdCE1) induced lower CD4(+)- and CD8(+)-T-cell responses than immunization with DC transduced with an adenovirus encoding NS3 (AdNS3). However, no differences in the strength of the immune response were detected when animals were immunized with mature DC subsequently transduced with AdCE1 or AdNS3. According to these findings, we observed that the expression of CE1 in DC inhibited the maturation caused by tumor necrosis factor alpha or CD40L but not that induced by lipopolysaccharide. Blockade of DC maturation by CE1 was manifested by a lower expression of maturation surface markers and was associated with a reduced ability of AdCE1-transduced DC to activate CD4(+)- and CD8(+)-T-cell responses in vivo. Our results suggest that HCV CE1 proteins modulate T-cell responses by decreasing the stimulatory ability of DC in vivo via inhibition of their physiological maturation pathways. These findings are relevant for the design of therapeutic vaccination strategies in HCV-infected patients.
- Vaccination with an adenoviral vector encoding hepatitis C virus (HCV) NS3 protein protects against infection with HCV-recombinant vaccinia virus(Elsevier, 2002) Paranhos-Baccala, G. (Glaucia); Bruña-Romero, O. (Oscar); Vales, A. (África); Borras-Cuesta, F. (Francisco); Lopez-Diaz-de-Cerio, A. (Ascensión); Casares, N. (Noelia); Prieto, J. (Jesús); Sarobe, P. (Pablo); Gorraiz, M. (Marta); Arribillaga, L. (Laura); Ruiz, J. (Juan); Lasarte, J.J. (Juan José)Cellular immune response plays an important role in the clearance of hepatitis C virus (HCV). Thus, development of efficient ways to induce anti-viral cellular immune responses is an important step toward prevention and/or treatment of HCV infection. With this aim, we have constructed a replication-deficient recombinant adenovirus expressing HCV NS3 protein (RAdNS3). The efficacy of RAdNS3 was tested in vivo by measuring the protection against infection with a recombinant vaccinia virus expressing HCV-polyprotein (vHCV1-3011). Immunisation with 10(9)pfu of RAdNS3 induced anti-NS3 humoral, T helper and T cytotoxic responses. We identified eight epitopes recognised by IFN-gamma producing cells, five of them exhibiting lytic activity. Moreover, we show that RAdNS3 immunised mice were protected against challenge with vHCV1-3011 and that this protection was mediated by CD8(+) cells. In conclusion, our results suggest that adenoviral vectors encoding NS3 might be useful for the induction of prophylactic and/or therapeutic anti-HCV immunity.
- A fusion protein between streptavidin and the endogenous TLR4 ligand EDA targets biotinylated antigens to dendritic cells and induces T cell responses in vivo(2013) Borras-Cuesta, F. (Francisco); Lozano-Moreda, T. (Teresa); Martinez, M. (Marta); Rudilla, F. (Francesc); Durantez, M. (Maika); Casares, N. (Noelia); Rehberger-Bescos, F. (Federico); Prieto, J. (Jesús); Sarobe, P. (Pablo); Villanueva, L. (Lorea); Gorraiz, M. (Marta); Arribillaga, L. (Laura); Lasarte, J.J. (Juan José)The development of tools for efficient targeting of antigens to antigen presenting cells is of great importance for vaccine development. We have previously shown that fusion proteins containing antigens fused to the extra domain A from fibronectin (EDA), an endogenous TLR4 ligand, which targets antigens to TLR4-expressing dendritic cells (DC), are highly immunogenic. To facilitate the procedure of joining EDA to any antigen of choice, we have prepared the fusion protein EDAvidin by linking EDA to the N terminus of streptavidin, allowing its conjugation with biotinylated antigens. We found that EDAvidin, as streptavidin, forms tetramers and binds biotin or biotinylated proteins with a ~ 2.6 × 10−14 mol/L. EDAvidin favours the uptake of biotinylated green fluorescent protein by DC. Moreover, EDAvidin retains the proinflammatory properties of EDA, inducing NF-κβ by TLR4-expressing cells, as well as the production of TNF-α by the human monocyte cell line THP1 and IL-12 by DC. More importantly, immunization of mice with EDAvidin conjugated with the biotinylated nonstructural NS3 protein from hepatitis C virus induces a strong anti-NS3 T cell immune response. These results open a new way to use the EDA-based delivery tool to target any antigen of choice to DC for vaccination against infectious diseases and cancer.
- A recombinant adenovirus encoding hepatitis C virus core and E1 proteins protects mice against cytokine-induced liver damage(Wiley-Blackwell, 2003) Borras-Cuesta, F. (Francisco); Lopez-Diaz-de-Cerio, A. (Ascensión); Boya, P. (Patricia); Casares, N. (Noelia); Prieto, J. (Jesús); Sarobe, P. (Pablo); Gorraiz, M. (Marta); Arribillaga, L. (Laura); Lasarte, J.J. (Juan José)Hepatitis C virus (HCV) infection has a strong tendency to evolve to chronicity despite up-regulation of proapoptotic cytokines in the inflamed liver. The mechanisms responsible for persistent viral replication in this inflammatory environment are obscure. It is conceivable that viral replication would be facilitated if the infected hepatocytes are rendered resistant to cytokine-induced cytotoxicity. In this study, we investigated if an adenovirus encoding HCV core and E1 (RAdCE1) could reduce liver cell injury in different in vivo models of cytokine-mediated hepatotoxicity in mice. We show that RAdCE1 markedly attenuates hepatocellular apoptosis and the increase in serum transaminase levels after concanavalin A (con A) challenge. This protective effect is accompanied by an inhibition of nuclear translocation of nuclear factor kappaB (NF-kappaB); reduced expression of inducible nitric oxide synthase (iNOS); decreased hepatic messenger RNA levels of chemokines macrophage inflammatory protein 2 (MIP-2), monocyte chemoattractant protein 1 (MCP-1), and interferon-inducible protein 10 (IP-10); and abrogation of liver leukocyte infiltration. RAdCE1 also causes a reduction in serum transaminase levels and inhibits hepatocellular apoptosis in mice given tumor necrosis factor (TNF)-alpha plus D-galactosamine. In conclusion, HCV structural antigens can protect liver cells against the proapoptotic effects of proinflammatory cytokines. The antiapoptotic status of infected liver cells may represent a mechanism favoring viral persistence. Our findings also suggest that, in chronic hepatitis C, the burden of hepatocellular damage mainly affects noninfected liver cells.
- Enhancement of CD4 and CD8 immunity by anti-CD137 (4-1BB) monoclonal antibodies during hepatitis C vaccination with recombinant adenovirus(Elsevier, 2005) Borras-Cuesta, F. (Francisco); Melero, I. (Ignacio); Prieto, J. (Jesús); Sarobe, P. (Pablo); Gorraiz, M. (Marta); Arribillaga, L. (Laura); Ruiz, J. (Juan); Chen, L. (Lieping); Arina, A. (Ainhoa); Lasarte, J.J. (Juan José)The induction of protective or therapeutic cellular immunity against hepatitis C virus (HCV) is a difficult goal. In a previous work we showed that immunization with a recombinant adenovirus encoding HCV-NS3 (RAdNS3) could partially protect mice from challenge with a vaccinia virus encoding HCV antigens. We sought to investigate whether systemic administration of an immunostimulatory monoclonal antibody directed against the lymphocyte surface molecule CD137 could enhance the immunity elicited by RAdNS3. It was found that treatment with anti-CD137 mAb after the administration of a suboptimal dose of RAdNS3 enhanced cytotoxic and T helper cell responses against HCV NS3. Importantly, the ability of RAdNS3 to induce protective immunity against challenge with a recombinant vaccinia virus expressing HCV proteins was markedly augmented. Thus, combination of immunostimulatory anti-CD137 mAb with recombinant adenoviruses expressing HCV proteins might be useful in strategies of immunization against HCV.
- A peptide inhibitor of FOXP3 impairs regulatory T cell Activity and improves vaccine efficacy in mice(American Association of Immunologists, 2010-11-25) Riezu-Boj, J.I. (José Ignacio); Borras-Cuesta, F. (Francisco); Lozano-Moreda, T. (Teresa); Rudilla, F. (Francesc); Casares, N. (Noelia); Guembe, L. (L.); Prieto, J. (Jesús); Llopiz, D. (Diana); Sarobe, P. (Pablo); Arribillaga, L. (Laura); Lopez-Sagaseta, J. (Jacinto); Lasarte, J.J. (Juan José)Immunosuppressive activity of regulatory T cells (Treg) may contribute to the progression of cancer or infectious diseases by preventing the induction of specific immune responses. Using a phage-displayed random peptide library, we identified a 15-mer synthetic peptide, P60, able to bind to forkhead/winged helix transcription factor 3 (FOXP3), a factor required for development and function of Treg. P60 enters the cells, inhibits FOXP3 nuclear translocation, and reduces its ability to suppress the transcription factors NF-κB and NFAT. In vitro, P60 inhibited murine and human-derived Treg and improved effector T cell stimulation. P60 administration to newborn mice induced a lymphoproliferative autoimmune syndrome resembling the reported pathology in scurfy mice lacking functional Foxp3. However, P60 did not cause toxic effects in adult mice and, when given to BALB/c mice immunized with the cytotoxic T cell epitope AH1 from CT26 tumor cells, it induced protection against tumor implantation. Similarly, P60 improved the antiviral efficacy of a recombinant adenovirus expressing NS3 protein from hepatitis C virus. Functional inhibition of Treg by the FOXP3-inhibitory peptide P60 constitutes a strategy to enhance antitumor and antiviral immunotherapies.