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Noelia Casares

Publications and source records attributed to Noelia Casares.

16 recordsLinked to original sources
PubMed2007

Upregulation of indoleamine 2,3-dioxygenase in hepatitis C virus infection.

Indoleamine 2,3-dioxygenase (IDO) is induced by proinflammatory cytokines and by CTLA-4-expressing T cells and constitutes an important mediator of peripheral immune tolerance. In chronic hepatitis C, we found upregulation of IDO expression in the liver and an increased serum kynurenine/tryptophan ratio (a reflection of IDO activity). Huh7 cells supporting hepatitis C virus (HCV) replication expressed higher levels of IDO mRNA than noninfected cells when stimulated with gamma interferon or when cocultured with activated T cells. In infected chimpanzees, hepatic IDO expression decreased in animals that cured the infection, while it remained high in those that progressed to chronicity. For both patients and chimpanzees, hepatic expression of IDO and CTLA-4 correlated directly. Induction of IDO may dampen T-cell reactivity to viral antigens in chronic HCV infection.

Antigens, CD↗
PubMed2007

The extra domain A from fibronectin targets antigens to TLR4-expressing cells and induces cytotoxic T cell responses in vivo.

Vaccination strategies based on the in vivo targeting of Ags to dendritic cells (DCs) are needed to improve the induction of specific T cell immunity against tumors and infectious agents. In this study, we have used a recombinant protein encompassing the extra domain A from fibronectin (EDA), an endogenous ligand for TLR4, to deliver Ags to TLR4-expressing DC. The purified EDA protein was shown to bind to TLR4-expressing HEK293 cells and to activate the TLR4 signaling pathway. EDA also stimulated the production by DC of proinflammatory cytokines such as IL-12 or TNF-alpha and induced their maturation in vitro and in vivo. A fusion protein between EDA and a cytotoxic T cell epitope from OVA efficiently presented this epitope to specific T cells and induced the in vivo activation of a strong and specific CTL response. Moreover, a fusion protein containing EDA and the full OVA also improved OVA presentation by DC and induced CTL responses in vivo. These EDA recombinant proteins protected mice from a challenge with tumor cells expressing OVA. These results strongly suggest that the fibronectin extra domain A may serve as a suitable Ag carrier for the development of antiviral or antitumoral vaccines.

Amino Acid Motifs↗
PubMed2006

Calreticulin exposure dictates the immunogenicity of cancer cell death.

Anthracyclin-treated tumor cells are particularly effective in eliciting an anticancer immune response, whereas other DNA-damaging agents such as etoposide and mitomycin C do not induce immunogenic cell death. Here we show that anthracyclins induce the rapid, preapoptotic translocation of calreticulin (CRT) to the cell surface. Blockade or knockdown of CRT suppressed the phagocytosis of anthracyclin-treated tumor cells by dendritic cells and abolished their immunogenicity in mice. The anthracyclin-induced CRT translocation was mimicked by inhibition of the protein phosphatase 1/GADD34 complex. Administration of recombinant CRT or inhibitors of protein phosphatase 1/GADD34 restored the immunogenicity of cell death elicited by etoposide and mitomycin C, and enhanced their antitumor effects in vivo. These data identify CRT as a key feature determining anticancer immune responses and delineate a possible strategy for immunogenic chemotherapy.

Animals↗
PubMed2006

Apoptosis regulation in tetraploid cancer cells.

Tetraploidy can result in cancer-associated aneuploidy. As shown here, freshly generated tetraploid cells arising due to mitotic slippage or failed cytokinesis are prone to undergo Bax-dependent mitochondrial membrane permeabilization and subsequent apoptosis. Knockout of Bax or overexpression of Bcl-2 facilitated the survival of tetraploid cells at least as efficiently as the p53 or p21 knockout. When tetraploid cells were derived from diploid p53 and Bax-proficient precursors, such cells exhibited an enhanced transcription of p53 target genes. Tetraploid cells exhibited an enhanced rate of spontaneous apoptosis that could be suppressed by inhibition of p53 or by knockdown of proapoptotic p53 target genes such as BBC3/Puma, GADD45A and ferredoxin reductase. Unexpectedly, tetraploid cells were more resistant to DNA damaging agents (cisplatin, oxaliplatin and camptothecin) than their diploid counterparts, and this difference disappeared upon inhibition of p53 or knockdown of p53-inducible ribonucleotide reductase. Tetraploid cells were also more resistant against UVC and gamma-irradiation. These data indicate the existence of p53-dependent alterations in apoptosis regulation in tetraploid cells.

Animals↗
PubMed2006

A novel dendritic cell subset involved in tumor immunosurveillance.

The interferon (IFN)-gamma-induced TRAIL effector mechanism is a vital component of cancer immunosurveillance by natural killer (NK) cells in mice. Here we show that the main source of IFN-gamma is not the conventional NK cell but a subset of B220(+)Ly6C(-) dendritic cells, which are atypical insofar as they express NK cell-surface molecules. Upon contact with a variety of tumor cells that are poorly recognized by NK cells, B220(+)NK1.1(+) dendritic cells secrete high levels of IFN-gamma and mediate TRAIL-dependent lysis of tumor cells. Adoptive transfer of these IFN-producing killer dendritic cells (IKDCs) into tumor-bearing Rag2(-/-)Il2rg(-/-) mice prevented tumor outgrowth, whereas transfer of conventional NK cells did not. In conclusion, we identified IKDCs as pivotal sensors and effectors of the innate antitumor immune response.

Adoptive Transfer↗
PubMed2005

Caspase-dependent immunogenicity of doxorubicin-induced tumor cell death.

Systemic anticancer chemotherapy is immunosuppressive and mostly induces nonimmunogenic tumor cell death. Here, we show that even in the absence of any adjuvant, tumor cells dying in response to anthracyclins can elicit an effective antitumor immune response that suppresses the growth of inoculated tumors or leads to the regression of established neoplasia. Although both antracyclins and mitomycin C induced apoptosis with caspase activation, only anthracyclin-induced immunogenic cell death was immunogenic. Caspase inhibition by Z-VAD-fmk or transfection with the baculovirus inhibitor p35 did not inhibit doxorubicin (DX)-induced cell death, yet suppressed the immunogenicity of dying tumor cells in several rodent models of neoplasia. Depletion of dendritic cells (DCs) or CD8+T cells abolished the immune response against DX-treated apoptotic tumor cells in vivo. Caspase inhibition suppressed the capacity of DX-killed cells to be phagocytosed by DCs, yet had no effect on their capacity to elicit DC maturation. Freshly excised tumors became immunogenic upon DX treatment in vitro, and intratumoral inoculation of DX could trigger the regression of established tumors in immunocompetent mice. These results delineate a procedure for the generation of cancer vaccines and the stimulation of anti-neoplastic immune responses in vivo.

Amino Acid Chloromethyl Ketones↗
PubMed2005

Inhibition of macroautophagy triggers apoptosis.

Mammalian cells were observed to die under conditions in which nutrients were depleted and, simultaneously, macroautophagy was inhibited either genetically (by a small interfering RNA targeting Atg5, Atg6/Beclin 1-1, Atg10, or Atg12) or pharmacologically (by 3-methyladenine, hydroxychloroquine, bafilomycin A1, or monensin). Cell death occurred through apoptosis (type 1 cell death), since it was reduced by stabilization of mitochondrial membranes (with Bcl-2 or vMIA, a cytomegalovirus-derived gene) or by caspase inhibition. Under conditions in which the fusion between lysosomes and autophagosomes was inhibited, the formation of autophagic vacuoles was enhanced at a preapoptotic stage, as indicated by accumulation of LC3-II protein, ultrastructural studies, and an increase in the acidic vacuolar compartment. Cells exhibiting a morphology reminiscent of (autophagic) type 2 cell death, however, recovered, and only cells with a disrupted mitochondrial transmembrane potential were beyond the point of no return and inexorably died even under optimal culture conditions. All together, these data indicate that autophagy may be cytoprotective, at least under conditions of nutrient depletion, and point to an important cross talk between type 1 and type 2 cell death pathways.

Adenine↗
PubMed2004

AIF deficiency compromises oxidative phosphorylation.

Apoptosis-inducing factor (AIF) is a mitochondrial flavoprotein that, after apoptosis induction, translocates to the nucleus where it participates in apoptotic chromatinolysis. Here, we show that human or mouse cells lacking AIF as a result of homologous recombination or small interfering RNA exhibit high lactate production and enhanced dependency on glycolytic ATP generation, due to severe reduction of respiratory chain complex I activity. Although AIF itself is not a part of complex I, AIF-deficient cells exhibit a reduced content of complex I and of its components, pointing to a role of AIF in the biogenesis and/or maintenance of this polyprotein complex. Harlequin mice with reduced AIF expression due to a retroviral insertion into the AIF gene also manifest a reduced oxidative phosphorylation (OXPHOS) in the retina and in the brain, correlating with reduced expression of complex I subunits, retinal degeneration, and neuronal defects. Altogether, these data point to a role of AIF in OXPHOS and emphasize the dual role of AIF in life and death.

Adenosine Triphosphate↗
PubMed2004

Increased immunogenicity of colon cancer cells by selective depletion of cytochrome C.

We and others have previously reported in an in vivo rat colon cancer cell model that cell death precedes and is necessary for the development of a specific antitumoral immune response. To sensitize colon cancer cells to death, we depleted cytochrome c by stable transfection with an antisense construct. Cytochrome c depletion sensitizes human and rat colon cancer cells to a nonapoptotic, nonautophagic death induced by various stimuli. This increased sensitization to a necrosis-like cell death may be related to a decrease in cellular ATP levels and an increase in reactive oxygen species production caused by cytochrome c depletion. In vivo, depletion of cytochrome c decreases the tumorigenicity of colon cancer cells in syngeneic rats without influencing their growth in immune-deficient animals. Furthermore, decreased expression of cytochrome c in tumor cells facilitates in vivo "necrotic" cell death and the induction of a specific immune response. These results delineate a novel strategy to sensitize colon cancer cells to chemotherapy and to increase their immunogenicity in immuno-competent hosts.

Animals↗
PubMed2003

CD4+/CD25+ regulatory cells inhibit activation of tumor-primed CD4+ T cells with IFN-gamma-dependent antiangiogenic activity, as well as long-lasting tumor immunity elicited by peptide vaccination.

CD25(+) regulatory T (T reg) cells suppress the activation/proliferation of other CD4(+) or CD8(+) T cells in vitro. Also, down-regulation of CD25(+) T reg cells enhance antitumor immune responses. In this study, we show that depletion of CD25(+) T reg cells allows the host to induce both CD4(+) and CD8(+) antitumoral responses following tumor challenge. Simultaneous depletion of CD25(+) and CD8(+) cells, as well as adoptive transfer experiments, revealed that tumor-specific CD4(+) T cells, which emerged in the absence of CD25(+) T reg cells, were able to reject CT26 colon cancer cells, a MHC class II-negative tumor. The antitumoral effect mediated by CD4(+) T cells was dependent on IFN-gamma production, which exerted a potent antiangiogenic activity. The capacity of the host to mount this antitumor response is lost once the number of CD25(+) T reg cells is restored over time. However, CD25(+) T reg cell depletion before immunization with AH1 (a cytotoxic T cell determinant from CT26 tumor cells) permits the induction of a long-lasting antitumoral immune response, not observed if immunization is conducted in the presence of regulatory cells. A study of the effect of different levels of depletion of CD25(+) T reg cells before immunization with the peptide AH1 alone, or in combination with a Th determinant, unraveled that Th cells play an important role in overcoming the suppressive effect of CD25(+) T reg on the induction of long-lasting cellular immune responses.

Adoptive Transfer↗
PubMed2003

A recombinant adenovirus encoding hepatitis C virus core and E1 proteins protects mice against cytokine-induced liver damage.

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.

Acute Disease↗
PubMed2003

Engineering Th determinants for efficient priming of humoral and cytotoxic T cell responses.

To engineer T(h) determinants (THd) to prime help for humoral or cytotoxic T cell responses, we modified ovalbumin [OVA(323-337)] and myoglobin [MYO(106-118)] eliciting T(h)1 and T(h)0 cytokine profiles respectively. Residues along the sequence of both THd were replaced with amino acids representative of different families. Replacements at positions P-1 and P5 pointing to the TCR in both THd afforded higher levels of IFN-gamma and IL-4 production. Peptides eliciting different proportions of IFN-gamma and IL-4 were co-immunized with a peptide hapten or a T cytotoxic determinant (TCd) respectively. OVA(323-337)- and MYO(106-118)-derived peptides afforded the best THd for the induction of cytotoxic T lymphocyte (CTL) and anti-hapten antibodies respectively. IFN-gamma and IL-4, primed by MYO(106-118)-derived peptides, correlated significantly with antibody production against the hapten (P < 0.05 for IFN-gamma and P < 0.05 for IL-4). Interestingly, two peptides derived from OVA(323-337), 323G and 327G, which induced the clearest T(h)2 cytokine profiles, were not the most efficient to prime cell help for the induction of anti-hapten antibodies. For CTL induction, OVA(323-337)-derived peptides, inducing a T(h)1-like profile, required a lower dose (5 nmol) than T(h)0 peptides (50 nmol). The dose of 50 nmol was detrimental for T(h)1-like peptides. Interestingly, IFN-gamma primed by the THd correlated significantly with that induced by the TCd (P < 0.01).

Animals↗
PubMed2002

Vaccination with an adenoviral vector encoding hepatitis C virus (HCV) NS3 protein protects against infection with HCV-recombinant vaccinia virus.

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.

Adenoviridae↗
PubMed2002

Abnormal priming of CD4(+) T cells by dendritic cells expressing hepatitis C virus core and E1 proteins.

Patients infected with hepatitis C virus (HCV) have an impaired response against HCV antigens while keeping immune competence for other antigens. We hypothesized that expression of HCV proteins in infected dendritic cells (DC) might impair their antigen-presenting function, leading to a defective anti-HCV T-cell immunity. To test this hypothesis, DC from normal donors were transduced with an adenovirus coding for HCV core and E1 proteins and these cells (DC-CE1) were used to stimulate T lymphocytes. DC-CE1 were poor stimulators of allogeneic reactions and of autologous primary and secondary proliferative responses. Autologous T cells stimulated with DC-CE1 exhibited a pattern of incomplete activation characterized by enhanced CD25 expression but reduced interleukin 2 production. The same pattern of incomplete lymphocyte activation was observed in CD4(+) T cells responding to HCV core in patients with chronic HCV infection. However, CD4(+) response to HCV core was normal in patients who cleared HCV after alpha interferon therapy. Moreover, a normal CD4(+) response to tetanus toxoid was found in both chronic HCV carriers and patients who had eliminated the infection. Our results suggest that expression of HCV structural antigens in infected DC disturbs their antigen-presenting function, leading to incomplete activation of anti-HCV-specific T cells and chronicity of infection. However, presentation of unrelated antigens by noninfected DC would allow normal T-cell immunity to other pathogens.

Adenoviridae↗
PubMed2002

Enhancing immunogenicity of a CTL epitope from carcinoembryonic antigen by selective amino acid replacements.

PURPOSE: Many epitopes from tumor antigens recognized by CTLs canbe poorly immunogenic. This low immunogenicity can be improved by carrying out amino acid replacements in their sequence. We have applied this strategy to enhance the immunogenicity of the HLA-A2-restricted CTL epitope CEA691 (IMIGVLVGV) from carcinoembryonic antigen (CEA), which is expressed by a wide variety of tumors. EXPERIMENTAL DESIGN: Substituted peptides from CEA691 were synthesized and tested in HLA-A2-binding assays, and in recognition by CEA691-specific CTL. Selected peptides were used to induce CTL responses in vivo in HLA-A2Kb transgenic mice and in vitro with human cells. RESULTS: Our experiments afforded several peptides with enhanced binding and/or recognition by CTL specific of CEA691. However, when HLA-A2Kb mice were immunized with these peptides only a few induced a CTL response that cross-reacted with CEA691. Additional replacement of their NH(2)-terminal amino acid by Y (tyrosine) afforded peptides YMIGMLVGV and YMIGVLLGV with enhanced in vivo and in vitro immunogenicity than CEA691. Indeed, they activated a greater number of precursor cells that recognized CEA691-pulsed cells and tumor cells expressing CEA. CONCLUSIONS: Our results widen the possibility of treating CEA-expressing tumors with enhanced efficacy.

Animals↗