Trypanosoma cruzi Adjuvants Potentiate T Cell-Mediated Immunity Induced by a NY-ESO-1 Based Antitumor Vaccine

Immunological adjuvants that induce T cell-mediate immunity (TCMI) with the least side effects are needed for the development of human vaccines. Glycoinositolphospholipids (GIPL) and CpGs oligodeoxynucleotides (CpG ODNs) derived from the protozoa parasite Trypanosoma cruzi induce potent pro-inflammatory reaction through activation of Toll-Like Receptor (TLR)4 and TLR9, respectively. Here, using mouse models, we tested the T. cruzi derived TLR agonists as immunological adjuvants in an antitumor vaccine. For comparison, we used well-established TLR agonists, such as the bacterial derived monophosphoryl lipid A (MPL), lipopeptide (Pam3Cys), and CpG ODN. All tested TLR agonists were comparable to induce antibody responses, whereas significant differences were noticed in their ability to elicit CD4+ T and CD8+ T cell responses. In particular, both GIPLs (GTH, and GY) and CpG ODNs (B344, B297 and B128) derived from T. cruzi elicited interferon-gamma (IFN-γ) production by CD4+ T cells. On the other hand, the parasite derived CpG ODNs, but not GIPLs, elicited a potent IFN-γ response by CD8+ T lymphocytes. The side effects were also evaluated by local pain (hypernociception). The intensity of hypernociception induced by vaccination was alleviated by administration of an analgesic drug without affecting protective immunity. Finally, the level of protective immunity against the NY-ESO-1 expressing melanoma was associated with the magnitude of both CD4+ T and CD8+ T cell responses elicited by a specific immunological adjuvant

High-Throughput Analysis of Synthetic Peptides for the Immunodiagnosis of Canine Visceral Leishmaniasis

Globally, the number of new human cases of visceral leishmaniasis (VL) is estimated to be approximately 500,000 per year. This is the most severe of all forms of leishmaniasis, and the zoonotic form of VL, caused by Leishmania infantum (also known as Leishmania chagasi), represents 20% of human visceral leishmaniasis worldwide; additionally, its prevalence is increasing in urban and peri-urban areas of the tropics. In Brazil, the identification and elimination of infected dogs, which act as a reservoir for Leishmania parasites, is a control measure employed in addition to the use of insecticides against the vectors and the identification and treatment of infected humans. Currently, the diagnostic methods employed to identify infected animals are not able to detect all of these dogs, which compromises the effectiveness of control measures. Moreover, one of the most important issues in controlling VL is the difficulty of diagnosing asymptomatic dogs, which act as parasite reservoirs. Therefore, to contribute to the improvement of the diagnostic methods for CVL, we aimed to identify and characterize new antigens that were more sensitive and specific and could be applied in epidemiologic surveys

Cross-priming of long lived protective CD8(+) T cells against Trypanosoma cruzi infection: Importance of a TLR9 agonist and CD4(+) T cells

We recently described that vaccination of mice with a gluthatione S transferase fusion protein representing amino acids 261-500 of the Amastigote Surface Protein-2 efficiently cross-primed protective CD8(+) T cells against a lethal challenge with the human protozoan parasite Trypanosoma cruzi. in this study, we initially established that this protective immunity was long lived. Subsequently, we studied the importance of TLR9 agonist CpG ODN 1826, TLR4 and CD4(+) T cells for the generation of these protective CD8(+) T cells. We found that: (i) the TLR9 agonist CpG ODN 1826 improved the efficiency of protective immunity; (ii) TLR4 is not relevant for priming of specific CD8(+) T cells; (iii) CD4(+) T cells are critical for priming of memory/protective CD8(+) T cells. (c) 2007 Elsevier B.V. All rights reserved.Universidade Federal de São Paulo, Escola Paulista Med, CINTERGEN, BR-04044010 São Paulo, BrazilUniversidade Federal de São Paulo, Escola Paulista Med, Dept Microbiol Imunol & Parasitol, BR-04044010 São Paulo, BrazilFiocruz MS, Ctr Pesquisas Rene Rachou, BR-30190002 Belo Horizonte, MG, BrazilUniv Fed Minas Gerais, BR-30190002 Belo Horizonte, MG, BrazilUniversidade Federal de São Paulo, Escola Paulista Med, CINTERGEN, BR-04044010 São Paulo, BrazilUniversidade Federal de São Paulo, Escola Paulista Med, Dept Microbiol Imunol & Parasitol, BR-04044010 São Paulo, BrazilWeb of Scienc

Distinct Kinetics of Effector CD8(+) Cytotoxic T Cells after Infection with Trypanosoma cruzi in Naïve or Vaccinated Mice

The kinetics of effector CD8(+)-T-cell responses to specific Trypanosoma cruzi epitopes was investigated after challenge. Our results suggest that the delayed kinetics differs from that observed in other microbial infections and facilitates the establishment of the disease in naïve mice. In contrast, in vaccinated mice, the swift CD8(+)-T-cell response helps host survival after challenge

CD8(+)-T-Cell-Dependent Control of Trypanosoma cruzi Infection in a Highly Susceptible Mouse Strain after Immunization with Recombinant Proteins Based on Amastigote Surface Protein 2

We previously described that DNA vaccination with the gene encoding amastigote surface protein 2 (ASP-2) protects approximately 65% of highly susceptible A/Sn mice against the lethal Trypanosoma cruzi infection. Here, we explored the possibility that bacterial recombinant proteins of ASP-2 could be used to improve the efficacy of vaccinations. Initially, we compared the protective efficacy of vaccination regimens using either a plasmid DNA, a recombinant protein, or both sequentially (DNA priming and protein boosting). Survival after the challenge was not statistically different among the three mouse groups and ranged from 53.5 to 75%. The fact that immunization with a recombinant protein alone induced protective immunity revealed the possibility that this strategy could be pursued for vaccination. We investigated this possibility by using six different recombinant proteins representing distinct portions of ASP-2. The vaccination of mice with glutathione S-transferase fusion proteins representing amino acids 261 to 500 or 261 to 380 of ASP-2 in the presence of the adjuvants alum and CpG oligodeoxynucleotide 1826 provided remarkable immunity, consistently protecting 100% of the A/Sn mice. Immunity was completely reversed by the in vivo depletion of CD8(+) T cells, but not CD4(+) T cells, and was associated with the presence of CD8(+) T cells specific for an epitope located between amino acids 320 and 327 of ASP-2. We concluded that a relatively simple formulation consisting of a recombinant protein with a selected portion of ASP-2, alum, and CpG oligodeoxynucleotide 1826 might be used to cross-prime strong CD8(+)-T-cell-dependent protective immunity against T. cruzi infection

<i>T. cruzi</i> derived GIPLs are TLR4 agonists and promote high levels of antigen-specific IgG2c antibodies as well as IFN-γ production by CD4⁺ T cells.

<p>(<b>A</b>) CHO cells control (TLR2⁻/TLR4⁻) or expressing TLR2 (TLR2⁺) or TLR4 (TLR4⁺) were either left untreated (solid gray) or exposed to 100 µg/ml of GIPLs from <i>Trypanosoma cruzi</i> Tulahuen (GTH), Y strain (GY) (black line). MALP-2 (10 ηg/ml) and LPS (200 ηg/ml) were used as positive controls for activation of TLR2⁺ or TLR4⁺, respectively. (<b>B</b>) TLR4⁺ cells were activated with different preparations of GIPLs in the presence of polymyxin B (PB). LPS was used as control. (<b>C</b>) OVA specific immune responses induced by immunization with TLR2 or TLR4 agonists associated with OVA absorbed in alum. Mice were immunized with three doses on days 0, 14 and 28. The production of total IgG, IgG1 and IgG2c were assessed by ELISA using the sera from immunized mice, at day 9 after the second boost. (<b>D</b>) To assess the levels of IFN-γ production by T lymphocytes from vaccinated mice, splenocytes were collected 21 days after the third immunization dose and stimulated with either CD4⁺ T or CD8⁺ T cell epitopes from OVA. The results are representative of two independent experiments yielding similar results. Asterisks indicate that differences were statistically significant, when comparing T cell response from mice receiving different vaccine formulations.</p