Identification of T. gondii epitopes,adjuvants, and host genetic factors that influence protection of mice and humans

Toxoplasma gondii is an intracellular parasite that causes severe neurologic and ocular disease in immune-compromised and congenitally infected individuals. There is no vaccine protective against human toxoplasmosis. Herein, immunization of L^d mice with HF10 (HPGSVNEFDF) with palmitic acid moieties or a monophosphoryl lipid A derivative elicited potent IFN-γ production from L^d-restricted CD8⁺ T cells in vitro and protected mice. CD8⁺ T cell peptide epitopes from T. gondii dense granule proteins GRA 3, 6, 7, and Sag 1, immunogenic in humans for HLA-A02⁺, HLA-A03⁺, and HLA-B07⁺ cells were identified. Since peptide repertoire presented by MHC class I molecules to CD8⁺ T cells is shaped by endoplasmic reticulum-associated aminopeptidase (ERAAP), polymorphisms in the human ERAAP gene ERAP1 were studied and associate with susceptibility to human congenital toxoplasmosis (p < 0.05). These results have important implications for vaccine development.     

Identification of two novel immunodominant UreB CD4 T cell epitopes in Helicobacter pylori infected subjects

An epitope-based vaccine is a promising option for treating Helicobacter pylori (H. pylori) infection. Epitope mapping is the first step in designing an epitope-based vaccine. A pivotal role of CD4⁺ T cells in protection against H. pylori has been accepted, but few Th epitopes have been identified. In this study, two novel UreB CD4⁺ T cell epitopes were identified using PBMCs obtained from two H. pylori infected subjects. We determined the restriction molecules by antibody blocking and used various Epstein–Barr virus-transformed B lymphocyte cell lines (BLCLs) with different HLA alleles as APCs to present peptides to CD4⁺ T cells. These epitopes were DRB1*1404-restricted UreB_373–385 and DRB1*0803-restricted UreB_438–452. The T cells specific to these epitopes not only recognized autologous DCs loaded with recombinant UreB but also those pulsed with H. pylori whole cell lysates, suggesting that these epitope peptides are naturally processed. These epitopes have important value for designing an effective H. pylori vaccine.     

CD4 T-cell responses among adults and young children in response to Streptococcus pneumoniae and Haemophilus influenzae vaccine candidate protein antigens

We characterized cytokine profiles of CD4⁺ T-helper (h) cells in adults and young children to ascertain if responses occur to next-generation candidate vaccine antigens PspA, PcpA, PhtD, PhtE, Ply, LytB of Streptococcus pneumonia (Spn) and protein D and OMP26 of non-typeable Haemophilus influenzae (NTHi). Adults had vaccine antigen-specific Th1 and Th2 cells responsive to all antigens evaluated whereas young children had significant numbers of vaccine antigen-specific CD4⁺ T cells producing IL-2, (p = 0.004). Vaccine antigen-specific CD4⁺ T-cell populations in adults were largely of effector (T_EM) and/or central memory (T_CM) phenotypes as defined by CD45RA⁻CCR7⁺ or CD45RA⁻CCR7⁻ respectively; however among young children antigen-specific IL-2 producing CD4⁺ T cells demonstrated CD45RA⁺ expression (non-memory cells). We conclude that adults have circulating memory CD4⁺ T cells (CD45RA⁻) that can be stimulated by all the tested Spn and NTHi protein vaccine candidate antigens, whereas young children have a more limited response.     

Ovalbumin lipid core peptide vaccines and their CD4 and CD8 T cell responses

The lipid core peptide (LCP) system has successfully been used in development of peptide-based vaccines against cancer and infectious diseases (such as group A streptococcal infection). CD8⁺ T cells are important targets for vaccines, however developing a vaccine that activates long-lasting immunity has proven challenging. The ability of LCP vaccines to activate antigen-specific CD8⁺ and/or CD4⁺ T cell responses was tested using compounds that contained two or four copies of OVA_257–264 and/or OVA_323–339 peptides conjugated to LCP, which are recognised by OTI (CD8⁺ specific) and OTII (CD4⁺ specific) T cells, respectively. The LCP–ovalbumin vaccines developed in this study were synthesised in 30% yields and showed no significant haemolytic effect on red blood cells (below 4% haemolysis when tested with compounds at up to 100 μM concentrations). Promising in vivo data in mice suggested that this LCP–ovalbumin vaccine system could act as a novel and potent vehicle for the stimulation of robust antigen-specific CD8⁺ T cell responses.     

A gold glyco-nanoparticle carrying a listeriolysin O peptide and formulated with Advax™ delta inulin adjuvant induces robust T-cell protection against listeria infection

In the search for an effective vaccine against the human pathogen, Listeria monocytogenes (Listeria), gold glyconanoparticles (GNP) loaded with a listeriolysin O peptide LLO_91–99 (GNP-LLO) were used to immunise mice, initially using a dendritic cell (DC) vaccine approach, but subsequently using a standard parenteral immunisation approach. To enhance vaccine immunogenicity a novel polysaccharide adjuvant based on delta inulin (Advax™) was also co-formulated with the GNP vaccine. Confirming previous results, DC loaded in vitro with GNP-LLO provided better protection against listeriosis than DC loaded in vitro using free LLO peptide. The immunogenicity of GNP-LLO loaded DC vaccines was further increased by addition of Advax™ adjuvant. However, as DC vaccines are expensive and impracticable for prophylactic use, we next asked whether the same GNP-LLO antigen could be used to directly target DC in vivo. Immunisation of mice with GNP-LLO plus Advax™ adjuvant induced LLO-specific T-cell immunity and protection against Listeria challenge. Protection correlated with an increased frequency of splenic CD4⁺ and CD8⁺ T cells, NK cells and CD8α⁺ DC, and Th1 cytokine production (IL-12, IFN-γ, TNF-α, and MCP-1), post-challenge. Enhanced T-cell epitope recruitment post-challenge was seen in the groups that received Advax™ adjuvant. Immunisation with GNP-LLO_91–99 plus Advax™ adjuvant provided equally robust Listeria protection as the best DC vaccine strategy but without the complexity and cost, making this a highly promising strategy for development of a prophylactic vaccine against listeriosis.     

Effectiveness of a novel immunogenic nanoparticle platform for Toxoplasma peptide vaccine in HLA transgenic mice

We created and produced a novel self-assembling nanoparticle platform for delivery of peptide epitopes that induces CD8⁺ and CD4⁺T cells that are protective against Toxoplasma gondii infection. These self-assembling polypeptide nanoparticles (SAPNs) are composed of linear peptide (LP) monomers which contain two coiled-coil oligomerization domains, the dense granule 7 (GRA7_20–28 LPQFATAAT) peptide and a universal CD4⁺T cell epitope (derived from PADRE). Purified LPs assemble into nanoparticles with icosahedral symmetry, similar to the capsids of small viruses. These particles were evaluated for their efficacy in eliciting IFN-γ by splenocytes of HLA-B*0702 transgenic mice and for their ability to protect against subsequent T. gondii challenge. This work demonstrates the feasibility of using this platform approach with a CD8⁺ epitope that binds HLA-B7 and tests the biological activity of potentially protective peptides restricted by human major histocompatibility complex (HLA) class I molecules in HLA transgenic mice.     

Anti-glioma effect of intracranial vaccination with tumor cell lysate plus flagellin in mice

The adjuvant effects of flagellin on regulation of immune response have been proved; whether flagellin could assist tumor cell lysate (TCL) to enhance anti-glioma immunity remains to be investigated. This study tests a hypothesis that therapeuticly intracranial administration with flagellin plus TCL enhances the effects of specific immunotherapy on glioma in mice. In this study, GL261 cells were transferred into C57BL/6 mice and the GL261-bearing mice were subcutaneously or intracranially inoculated with flagellin plus TCL, flagellin, TCL or saline. Our results showed that prophylacticly subcutaneous administration with TCL and flagellin could induce potent cytotoxic T lymphocyte (CTL) and prolong the survival of GL261-bearing mice significantly, but therapeuticly subcutaneous administration failed to. However, therapeuticly intracranial administration of TCL plus flagellin could prolong the survival. Moreover, intracranial administration of flagellin could recruit CD4⁺ T cells and CD8⁺ T cells to brain tissues, induce proliferation of natural killer (NK) cells, CD4⁺ T cells and CD8⁺ T cells in peripheral blood mononuclear cells and induce to splenomegaly. The results suggested that flagellin could be acted as an efficient adjuvant for TCL based vaccine.     

Novel influenza virus vectors expressing Brucella L7/L12 or Omp16 proteins in cattle induced a strong T-cell immune response,as well as high protectiveness against B. abortus infection

This paper presents the results of a study of the immunogenicity and protectiveness of new candidate vector vaccine against Brucella abortus – a bivalent vaccine formulation consisting of a mixture of recombinant influenza A subtype H5N1 or H1N1 (viral constructs vaccine formulation) viruses expressing Brucella ribosomal protein L7/L12 and Omp16, in cattle. To increase the effectiveness of the candidate vaccine, adjuvants such as Montanide Gel01 or chitosan were included in its composition. Immunization of cattle (heifers aged 1–1.5 years, 5 animals per group) with the viral constructs vaccine formulation only, or its combination with adjuvants Montanide Gel01 or chitosan, was conducted via the conjunctival method using cross prime (influenza virus subtype H5N1) and booster (influenza virus subtype H1N1) vaccination schedules at an interval of 28 days. Vaccine candidates were evaluated in comparison with the positive (B. abortus S19) and negative (PBS) controls. The viral constructs vaccine formulations, particularly in combination with Montanide Gel01 adjuvant promoted formation of IgG antibodies (with a predominance of antibodies of isotype IgG2a) against Brucella L7/L12 and Omp16 proteins in ELISA. Moreover, these vaccines in cattle induced a strong antigen-specific T-cell immune response, as indicated by a high number of CD4⁺ and CD8⁺ cells, as well as the concentration of IFN-γ, and most importantly provided a high level of protectiveness comparable to the commercial B. abortus S19 vaccine and superior to the B. abortus S19 vaccine in combination with Montanide Gel01 adjuvant. Based on these findings, we recommended the bivalent vaccine formulation containing the adjuvant Montanide Gel01 for practical use in cattle.     

Flagellin is a Th1 polarizing factor for human CD4+ T cells and induces protection in a murine neonatal vaccination model of rotavirus infection

Neonates have an increased susceptibility to infections, particularly those caused by intracellular pathogens, leading to high morbidity and mortality rates. This is partly because of a poor response of neonatal CD4⁺ T cells, leading to deficient antibody production and a low production of IFN-γ, resulting in deficient elimination of intracellular pathogens. The poor memory response of human neonates has underpinned the need for improving vaccine formulations. Molecular adjuvants that improve the response of neonatal lymphocytes, such as the ligands of toll-like receptors (TLRs), are attractive candidates. Among them, flagellin, the TLR5 ligand, is effective at very low doses; prior immunity to flagellin does not impair its adjuvant activity. Human CD4⁺ and CD8⁺ T cells express TLR5. We found that flagellin induces the expression of IFN-γ, IL-1β and IL-12 in mononuclear cells from human neonate and adult donors. When human naïve CD4⁺ T cells were activated in the presence of flagellin, there was high level of expression of IFN-γ in both neonates and adults. Furthermore, flagellin induced IFN-γ production in Th1 cells obtained from adult donors; in the Th2 population, it inhibited IL-4 cytokine production. Flagellin also promoted expression of the IFN-γ receptor in naive CD4⁺ T cells from neonates and adults. To test the adjuvant capacity of flagellin in vivo, we used a murine neonate vaccination model for infection with rotavirus, a pathogen responsible for severe diarrhea in young infants. Using the conserved VP6 antigen, we observed an 80% protection against rotavirus infection in the presence of flagellin, but only in those mice previously primed in the neonatal period. Our data suggest that flagellin could be an attractive adjuvant for achieving a Th1 response.     

Immune response and protective efficacy of live attenuated Salmonella vaccine expressing antigens of Mycobacterium avium subsp. paratuberculosis against challenge in mice

Mycobacterium avium subsp. paratuberculosis (MAP) causes chronic granulomatous enteritis in ruminants that leads to diarrhea and eventually death. Existing vaccines have proven useful in limiting disease progression but have not been effective in preventing infection. To address this problem we constructed an attenuated Salmonella (ΔyejE; ΔssaV) strain harboring a plasmid that expressed a fusion protein comprised of the Salmonella Type III secretion system (T3SS) effector SopE and MAP antigens (85A, 85B, SOD, 74F) and evaluated its potential as vaccine candidate against MAP infection in mice. Of various SopE-MAP fusion proteins analyzed, only SopE104-Ag85A C-terminal^202–347-SOD N-terminal^1–72-Ag85B C-terminal^173–330 and SopE104-74F^{1–148 + 669–786}were successfully expressed and secreted into culture media as revealed by western blot analysis. Mice immunized with attenuated Salmonella (ΔyejE; ΔssaV) harboring the SopE104-Ag85A C-terminal^202–347-SOD N-terminal^1–72-Ag85B C-terminal^173–330 and SopE104-74F^{1–148 + 669–786}plasmid generated a potent and long lasting Th1 response characterized by production of IFN-γ. The cytokine profile varied at various time points after immunization and challenge, which showed down regulation of Th2 cytokines (IL-4, IL-10) and up-regulation of proinflammatory cytokines (IL-12 and IL-17). Further, the immune response correlated with protection as revealed by reduced bacterial load and improved histopathology of spleen and liver, which showed fewer granulomas and lower numbers of acid-fast bacilli as compared to PBS controls. Interestingly, vaccination with antigens mixed with Ribi adjuvant (Agmix + Ribi) imparted better protection than the attenuated salmonella vectored vaccine. Thus, priming with a live recombinant Salmonella strain that secretes MAP antigens represents a promising approach that could lead to development of an efficacious and cost effective vaccine for Johne's disease.     

PED subunit vaccine based on COE domain replacement of flagellin domain D3 improved specific humoral and mucosal immunity in mice

Porcine epidemic diarrhea (PED) is an important re-emergent infectious disease and inflicts huge economic losses to the swine industry worldwide. To meet the pressing need of developing a safe and cost-efficient PED maternal vaccine, we generated three PED subunit vaccine candidates, using recombined Salmonella flagellin (rSF) as a mucosal molecular adjuvant. Domain D3 in rSF was replaced with COE domain of PEDV to generate rSF-COE-3D. COE fused to the flanking C′/N′ terminal of rSF yielded rSF-COE-C and rSF-COE-N. As a result, rSF-COE-3D could significantly improve COE specific antibody production including serum IgG, serum IgA, mucosal IgA and PEDV neutralizing antibody. Furthermore, rSF-COE-3D elicited more CD3⁺CD8⁺ T cell and cytokine production of IFN-γ and IL-4 in mouse splenocytes. In summary, our data showed that rSF-COE-3D could improve specific humoral and mucosal immunity in mice, thus suggesting that rSF-COE-3D could be applied as a novel efficient maternal PED vaccine.     

An IL-15 adjuvant enhances the efficacy of a combined DNA vaccine against Brucella by increasing the CD8 cytotoxic T cell response

We investigated whether a combined DNA vaccine delivered together with the IL-15 gene (DNA-IL-15(+)) enhanced the immune response against Brucella abortus in mice. Mice vaccinated with DNA-IL-15(+) developed a robust humoral response; Brucella-specific antibodies exhibited a dominance of immunoglobulin G2a (IgG2a) over IgG1. Splenocytes from DNA-IL-15(+)-vaccinated mice induced significantly higher levels of IFN-γ (P < 0.01) and CD8⁺ T cell response (P < 0.01), suggesting induction of a T-helper-1-dominated immune response. In a specific cytotoxic-T-lymphocyte activity assay, DNA-IL-15(+) immunization elicited mainly CD8⁺ T cells, which mediate cytotoxicity, but also CD4⁺ T cells. In vivo depletion of T cell subsets showed that the DNA-IL-15(+)-induced protection against Brucella infection is mediated predominantly by CD8⁺ T cells, although CD4⁺ T cells also contribute. These data indicate that plasmid-delivered IL-15 increases the efficacy of the Brucella DNA vaccine.     

Cationic lipid/DNA complexes (JVRS-100) combined with influenza vaccine (Fluzone) increases antibody response,cellular immunity,and antigenically drifted protection

Safe and effective adjuvants for influenza vaccines that could increase both the levels of neutralizing antibody, including against drifted viral subtypes, and T-cell immunity would be a major advance in vaccine design. The JVRS-100 adjuvant, consisting of DOTIM/cholesterol cationic liposome–DNA complexes, is particularly promising for vaccines that require induction of high levels of antibody and T-cell immunity, including CD8⁺ cytotoxic T lymphocytes (CTL). Inclusion of protein antigens with JVRS-100 results in the induction of enhanced humoral and cell-mediated (i.e., CD4⁺ and CD8⁺ T cells) immune responses. The JVRS-100 adjuvant combined with a split trivalent influenza vaccine (Fluzone^®—sanofi pasteur) elicited increased antibody and T-cell responses in mice and non-human primates compared to vaccination with Fluzone^® alone. Mice vaccinated with JVRS-100–Fluzone^® and challenged with antigenically drifted strains of H1N1 (PR/8/34) and influenza B (B/Lee/40) viruses had higher grade protection, as measured by attenuation of weight loss and increased survival, compared to recipients of unadjuvanted vaccine. The results indicate that the JVRS-100 adjuvant substantially increases immunogenicity and protection from drifted-strain challenge using an existing influenza vaccine.     

A synthetic peptide from Trypanosoma cruzi mucin-like associated surface protein as candidate for a vaccine against Chagas disease

Chagas disease, caused by Trypanosoma cruzi, is responsible for producing significant morbidity and mortality throughout Latin America. The disease has recently become a public health concern to nonendemic regions like the U.S. and Europe. Currently there are no fully effective drugs or vaccine available to treat the disease. The mucin-associated surface proteins (MASPs) are glycosylphosphatidylinositol (GPI)-anchored glycoproteins encoded by a multigene family with hundreds of members. MASPs are among the most abundant antigens found on the surface of the infective trypomastigote stage of T. cruzi, thus representing an attractive target for vaccine development. Here we used immunoinformatics to select a 20-mer peptide with several predicted overlapping B-cell, MHC-I, and MHC-II epitopes, from a MASP family member expressed on mammal-dwelling stages of T. cruzi. The synthetic MASP peptide conjugated to keyhole limpet hemocyanin (MASPpep-KLH) was tested in presence or not of an adjuvant (alum, Al) as a vaccine candidate in the C3H/HeNsd murine model of T. cruzi infection. In considerable contrast to the control groups receiving placebo, Al, or KLH alone or the group immunized with MASPpep-KLH/Al, the group immunized with MASPpep-KLH showed 86% survival rate after challenge with a highly lethal dose of trypomastigotes. As evaluated by quantitative real-time polymerase chain reaction, MASPpep-KLH-immunized animals had much lower parasite load in the heart, liver, and spleen than control animals. Moreover, protected animals produced trypanolytic, protective antibodies, and a cytokine profile conducive to resistance against parasite infection. Finally, in vivo depletion of either CD4⁺ or CD8⁺ T cells indicated that the latter are critical for protection in mice immunized with MASPpep-KLH. In summary, this new peptide-based vaccine with overlapping B- and T-cell epitopes is able to control T. cruzi infection in mice by priming both humoral and cellular immunity.     

Effective CD8 T cell priming and tumor protection by enterotoxin B subunit-conjugated peptides targeted to dendritic cells

In our previous studies we have shown that bacterial enterotoxin B subunits are effective vehicles to deliver antigen into the MHC class I processing route. Here we have used the non-toxic Escherichia coli heat labile enterotoxin B subunit (EtxB) conjugated to OVA peptide (EtxB–peptide) to address the impact on induction of specific CD8⁺ T cells in vivo. Although incubation of DCs with these EtxB–peptide conjugates as such did not induce DC maturation in vitro MHC class I antigen presentation was much more efficient as compared to peptide alone. Antigen presentation was further enhanced upon DC maturation with the TLR-4 ligand LPS. Injection of matured DCs incubated with EtxB–peptide conjugates lead to strong induction of OVA-specific CD8⁺ T lymphocytes and fully prevented the outgrowth of lethal B16 melanoma in wild type mice. Our data demonstrate that bacterial non-toxic B subunit–peptide conjugates are potent vaccine vehicles for induction of protective CD8⁺ T cell responses.     

Improving the Th1 cellular efficacy of the lead Yersinia pestis rF1-V subunit vaccine using SA-4-1BBL as a novel adjuvant

The lead candidate plague subunit vaccine is the recombinant fusion protein rF1-V adjuvanted with alum. While alum generates Th₂ regulated robust humoral responses, immune protection against Yersinia pestis has been shown to also involve Th₁ driven cellular responses. Therefore, the rF1-V-based subunit vaccine may benefit from an adjuvant system that generates a mixed Th₁ and humoral immune response. We herein assessed the efficacy of a novel SA-4-1BBL costimulatory molecule as a Th₁ adjuvant to improve cellular responses generated by the rF1-V vaccine. SA-4-1BBL as a single adjuvant had better efficacy than alum in generating CD4⁺ and CD8⁺ T cells producing TNFα and IFNγ, signature cytokines for Th₁ responses. The combination of SA-4-1BBL with alum further increased this Th₁ response as compared with the individual adjuvants. Analysis of the humoral response revealed that SA-4-1BBL as a single adjuvant did not generate a significant Ab response against rF1-V, and SA-4-1BBL in combination with alum did not improve Ab titers. However, the combined adjuvants significantly increased the ratio of Th₁ regulated IgG_2c in C57BL/6 mice to the Th₂ regulated IgG₁. Finally, a single vaccination with rF1-V adjuvanted with SA-4-1BBL + alum had better protective efficacy than vaccines containing individual adjuvants. Taken together, these results demonstrate that SA-4-1BBL improves the protective efficacy of the alum adjuvanted lead rF1-V subunit vaccine by generating a more balanced Th₁ cellular and humoral immune response. As such, this adjuvant platform may prove efficacious not only for the rF1-V vaccine but also against other infections that require both cellular and humoral immune responses for protection.     

In vitro stimulation of human influenza-specific CD8 T cells by dendritic cells pulsed with an influenza virus-like particle (VLP) vaccine

The purpose of this in vitro study was to determine if a virus-like particle (VLP) influenza vaccine stimulated human CD8⁺ T cells in a dendritic cell (DC): T cell co-culture system. VLP-pulsed DCs were co-cultured with autologous CD8⁺ T cells from five donors. Functional CD8⁺ T cells were detected via cell surface and intracellular cytokine staining. T cells from four of the five donors showed ≥2-fold increase over background in the % activated CD8⁺ cells. These results indicate that the influenza VLP vaccine can stimulate CD8⁺ T cells via DC antigen presentation, likely through the MHC-I pathway, thus broadening the immunological response induced by this promising influenza vaccine.     

Evaluation of hepatic changes and local and systemic immune responses in goats immunized with recombinant Peroxiredoxin (Prx) and challenged with Fasciola hepatica

Protection against Fasciola hepatica in goats immunized with Peroxiredoxin (Prx) was assessed. The experimental trial consisted of three groups of seven animals; group 1 were unimmunized and uninfected, group 2 were immunized with adjuvant only and group 3 were immunized with recombinant Prx in adjuvant (immunized and infected). Immunization with Prx in Quil A adjuvant, group 3, induced a reduction in fluke burden of 33.04% when compared to adjuvant control, group 2, although this difference was not significant. The hepatic gross and microscopical morphometric study revealed lower damage in the Prx-immunized compared to group 2 (p < 0.05). Furthermore, immunohistochemical studies revealed that the Prx-immunized group exhibited reduced infiltration of CD4⁺, CD8⁺, IFN-γ⁺ and TCR⁺ (p < 0.05); and CD2⁺ and IL-4⁺ (p < 0.001) in hepatic lesions. Levels of anti-Prx serum IgG in group 3 showed a significant increase at the 4th week after challenge infection compared with group 2 (p < 0.0001). This is the first report of ruminant immunization with recombinant Prx of F. hepatica. The study shows that this vaccine significantly reduces hepatic damage and encourages further studies to improve the vaccine efficacy.     

Delivery of antigenic candidates by a DNA/MVA heterologous approach elicits effector CD8T cell mediated immunity against Trypanosoma cruzi

In this study, we have characterized the immune mechanisms elicited by antigenic candidates, TcG2 and TcG4, delivered by a DNA-prime/MVA-boost approach, and evaluated the host responses to Trypanosoma cruzi infection in C57BL/6 mice. Immunization of mice with antigenic candidates elicited antigen-specific, high-avidity, trypanolytic antibody response (IgG2b > IgG1) and CD8⁺T cells that exhibited type-1 cytolytic effector (CD8⁺CD107a⁺IFN-γ⁺Perforin⁺) phenotype. The extent of TcG2-dependent type 1 B and T cell immunity was higher than that noted in TcG4-immunized mice, and expanded accordingly in response to challenge infection with T. cruzi. The progression of chronic phase in immunized mice was associated with persistence of IgGs, 55–90% reduction in the frequency of proinflammatory (IFN-γ⁺ or TNF-α⁺) CD8⁺T cells, and an increase or emergence of immunoregulatory (IL-10⁺) CD4/CD8 T cells. The tissue parasitism, infiltration of inflammatory infiltrate, parasite persistence, and fibrosis were decreased by 82–92% in heart and skeletal muscle of immunized/chronically infected mice. Control mice exhibited a significantly low antibody response, consistent activation of effector CD8⁺T cells dominated by pro-inflammatory phenotype and mixed cytokine profile (IFN-γ + TNF-α > IL-4 + IL-10), parasite persistence and pathologic damage in chagasic hearts. We conclude that delivery of TcG2 or TcG4 by DNA-rMVA approach elicits effective antibody and CD8⁺T cell mediated immunity against T. cruzi and Chagas disease. The emergence of type 2 cytokine and T cell response in chronic phase was indicative of prevention of clinical disease.     

CD11a and CD49d enhance the detection of antigen-specific T cells following human vaccination

BackgroundDetermining the efficacy of human vaccines that induce antigen-specific protective CD4 T cell responses against pathogens can be particularly challenging to evaluate. Surface expression of CD11a and CD49d has been shown to identify antigen-specific CD4 T cells against viral pathogens in mice. We hypothesized that CD11a and CD49d would also serve as markers of human antigen-specific T cells responding to vaccination.MethodsA phase I vaccine trial enabled us to evaluate a novel gating strategy based on surface expression of CD11a and CD49d as a means of detecting antigen-specific, cytokine producing CD4 and CD8 T cells induced after vaccination of naïve individuals against leishmaniasis. Three study groups received LEISH-F3 recombinant protein combined with either squalene oil-in-water emulsion (SE) alone, SE with the synthetic TLR-4 ligand glucopyranosyl lipid adjuvant (GLA-SE), or SE with Salmonella minnesota-derived monophosphoryl lipid A (MPL-SE). Individuals were given 3 vaccine doses, on days 0, 28 and 168.ResultsStarting after the first vaccine dose, the frequency of both CD11a^hiCD49d⁺ CD4 and CD11a^hiCD49d⁺ CD8 T cells significantly increased over time throughout the 24-week trial. To confirm the role of CD11a^hiCD49d⁺ expression in the identification of the antigen-specific T cells, cytokine production was measured following LEISH-F3 stimulation. All of the IFN-γ, TNF-α, and IL-2 producing cells were found within the CD11a^hiCD49d⁺ population.ConclusionsOur results suggest that the change in the frequency of CD11a^hiCD49d⁺ T cells can be used to track antigen-specific CD4 and CD8 T cell responses following T cell-targeted vaccination.