Vaccination with a CD4+ and CD8+ T-cell epitopes-based recombinant chimeric protein derived from Leishmania infantum proteins confers protective immunity against visceral leishmaniasis

Abstract

Vaccination seems to be the best approach to control visceral leishmaniasis (VL). Resistance against infection is based on the development of a Th1 immune response characterized by the production of interferons-γ (IFN-γ), interleukin-12 (IL-12), granulocyte-macrophage-colony-stimulating factor (GM-CSF), and tumor necrosis factor-α (TNF-α), among others. A number of antigens have been tested as potential targets against the disease; few of them are able to stimulate human immune cells. In the present study, 1 prediction of MHC class I and II molecules-specific epitopes in the amino acid sequences of 3 Leishmania proteins: 1 hypothetical, prohibitin, and small glutamine-rich tetratricopeptide repeat-containing proteins, was performed using bioinformatics tools, and a T-cell epitopes-based recombinant chimeric protein was constructed, synthetized and purified to be evaluated in invitro and in vivo experiments. The purified protein was tested regarding its immunogenicity in peripheral blood mononuclear cells (PBMCs) from healthy subjects and VL patients, as well as to its immunogenicity and protective efficacy in a murine model against Leishmania infantum infection. Results showed a Th1 response based on high IFN-γ and low IL-10 levels derived from in chimera-stimulated PBMCs in both healthy subjects and VL patients. In addition, chimera and/or saponin-immunized mice presented significantly lower parasite burden in distinct evaluated organs, when compared to the controls, besides higher levels of IFN-γ, IL-2, IL-12, and GM-CSF, and an IgG2a isotype-based humoral response. In addition, the CD4 and CD8 T-cell subtypes contributed to IFN-γ production in the protected animals. The results showed the immunogenicity in human cells and the protective efficacy against L. infantum in a murine model, and well indicate that this recombinant chimera can be considered as a promising strategy to be used against human disease.

An immunoinformatic approach driven by experimental proteomics: in silico design of a subunit candidate vaccine targeting secretory proteins of Leishmania donovani amastigotes

“…Nevertheless, the outcome can be considered consistent with the general trend of in vitro immune response (compared to SLA), with peptide 2 being more prominent IL-10 inducer compared to SLA as reported in [47]. In terms of cytokine induction potential, simulation outcome of our designed construct conformed more closely to that of peptide 1, which did not induce IL-10 level higher than that by SLA in vitro [46] (Fig. 2).…”

“…The structure was found thermodynamically stable and surface-soluble, while the core is hydrophobic, a favorable feature for antigen processing. Vaccine-specific, but not parasite protein-specific humoral response was predicted, and this can be used as a biomarker of vaccine efficacy [46,73] without eliciting a parasite-specific B cell response. Moreover, the construct structure showed a good binding affinity in previously reported binding cavity of TLR4 [74][75][76][77].…”

“…The capability to induce IFN-γ and IL-10 by the chimera was predicted by scanning in IFNepitope [44] and IL-10Pred module, respectively. Simulation of immune response based exclusively on the chimeric construct was performed in C-ImmSim [45] server, whereas two previously reported candidate Leishmania vaccine peptides [46,47] were used to evaluate whether C-ImmSim prediction corroborates to the dynamicity of antigenic constructs. For structural analysis, the tertiary structure of the construct was produced in I-TASSER [48] modeling server followed by refinement using YASARA [49] force-field and GalaxyRefine [50] web tools.…”

“…Since the hypothetical cytokine levels in simulated immune response represent only the outcome of algorithmically set dynamic cellular interactions for a defined time period after antigen priming [37], it was important to evaluate whether the simulation module can respond dynamically to different constructs [45]. Therefore, we simulated immune responses of two additional peptide vaccine candidates: peptide 1 (L. infantum derived fusion peptide [46]) and peptide 2 (L. donovani GP63 derived peptide [47]), which were experimentally found to exhibit varying cytokine response in comparison to soluble Leishmania antigen (SLA). Substantial difference was observed in terms of immunosuppressive IL-10 and TGF-β induction capacity between peptide 1 and peptide 2; however, determination of statistical significance was not possible in the simulation module.…”

An immunoinformatic approach driven by experimental proteomics: in silico design of a subunit candidate vaccine targeting secretory proteins of Leishmania donovani amastigotes

“…The capability to induce IFN-γ and IL-10 by the chimera was predicted by scanning in IFNepitope [44] and IL-10Pred module, respectively. Simulation of immune response based exclusively on the chimeric construct was performed in C-ImmSim [45] server, whereas two previously reported candidate Leishmania vaccine peptides [46,47] were used to evaluate whether C-ImmSim prediction corroborates to the dynamicity of antigenic constructs. For structural analysis, tertiary structure of the construct was produced in I-TASSER [48] modeling server followed by refinement using YASARA [49] force field and GalaxyRefine [50] web tool.…”