In silico identification and ex vivo evaluation of Toxoplasma gondii peptides restricted to HLA-A*02, HLA-A*24 and HLA-B*35 alleles in human PBMC from a Colombian population

Toxoplasma gondii infects approximately 30% of the population, and there is currently no approved vaccine. Identifying immunogenic peptides with high affinity to different HLA molecules is a promising vaccine strategy. This study used an in silico approach using artificial neural networks to identify T. gondii peptides restricted to HLA-A*02, HLA-A*24, and HLA-B*35 alleles. Proteomes from seven T. gondii strains and transcriptomic data of overexpressed genes from T. gondii-RH in human PBMC were also used. Parasite protein sequences were analyzed with R 'Epitope Prediction' library. Peptide candidates were evaluated in the artificial neural networks based on the probabilities of output neurons (p > 0.5). The IFN-γ responses in PBMC from T. gondii seronegative and seropositive individuals were evaluated by ELISpot. Peptides with higher IFN-γ induction were evaluated to identify cytotoxic response in CD8⁺ T cells (CD107a). In silico analysis identified 36 peptides from T. gondii proteins with predicted affinity to HLA-A*02, A*24, and B*35 alleles. Experiments with PBMCs revealed that a peptide restricted to HLA-A02 (P1: FLFAWITYV) induced a significant increase in IFN-γ-producing cells (p = 0.004). For HLA-A24, a peptide (P8: VFAFAFAFFLI) also induced a significant IFN-γ response (p = 0.004), while for the HLA-B*35 allele, the P6 peptide (YPIAPSFAM) induced a response that differed significantly from the control (p = 0.05). These peptides induced also a significant percentage of central memory CD8 + T cells expressing the degranulation marker CD107a (p < 0.05). Finally, we identified three T. gondii peptides that induced IFN-γ response, and a cytotoxic response measured by CD107a expression on CD45RAneg-CD8 cells. These peptides could be considered part of a multi-epitope vaccine against toxoplasmosis in humans.