Long-Lasting Antibody and CD8+ Memory T Cell Responses Induced by N-Tc52/TSKb20 Vaccination upon Trypanosoma cruzi Antigen Re-Encounter.

Background: Chagas disease, caused by Trypanosoma cruzi, remains a major public health problem in Latin America and an emerging concern worldwide. Current chemotherapies show limited efficacy during chronic infection, and no licensed vaccine is currently available. We previously developed the chimeric antigen N-Tc52/TSKb20 as a vaccine candidate against T. cruzi infection. In a murine model, this vaccine induced robust antigen-specific immune response associated with protection shortly after vaccination. Objectives: Here, we investigated the long-term persistence and effector functions of the immune responses elicited by this vaccine candidate. Methods: Both female and male C57BL/6 mice were immunized with three doses of N-Tc52/TSKb20 formulated with QuilA adjuvant. Serum samples collected 170 days post-immunization were analyzed for antigen-specific antibodies by ELISA and for trypanolytic activity against cell-derived trypomastigotes using an in vitro functional assay. Cellular immune responses were evaluated by measuring cytokine production, T cell activation, and memory T cell responses following in vitro re-stimulation with the vaccine antigen or T. cruzi antigens. Results: N-Tc52/TSKb20 vaccination induced a sustained antigen-specific humoral response, characterized by long-lasting IgG2c antibodies and functional activity persisting for up to 170 days post-immunization. In parallel, vaccination promoted long-term activation of antigen-specific CD8+ T cells and production of TNF-α and IFN-γ upon antigen re-encounter. A sex-dependent tendency was observed for IL-10, with increased production in vaccinated female mice. Moreover, vaccinated animals exhibited increased frequencies of central and effector memory CD4+ and CD8+ T cells in response to T. cruzi antigens, with a predominant contribution of CD8+ T cells, indicating the establishment of parasite-specific T cell memory. Conclusions: Together, these findings demonstrate that vaccination with N-Tc52/TSKb20 induces a long-lasting Th1-biased immune response characterized by trypanolytic antibodies, functional and durable T cell responses, and parasite-specific memory T cells. This immunological profile supports the potential of N-Tc52/TSKb20 as a promising vaccine candidate for Chagas disease and highlights its capacity to elicit immune mechanisms that have been associated with protection against T. cruzi infection.