Evaluation of the safety and immunogenicity of a peptide vaccine against canine leishmaniosis: a double-blind, multicenter, controlled clinical trial in dogs

Current vaccines for canine leishmaniosis (CanL) provide limited protection, underscoring the need for improved immunization strategies. Multi-epitope peptide vaccine delivered <i>via</i> nanoparticle systems represents a promising alternative but remains underexplored in canine clinical trials. Here, we report the results of a double-blind clinical trial (499/ECV) evaluating the safety and immunogenicity of HisDTC, a peptide vaccine targeting <i>Leishmania infantum</i>, encapsulated in poly(lactic-co-glycolic acid) PLGA nanoparticles and adjuvanted with Toll-like receptor 2 (TLR2) and TLR3 ligands. Forty healthy dogs were immunized with different vaccine formulations and monitored over 12 months. Immune responses were assessed by flow cytometry, ELISA, and <i>in vitro</i> macrophage infection assays, while safety was evaluated through clinical follow-up. Vaccination with HisDTC elicited a protective cellular response, including sustained IFN-γ production by CD4⁺ and CD8⁺ T cells, an IgG2a-skewed humoral response, and expansion of CD4⁺CD8α⁺ double-positive memory T cells. Notably, only HisDTC-vaccinated dogs exhibited a >80% reduction in <i>in vitro</i> macrophage infection, with protective effects persisting for up to 8 months post-immunization. Importantly, the formulation was well tolerated, with no adverse effects reported. These findings demonstrate that HisDTC delivered <i>via</i> PLGA nanoparticles induces durable, protective immunity against <i>L. infantum</i> in dogs and supports its further evaluation under natural exposure conditions.