Caracterización in vitro de vacunas liposomales antiLeishmania basadas en plásmidos de ADN

Leishmaniasis is a disease prevalent worldwide, particularly affecting dogs and rats with a high incidence. Various vaccines exist for its prevention, especially recommended when traveling to endemic zones. Lipid-based nanoparticles have been used in vaccine development for other diseases, such as the several releases for COVID-19. These nanoparticles enable the encapsulation of both DNA and RNA sequences, triggering the necessary immunological response for disease prevention once inside the body. In this study, we encapsulated a modified Leishmania DNA plasmid within lipid nanoparticles. The original plasmid, pVAX1, was altered to create pVAX1::HisAK70-asd, which includes a continuous sequence of genes encoding seven different Leishmania antigens (H2A, H2B, H3, H4, A2, KMP11, and HSP70), followed by an E-Tag for detection. The lipid nanoparticles were synthesized using various lipids, including DOPC, S100, DPP+PEG 2000, and PC-3, to evaluate their effectiveness in encapsulating the plasmid and serving as vaccine adjuvants. DOTAP-Cl was used as the cationic lipid to neutralize the negative charges of the plasmid. The synthesis method used was extrusion, and their physicochemical properties were analyzed by measuring hydrodynamic size and surface charge. The morphology and size of the lipid nanoparticles were further evaluated using transmission electron microscopy. Subsequently, in vitro assays were performed to assess cytotoxicity and possible alterations in cellular and nuclear morphology in DH82 canine macrophage cells. The results demonstrated that both DOPC and S100 are promising candidates for further studies. In vitro cell viability assays, using controls of both DOPC and S100, showed that these lipid nanoparticles are non-toxic and can be used as adjuvants in Leishmania vaccines. It is important to note that the plasmid was only included in the DOPC nanoparticles. This study highlights the potential of DOPC liposomal vaccines in enhancing the efficacy of targeted antigen delivery to antigenpresenting cells, addressing the challenges in developing effective vaccines against Leishmaniasis.