Prime and boost immunization protocol employing novel vaccine candidates for Chagas’disease.

ZILIANI, M; SANCHEZ DO; TEKIEL, V

Foz do Iguacu

Congreso; XXVII Annual Meeting of the Brazilian Society of protozoology - XXXVIII Annual Meeting on Basic Research in Chagas' disease; 2011

Sociedad Brasilera de Protozoologia

[Cod. Trabalho: 71]PÔSTERImunologia - ImmunologyPRIME AND BOOST IMMUNIZATION PROTOCOL EMPLOYING NOVEL VACCINE CANDIDATES FOR CHAGAS’ DISEASE.ZILIANI, M.; SÁNCHEZ, D.O.; TEKIEL, V..INSTITUTO DE INVESTIGACIONES BIOTECNOLÓGICAS, UNSAM, BUENOS AIRES - ARGENTINA.Agência Financiadora: Agencia Nacional de promoción Científica y Tecnológica (ANPCyT): PICT-488.Palavras-chave: vaccine, T. cruzi;prime&boost;recombinant protein expressionResumoWe have previously screened an epimastigote-subtracted trypomastigote cDNA expression library of Trypanosoma cruzi by genetic immunization and challenge. This approach allowed us to identify 22 novel vaccine candidates. Sequence analysis showed that most of them are fragments of hypothetical proteins or unannotated T. cruzi open reading frames (ORFs). As the candidates identified were gene fragments that produce peptides ranging 4-21 kDa, we expanded the original sequences to include a greater number of T and B epitopes and to increase their immunogenicity.For this study, we selected the T. cruzi annotated proteins (n=4) and the ORFs whose products have high density of T and B epitopes and/or trans-membrane domains (n=5). They were amplified from CL Brener DNA, cloned and expressed into pQE or pGEX expression vectors and purified by affinity chromatography. As most of the recombinant proteins were insoluble in all the assayed protocols, they were purified from inclusion bodies. Mice (n=6) were immunized with 2 doses of mammalian-expression plasmid DNA carrying the gene fragments of T. cruzi genes/ORFs, administered with a plasmid coding for murine GM-CSF as adjuvant (control mice received empty plasmids plus GM-CSF; n=6). Animals were boosted with a 3rd dose of recombinant proteins coupled to aluminum hydroxide (control mice: GST + his-tagged protein + aluminum hydroxide). Sera were collected to analyze the humoral response. Animals were challenged with a lethal dose of trypomastigotes of the RA strain. Levels of bloodstream trypomastigotes were lower in the vaccinated mice than in control group. This data suggests that the immunization protocol employed allows partial control and/or delay of parasitemia.