Immunization with an engineered mutant trans-sialidase highly protects mice from experimental Trypanosoma cruzi infection: a vaccine candidate.
ABSTRACT Chagas' disease is a major tropical disease for which a cure for chronic phase does not exist yet. Trypanosoma cruzi trans-sialidase (TS) seems to be involved in relevant processes such as infectivity, host survival and, very importantly, disease pathogenesis. In this study, we show that mice vaccinated with an engineered enzymatically deficient mutant TS containing the catalytic domain without the immunodominant SAPA (Shed Acute Phase Antigen) repeats, were highly protected against T. cruzi infection. Adult male BALB/c mice were immunized with mutant protein, purified from Pichia pastoris yeast, using three inoculations in Freund's adjuvant. All immunized mice were protected against challenge with a lethal dose of T. cruzi trypomastigotes. The protected immunized mice developed no clinical or tissue evidence of infection throughout the study. In contrast, 60-90% mortality and 100% occurrence of myocardial lesions were observed in the non-immunized counterparts. Titers of circulating antibody against TS did not correlate with protection, while anti-SAPA antibodies were coincident with disease severity. Further studies indicated that a single inoculation of mutant recombinant protein in Freund's complete adjuvant was not associated with blood or organic alterations, per se. Mutant TS vaccination seems to be a promising tool for immune intervention strategies in Chagas' disease, aimed at preventing T. cruzi-related heart tissue damage.
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Article: Carbohydrates in transplantation.[show abstract] [hide abstract]
ABSTRACT: Carbohydrate materials have become increasingly utilized in transplantation and cell/tissue engineering within the past year. This has been well documented in recent applications of immobilized or soluble alpha-galactosyl epitopes (i.e. oligosaccharides with a terminal Galalpha1-3Gal sequence) in preventing hyperacute rejection in pig-to-primate xenotransplantation. In addition, alpha-galactosyl polymers have been shown to exhibit much greater activity (up to 10(4) times) than alpha-galactosyl monomers in inhibiting the binding of anti-galactosyl antibodies to pig kidney epithelial cells and assisting in the prevention of cytotoxicity in human serum.Current Opinion in Chemical Biology 01/2000; 3(6):650-8. · 9.47 Impact Factor
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ABSTRACT: Protective immunity against lethal infection is developed when BALB/c or C57BL/6 mice are immunized with plasmids containing genes from the protozoan parasite Trypanosoma cruzi. However, genetic vaccination of the highly susceptible mouse strain A/Sn promoted limited survival after challenge. This observation questioned whether this type of vaccination would be appropriate for highly susceptible individuals. Here, we compared the protective efficacy and the immune response after individual or combined genetic vaccination of A/Sn mice with genes encoding trans-sialidase (TS) or the amastigote surface protein-2 (ASP-2). After challenge, a significant proportion of A/Sn mice immunized with either the asp-2 gene or simultaneously with asp-2 and ts genes, survived infection. In contrast, the vast majority of mice immunized with the ts gene or the vector alone died. Parasitological and histological studies performed in the surviving mice revealed that these mice harbored parasites; however, minimal inflammatory responses were seen in heart and striated muscle. We used this model to search for an in vitro correlation for protection. We found that protective immunity correlated with a higher secretion of interferon- by spleen cells on in vitro restimulation with ASP-2 and the presence of ASP-2-specific CD8 cells. Depletion of either CD4 or CD8 or both T-cell subpopulations prior to the challenge rendered the mice susceptible to infection demonstrating the critical contribution of both cell types in protective immunity. Our results reinforce the prophylactic potential of genetic vaccination with asp-2 and ts genes by describing protective immunity against lethal T. cruzi infection and chronic tissue pathology in a highly susceptible mouse strain.Human Gene Therapy 10/2004; 15(9):878-86. · 4.02 Impact Factor
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ABSTRACT: Therapeutic glycoprotein production in the widely used expression host Pichia pastoris is hampered by the differences in the protein-linked carbohydrate biosynthesis between this yeast and the target organisms such as man. A significant step towards the generation of human-compatible N-glycans in this organism is the conversion of the yeast-type high-mannose glycans to mammalian-type high-mannose and/or complex glycans. In this perspective, we have co-expressed an endoplasmic reticulum-targeted Trichoderma reesei 1,2-alpha-D-mannosidase with two glycoproteins: influenza virus haemagglutinin and Trypanosoma cruzi trans-sialidase. Analysis of the N-glycans of the two purified proteins showed a >85% decrease in the number of alpha-1,2-linked mannose residues. Moreover, the human-type high-mannose oligosaccharide Man(5)GlcNAc(2) was the major N-glycan of the glyco-engineered trans-sialidase, indicating that N-glycan engineering can be effectively accomplished in P. pastoris.FEBS Letters 09/2001; 503(2-3):173-8. · 3.58 Impact Factor