Production of recombinant TSA-1 and evaluation of its potential for the immuno-therapeutic control of Trypanosoma cruzi infection in mice.

A therapeutic vaccine for human Chagas disease (American trypanosomiasis caused by Trypanosoma cruzi) is under development based on the success of vaccinating mice with DNA constructs expressing the antigens Tc24 and Tc-TSA-1. However, because DNA and nucleic acid vaccines produce less than optimal responses in humans, our strategy relies on administering a recombinant protein-based vaccine, together with adjuvants that promote Th1-type immunity. Here we describe a process for the purification and refolding of recombinant TSA-1 expressed in Escherichia coli. The overall yield (20-25%) and endotoxin level of the purified recombinant TSA-1 (rTSA-1) is suitable for pilot scale production of the antigen for use in phase 1 clinical trials. Mice infected with T. cruzi were treated with rTSA-1, either alone or with Toll-like receptor 4 (TLR-4) agonist adjuvants including monophosphoryl lipid A (MPLA), glucopyranosyl lipid A (GLA, IDRI), and E6020 (EISEI, Inc). TSA-1 with the TLR-4 agonists was effective at reducing parasitemia relative to rTSA-1 alone, although it was difficult to discern a therapeutic effect compared to treatment with TLR-4 agonists alone. However, rTSA-1 with a 10 ug dose of MPLA optimized reductions in cardiac tissue inflammation, which were significantly reduced compared to MPLA alone. It also elicited the lowest parasite burden and the highest levels of TSA-1-specific IFN-gamma levels and IFN-gamma/IL-4 ratios. These results warrant the further evaluation of rTSA-1 in combination with rTc24 in order to maximize the therapeutic effect of vaccine-linked chemotherapy in both mice and non-human primates before advancing to clinical development.

Immune responses to gp82 provide protection against mucosal Trypanosoma cruzi infection.

The potential use of the Trypanosoma cruzi metacyclic trypomastigote (MT) stage-specific molecule glycoprotein-82 (gp82) as a vaccine target has not been fully explored. We show that the opsonization of T. cruzi MT with gp82-specific antibody prior to mucosal challenge significantly reduces parasite infectivity. In addition, we investigated the immune responses as well as the systemic and mucosal protective immunity induced by intranasal CpG-adjuvanted gp82 vaccination. Spleen cells from mice immunized with CpG-gp82 proliferated and secreted IFN-γ in a dose-dependent manner in response to in vitro stimulation with gp82 and parasite lysate. More importantly, these CpG-gp82-immunized mice were significantly protected from a biologically relevant oral parasite challenge.

Current status of vaccination against African trypanosomiasis.

Anti-trypanosomiasis vaccination still remains the best theoretical option in the fight against a disease that is continuously hovering between its wildlife reservoir and its reservoir in man and livestock. While antigenic variation of the parasite surface coat has been considered the major obstacle in the development of a functional vaccine, recent research into the biology of B cells has indicated that the problems might go further than that. This paper reviews past and current attempts to design both anti-trypanosome vaccines, as well as vaccines directed towards the inhibition of infection-associated pathology.

Immunotherapy of Trypanosoma cruzi infection with DNA vaccines in mice.

The mechanisms involved in the pathology of chronic chagasic cardiomyopathy are still debated, and the controversy has interfered with the development of new treatments and vaccines. Because of the potential of DNA vaccines for immunotherapy of chronic and infectious diseases, we tested if DNA vaccines could control an ongoing Trypanosoma cruzi infection. BALB/c mice were infected with a lethal dose (5 x 10(4) parasites) as a model of acute infection, and then they were treated with two injections of 100 microg of plasmid DNA 1 week apart, beginning on day 5 postinfection. Control mice had high levels of parasitemia and mortality and severe cardiac inflammation, while mice treated with plasmid DNA encoding trypomastigote surface antigen 1 or Tc24 had reduced parasitemia and mild cardiac inflammation and >70% survived the infection. The efficacy of the immunotherapy also was significant when it was delayed until days 10 and 15 after infection. Parasitological analysis of cardiac tissue of surviving mice indicated that most mice still contained detectable parasite kinetoplast DNA but fewer mice contained live parasites, suggesting that there was efficient but not complete parasite elimination. DNA vaccine immunotherapy was also evaluated in CD1 mice infected with a low dose (5 x 10(2) parasites) as a model of chronic infection. Immunotherapy was initiated on day 70 postinfection and resulted in improved survival and reduced cardiac tissue inflammation. These results suggest that DNA vaccines have strong potential for the immunotherapy of T. cruzi infection and may provide new alternatives for the control of Chagas' disease.

Nota sobre a variaçäo do comportamento da patogenicidade de imunogecidade de cepas de T. cruzi variedade hastatus E y mutante após longo período de manutençäo em cultura / Note about variation of the patogenicity and immunogenicity of strains of T. cruzi, hastatus and Y, maintained in cultures for longer periods

Neste trabalho nos reportamos a falta absoluta de proteçäo de camundongos isogênicos Balc-C, desafiados com cepa Y de T. cruzi patogênica, que se deu tanto com formas de cultivo epimastigotas e tripomastigotas induzidas em cultivo e delas associadas, quanto com as glicoproteínas de superfície do T.hastatus mais adjuvantes. A cepa do T. hastatus foi a mesma que há anos atrás havia protegido camundongos näo isogênicos desafiados com a mesma cepa Y patogênica. Estes dados sugeriram a perda da imunogenicidade da cepa de T. hastatus, já näo infectante, ao longo de sua manutençäo em cultura. Na mesma ocasiäo, foi constatada a perda da infectividade da cepa Y mutante de T. cruzi mantida por 15 anos em cultura. Consideraçöes de natureza teórica foram feitas admitindo-se que as variaçöes biológicas e de natureza bioquímica ocorrentes com T. cruzi se desenvolvem permanente e evolutivamente, provavelmente em funçäo de mutaçöes na dependência da interaçäo entre o parasita e os componentes do meio em que ele vive