Functional genome of Schistosoma mansoni applied to development of vaccines

Abstract

Schistosoma mansoni is the predominant parasite responsible for schistosomiasis which affects 200 million individuals in tropical regions of the world, being endemic in 74 countries, including Brazil. It is estimated that 600 million are in areas at risk. The development of the anti-parasite drug, Praziquantel, was essential for the reduction in morbidity and mortality due to schistosomiasis. However, this drug has little effect on reinfeccion, which leads to the requirement of repeated treatments. Furthermore, the description of strains resistant to this drug has raised concerns as to its future efficacy in controlling this parasite. It is therefore imperative to search for control mechanisms complementary to chemotherapy. Vaccination would be the most cost-effective approach. It is considered that it is possible to obtain an efficient vaccination against schistosomiasis, since experimental animals immunized with irradiated cercaria show up to 80% protection against a subsequent challenge with an elevated dose of the parasite, as measured by a reduction in parasitemia in immunized animals. Contrary to other diseases, sterilizing immunity is not possible, nor necessary, since serious morbidity requires high levels of infection for prolonged periods. Mathematical models show that a reduction in bacterial load of 45% would cause an enormous effect on endemicity. More than 100 antigens have been identified and tested and a dozen showed some level of protection, between 35-50%. Although some antigens have show potential and are progressing into clinical trials, an effective vaccine against schistosomiasis is still to be established, and it will probably require a combination of antigens that induce a variety of specific immune responses to reach higher levels of protection. The Schistosoma mansoni EST Genome Project, in São Paulo, Brazil, has recently generated 163.000 ORESTES (open reading frame-expressed sequence tags) from 6 stages of the parasite, resulting in 31.000 assembled gene fragments. It is estimated that this dataset represents 92% of a total 14,000 genes of the parasite, increasing 10-fold the number of ESTs available and increasing to 522 the number of complete genes of the parasite. Automatic annotation based on attributions within the Gene Ontology system, provided a vision of important biological functions of the parasite and permitted identification of a set of proteins with characteristics which could indicate some potential as vaccine candidates. The gene/proteins were selected based on their attributed functions, which may indicate a potential role in the survival of the parasite in the host and possible exposure for interaction with the immune system. Of special interest were proteins with functions such as toxins, surface receptors for cellular adhesion, surface proteins and ezymes, and receptors for host factors. Expression of the genes in the stages interacting with the host was evaluated. Furthermore, the transcriptome of the parasite revealed that ~55% of the sequences coded for protein with no definable function. Part of these may be associated with adaptation to parasitism; within these, genes which have increased expression in transition between cercaria and schistosomula/adult worms, could have interest as vaccine candidates. In parallel to the Transcriptome sequencing, we carried out a pilot project for characterization of the potential protective effect of some possible antigens identified in the beginning of the project, using DNA vaccines for presentation of the antigens to the immune system. This pilot project permitted establishing the basis of a Functional Genomics project for characterization of the protective potential of the identified genes. The present project proposes to investigate a selection of genes/proteins as potential vaccine candidates, presented as DNA vaccines. It will be organized in 3 subprojects with the following objectives: 1) construction of DNA vaccines containing 25 candidate genes, characterization of the immune response induced against the respective antigens and evaluation of the protection induced against challenge with live cercaria; 2) characterization of the differential expression of the genes in the different life stages of the parasite by microarray using a panel of 4000 selected genes; differential expression in parasites submitted to sera or cells from the immune system of immunized mice; 3) immunolocalization of the vaccine candidates in histological cuts of the parasite in the intra-host stages. This project should provide a first evaluation of the protective potential of a set of selected genes/proteins as vaccine candidates. (AU)