Identification of Schistosoma mansoni fucosyltransferases and their evaluation as therapeutic targets

The fucosylated glycoconjugates of the parasite Schistosoma mansoni are directly involved in many important aspects of the parasite's life cycle, as well as in the pathogenesis associated with Schistosoma infection. Therefore, the enzymes involved in the synthesis of these fucosylated carbohydrates are promising targets for the development of new therapies. The presence of fucosyltransferase activities was investigated in various developmental stages of Schistosoma mansoni by the use of a new enzymatic assay which takes advantage of the resolution of fluorophore-assisted carbohydrate electrophoresis and the sensitivity of phosphoimager detection of radioactive fucose. The total fucosyltransferase specific activity in egg extracts was 50-fold higher than in the other live-stages tested, and the presence of putative fucosyltransferases with alpha1,3 and alpha1,2 activities and a low level of alpha1,4 activity was determined.;A molecular probe was used to isolate one fucosyltransferase of Schistosoma mansoni with a DNA sequence similarity of 84.6% and 63.7% to mouse and human fucosyltransferase type VII. The enzymatic activity of this SmFuc-T fucosyltransferase was shown to catalyze the synthesis of the sialo-Lewisx (SLex) structure in vitro and in vivo. Since the presence of sialic acid in schistosomes has not been clearly demonstrated, the developmental expression of the SmFuc-T in schistosomes was investigated in order to help elucidate the role played by the enzyme in vivo. The enzymatic activity of the fucosyltransferase during specific life stages of the parasite was determined in vitro using sialylated acceptor sugars. The enzymatic activity was about 100-fold higher in egg extracts than in adults or cercariae. A peak that co-migrated with a sialic acid standard was identified in egg membranes by high-performance anion exchange chromatography (HPAEC-AD); this peak was abolished by treatment with neuraminic acid aldolase, suggesting that the SLex epitope may be present on schistosome eggs.;The effect of fucose-type iminosugars on schistosome fucosyltransferases was then investigated in vitro and in vivo. Fucose-type 1-N-iminosugars were 4 to 6-fold more potent inhibitors of the schistosome fucosyltransferases in vitro than were the conventional deoxyfuconojirimycin iminosugars. 1-N-Iminosugar A at a concentration of 5 muM was able to block the expression of the pathogenic fucosylated epitope recognized by mAb 128C3/3 in adult worms in vivo. The mAb 128C3/3 was also found to reduce the worm burden in infected mice by 48.5% in a passive transfer experiment strongly suggesting that the carbohydrate target of this mAb is a viable candidate antigen for the development of an anti-schistosome vaccine.