Enterotoxigenic E. coli that produce heat labile (LT) and heat stable (ST) enterotoxins are a major cause of enteric disease in many areas of the world. We have found that ST is inhibited by human milk in vitro and in vivo. All of this inhibition resides in the ST inhibitory fucosyloligosaccharide (STIF). The oligosaccharide binds to the extracellular domain of guanylyl cyclase, thereby inhibiting binding of ST. The protective oligosaccharide is a large structure present in too low a concentration to be routinely measured directly by HPLC and the molecule is too large to be synthesized routinely. However, its concentration in milk may be inferred by measuring small, more plentiful, structurally homologous oligosaccharides or might be measured directly by a solid phase assay. We hypothesize that the activity of STIF resides in the fucose containing moieties at the non-reducing terminus of the molecule and, thus, can be mimicked by structurally homologous compounds. Thus, the following aims are proposed: Determine the three-dimensional conformation of the ST inhibitory fucosyloligosaccharide. Chemically synthesize STIF, as well as a series of STIF homologs designed to incorporate or approximate structural features of the natural compound to define the minimum structure necessary for activity. Search for natural homologs and analogs to STIF from milk of other species. Utilize pure STIF homolog or analog to study details of the inhibition of ST-induced transmembrane signaling. Investigate the survival of STIF oligosaccharide structure and biological activity after transit through the gastrointestinal tract of the infant. Assess the clinical relevance of STIF levels in human milk. Test the safety, tolerance and efficacy of STIF or its analogs in prevention or ameliorating ST-related enteric disease in a population of Mexican infants at high risk for ST-related disease.
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