Synthesis of fragments of salmonella Typhi capsular polysaccharide and their zwitterionic analogues

Capsular polysaccharides (CPS) are T-independent antigens, and therefore they are not able to induce the formation of memory B cells. Their immunogenicity can be enhanced by conjugation with a immunogenic protein, forming a T-dependent glycoconjugate. Recently it has been discovered a new group of CPS that can directly activate T cells through the traditional MHC-II-dependent mechanism. Although these molecules show a wide diversity of chemical structure, they share the common characteristic of presenting a zwitterionic charge motif distributed along the chain, i.e. they contain both positive (e.g. NH3+) and negative (e.g. phosphate or carboxylate) charge centers within a repeating unit structure (zwitterionic polysaccharides, ZPS). This zwitterionic charge motif is believed to be responsible of their peculiar immunological activity, which is unique among bacterial polysaccharides. ZPS might offer previously unrecognized opportunities for the design of new classes of vaccines, based on the artificial introduction of a zwitterionic charge motif by chemical modification of surface glycans of pathogens. The design of new T cell dependent, ZPS-based antigens for vaccine formulation, however, requires a better understanding of how ZPS antigens stimulate the host immune system and a correlation of the ZPS structural and conformational properties with their biological activity. We report the preparation of fully synthetic oligomers of Salmonella enterica serovar Typhi (often called S. Typhi) CPS, and their zwitterionic analogues. S. Typhi is a motile Gram-negative bacterium, whose CPS (often referred to as Vi antigen) is an anionic polymer composed by α-(1-4)-linked N-acetyl galactosaminuronic acid repeating units predominantly O-acetylated at position 3. The synthesis of Vi oligomers required a strategy based on two versatile intermediates suitably protected for the systematic introduction of charge centres on each repeating unit, and containing non participating protecting groups onto 2-amino functions to allow the formation of 1,2-cis glycosidic linkages. Both are easily attainable from commercially available D-galactosamine hydrochloride. We employed the non participating azide group, which can be converted into the amino group (ZPS) or into the acetamido function (natural Vi) in the late stages of the synthesis. After extensive exploration, the glycosylation reactions were carried out using N-phenyl-trifluoroacetimidates as glycosyl donors, yielding stereoselectively the desired α product. The immunological properties of the synthetic oligomers will be investigated in order to correlate the structural features with their biological behaviour.