Neoglycoconjugates from synthetic tetra- and hexasaccharides that mimic the terminus of the O-PS of Vibrio cholerae O:1, serotype Inaba.

A glycosyl acceptor and a glycosyl donor having the N-3-deoxy-L-glycero-tetronic acid side chain already attached have been prepared and used for the synthesis of the di-through to the hexasaccharide that mimic the upsteam terminus of the O-specific polysaccharide of Vibrio cholerae O:1, serotype Inaba. The target tetra- and the hexasaccharide, which were obtained in the form of 5-methoxycarbonylpentyl glycosides, were linked to BSA using squaric acid diester chemistry. The conjugation reactions were monitored by surface enhanced laser desorption ionization-time of flight mass spectrometry (SELDI-TOF MS). This allowed the progression of the conjugation of the synthetic oligosaccharides in a controlled way and termination of the reaction when the desired molar hapten/BSA ratio had been reached, yielding neoglycoconjugates with predetermined carbohydrate/carrier ratios. The ability to monitor the conjugation by the SELDI-TOF MS technique made it possible to prepare, from one hapten in a one-pot reaction, several neoglycoconjugates having different, predetermined carbohydrate/carrier ratios.

Induction of protective immunity by synthetic Vibrio cholerae hexasaccharide derived from V. cholerae O1 Ogawa lipopolysaccharide bound to a protein carrier.

Synthetic antigens that mimic the terminal hexasaccharide epitope of the O-specific polysaccharide of Vibrio cholerae O1, serotype Ogawa, were conjugated to bovine serum albumin (BSA). Conjugates with carbohydrate-to-carrier molar ratios of 15.5:1, 9.2:1, and 4.6:1 were tested for immunogenicity and efficacy in mice. The role of preimmunity to BSA and the use of adjuvant in the generation of the serologic response to the O-specific polysaccharide and protection against virulent V. cholerae was examined. Preimmunity to BSA did not affect the anti-Ogawa titers but seemed to enhance the protective capacity of antiserum. All 3 conjugates were immunogenic, but adjuvant was effective at inducing higher and earlier antibody responses. In tertiary serum samples, a correlation was found between vibriocidal activity and protection. The protective capacity of antiserum was evident in serum from mice immunized with all conjugates, but it was highest in the groups that received the conjugate with the lowest level of substitution. Further studies are required to increase understanding of the reason for differential protection.