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

Abstract

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.

Glycosylation under Thermodynamic Control: Synthesis of the Di‐ and the Hexasaccharide Fragments of the O‐SP of Vibrio Cholerae O:1 Serotype Ogawa from Fully Functionalized Building Blocks

“…Our previous syntheses [11,[18][19][20][21][22] of higher oligosaccharides in the Vibrio cholerae O:1 series had their shortcomings due to either instability of protecting groups, lack of stereoselectivity of glycosylation reactions, or separation problems involved. Also, losses resulting from multiple chemical manipulations with high oligosaccharides we experienced during some previous approaches suggested that, rather than a stepwise approach, a suitable block wise strategy using fully functionalized intermediates as building blocks should be pursued.…”

Immunogens related to the synthetic tetrasaccharide side chain of the Bacillus anthracis exosporium

“…To minimize waste of the precious oligosaccharides, conjugations described here were performed at a considerably lower initial hapten-carrier ratio (20:1) than in our earlier work. [29][30][31] The expected slower reaction rate due to conducting the conjugation under these conditions 32 was fully compensated for by running conjugations at a higher concentration only in the case of hapten 84 (Table 3). We have no other explanation for the much slower rate of conjugation with haptens 2, 12, and 35, other than, as we pointed out previously, 29 that we may not be aware of some parameters that affect conjugation.…”

Synthetic Fragments ofVibrio choleraeO1 Inaba O-Specific Polysaccharide Bound to a Protein Carrier Are Immunogenic in Mice but Do Not Induce Protective Antibodies

“…Briefly (Fig. 1A), condensation (23) of the glycosyl acceptor 5 and glycosyl donor 4, prepared from the corresponding amine (7) and 2,4-O-benzylidene-3-deoxy-L-glycero-tetronic acid (24,28), gave the fully protected disaccharide 6. Partial deprotection by Zemplén deacetylation afforded alcohol 7, which was used as a glycosyl acceptor in the next coupling with 4 to extend the oligosaccharide chain and obtain the trisaccharide 8.…”

“…Immunogens 1a to 3c were prepared by linking the chemically synthesized di-, tetra-, and hexasaccharide fragments of the O-SP of V. cholerae O1 serotype Inaba to BSA by using squaric acid chemistry (20,36,38). The oligosaccharides were assembled in a stepwise manner (28) (Fig. 1) from the monosaccharide glycosyl donor 4 and the monosaccharide glycosyl acceptor 5.…”