Yu. E. Tsvetkov

Synthesis of oligosaccharide fragments of mannan from Candida albicans cell wall and their BSA conjugates

Abstract3-Aminopropyl glycosides of α-D-mannopyranosyl-(1→2)-α-D-mannopyranosyl-(1→2)-α-D-mannopyranosyl-(1→2)-α-D-mannopyranose, α-D-mannopyranosyl-(1→3)-α-D-mannopyranosyl-(1→2)-α-D-mannopyranosyl-(1→2)-α-D-mannopyranose, and α-D-mannopyranosyl-(1→2)-[α-D-mannopyranosyl-(1→3)]-α-D-mannopyranosyl-(1→2)-α-D-mannopyranosyl-(1→2)-α-D-mannopyranose were efficiently synthesized starting from ethyl 2-O-acetyl(benzoyl)-3,4,6-tri-O-benzyl-1-thio-α-D-mannopyranoside, ethyl 4,6-di-O-benzyl-2-O-benzoyl-1-thio-α-D-mannopyranoside, ethyl 4,6-di-O-benzyl-2,3-di-O-benzoyl-1-thio-α-D-mannopyranoside, and 2,3,4,6-tetra-O-benzoyl-α-D-mannopyranosyl bromide. The oligosaccharide chains synthesized correspond to the three structural types of side chains of mannan from Candida albicans cell wall. A conjugate of the third pentasaccharide with bovine serum albumin was prepared using the squarate method.

Synthesis of oligosaccharide fragments of the Streptococcus pneumoniae type 14 capsular polysaccharide and their neoglycoconjugates with bovine serum albumin

Abstract2-Aminoethyl glycosides of tetra-, hexa- and octasaccharide fragments of the bacteria Streptococcus pneumoniae type 14 capsular polysaccharide were synthesized within a project directed to the development of pneumococcal conjugated vaccine based on synthetic carbohydrate ligands. Squarate method was used to obtain neoglycoconjugates of the synthesized oligosaccharides with bovine serum albumin.

The study of the reaction of terminated oligomerization in the synthesis of oligo-(β1-6)-glucosamines

The applicability of terminated oligomerization to the synthesis of oligo-(β1-6)-glucosamines, fragments of the intercellular polysaccharide adhesin of staphylococci, was studied. The reactions of terminated oligomerization were carried out with mono-, di-, and trisaccharide monomers and N-protected aminopropanol; and spacered mono-and disaccharides as terminating molecules were also attempted. The primary formation of cyclic products of monomer intramolecular glycosylation was observed in almost all the reactions. Only the experiments with the monomer based on the disaccharide bromide under the conditions of the Helferich reaction led to reduced yields (30%) of the cyclic products. However, even in this case, the desired terminated oligosaccharides were generated in approximately 10% yield and mainly were the products of single glycosylation of the terminator by the monomer. These experiments allow the conclusion that, under the examined conditions, the reaction of terminated oligomerization could not result in the synthesis of oligoglucosamines with a high molecular mass.

P-selectin blocking potency of multimeric tyrosine sulfates in vitro and in vivo

P-selectin blocking potency was investigated using synthetic monomeric and polymeric anionic compounds containing sulfate groups such as O-sulfotyrosine (sTyr) and/or sulfated Lewis structures. A non-carbohydrate-containing polyacrylamide conjugate sTyr-PAA (80% mol of sTyr) was a remarkably potent inhibitor of P-selectin binding in vitro, having an IC(50) value of 6 ng/mL (equivalent to 10 nM calculated on the basis of sTyr residues or 0.1 nM calculated by the mass of the macromolecule). The inhibitory effect of sTyr-PAA (80%) towards P-selectin is significantly greater than that of fucoidan (IC(50), 100 ng/mL). However, sTyr-PAA (80%) was less effective than fucoidan at reducing neutrophil extravasation in an in vivo rat model of peritonitis.

Synthetic β-(1→3)-d-glucooligosaccharides: model compounds for the mechanistic study of β-(1→3)-d-glucan bioactivities and design of antifungal vaccines

Synthetic methods used for the preparation of linear β-(1→3)-d-glucooligosaccharides with three and more monosaccharide units and their conjugates with carrier proteins, as well as the application of such derivatives in the mechanistic study of bioactivites of natural β-(1→3)-d-glucans and in the design of conjugated antifungal vaccines are considered.

Synthesis of Aminoethyl Glycosides of the Ganglioside GM1 and Asialo-GM1 Oligosaccharide Chains

Abstract4′-O-Glycosylation of 2-azidoethyl 2,3,6-tri-O-benzyl-4-O-(2,3-di-O-benzyl-6-O-benzoyl-β-D-galactopyranosyl)-β-D-glucopyranoside with a disaccharide donor, 4-trichloroacetamidophenyl 4,6-di-O-acetyl-2-deoxy-3-O-(2,3,4,6-tetra-O-acetyl-β-D-galactopyranosyl)-1-thio-2-trichloroacetamido-β-D-galactopyranoside, in dichloromethane in the presence of N-iodosuccinimide and trifluoromethanesulfonic acid resulted in a tetrasaccharide, 2-azidoethyl (2,3,4,6-tetra-O-acetyl-β-D-galactopyranosyl)-(1 → 3)-(4,6-di-O-acetyl-2-deoxy-2-trichloroacetamido-β-D-galactopyranosyl)-(1 → 4)-(2,3-di-O-benzyl-6-O-benzoyl-β-D-galactopyranosyl)-(1 → 4)-2,3,6-tri-O-benzyl-β-D-glucopyranoside, in 69% yield. The complete removal of O-protecting groups in the tetrasaccharide, the replacement of N-trichloroacetyl by N-acetyl group, and the reduction of the aglycone azide group to amine led to the target aminoethyl glycoside of β-D-Gal-(1 → 3)-β-D-GalNAc-(1 → 4)-β-D-Gal-(1 → 4)-β-D-Glc-OCH2CH2NH2 containing the oligosaccharide chain of asialo-GM1 ganglioside in 72% overall yield. Selective 3′-O-glycosylation of 2-azidoethyl 2,3,6-tri-O-benzyl-4-O-(2,6-di-O-benzyl-β-D-galactopyranosyl)-β-D-glucopyranoside with thioglycoside methyl (ethyl 5-acetamido-4,7,8,9-tetra-O-acetyl-3,5-dideoxy-2-thio-D-glycero-α-D-galacto-2-nonulopyranosyl)oate in acetonitrile in the presence of N-iodosuccinimide and trifluoromethanesulfonic acid afforded 2-azidoethyl [methyl (5-acetamido-4,7,8,9-tetra-O-acetyl-3,5-dideoxy-D-glycero-α-D-galacto-2-nonulopyranosyl)oate]-(2 → 3)-(2,6-di-O-benzyl-β-D-galactopyranosyl)-(1 → 4)-2,3,6-tri-O-benzyl-β-D-glucopyranoside, the selectively protected derivative of the oligosaccharide chain of GM3 ganglioside, in 79% yield. Its 4′-O-glycosylation with a disaccharide glycosyl donor, (4-trichloroacetophenyl-4,6-di-O-acetyl-2-deoxy-3-O-(2,3,4,6-tetra-O-acetyl-β-D-galactopyranosyl) 1-thio-2-trichloroacetamido-β-D-galactopyranoside in dichloromethane in the presence of N-iodosuccinimide and trifluoromethanesulfonic acid gave 2-azidoethyl (2,3,4,6-tetra-O-acetyl-β-D-galactopyranosyl)-(1 → 3)-(4,6-di-O-acetyl-2-deoxy-2-trichloroacetamido-β-D-galactopyranosyl)-(1 → 4)-{[methyl (5-acetamido-4,7,8,9-tetra-O-acetyl-3,5-dideoxy-D-glycero-α-D-galacto-2-nonulopyranosyl)onate]-(2 → 3)}-(2,6-di-O-benzyl-β-D-galactopyranosyl)-(1 → 4)-2,3,6-tri-O-benzyl-β-D-glucopyranoside in 85% yield. The resulting pentasaccharide was O-deprotected, its N-trichloroacetyl group was replaced by N-acetyl group, and the aglycone azide group was reduced to afford in 85% overall yield aminoethyl glycoside of β-D-Gal-(1 → 3)-β-D-GalNAc-(1 → 4)-[α-D-Neu5Ac-(2 → 3)]-β-D-Gal-(1 → 4)-β-D-Glc-OCH2CH2NH2 containing the oligosaccharide chain of GM1 ganglioside.

The use of biotinylated oligosaccharides related to fragments of capsular polysaccharides from Streptococcus pneumoniae serotypes 3 and 14 as a tool for assessment of the level of vaccine-induced antibody response to neoglycoconjugates

In this report, the data are presented on the use of synthetic biotinylated oligosaccharides corresponding to different fragments of capsular polysaccharides (CP) from S. pneumoniae serotypes 3 and 14 for evaluation of the level of vaccine-induced antibodies in the serum of mice immunized with conjugates of oligosaccharide ligands with bovine sera albumin (BSA). The level of antigen-specific antibodies assessed by enzyme-linked immunosorbent assay (ELISA) using biotinylated oligosaccharides immobilized on the surface of streptavidin precoated ELISA plates was substantially higher than that assessed using conventional ELISA plates with the bacterial CP as a coating antigen. Hence, the ELISA protocol using biotinylated oligosaccharides is much more sensitive as compared to the conventional protocol, and it can be used for the detection of antibodies in highly diluted antisera. The highest antibody titer against biotinylated oligosaccharides was detected in the antisera of mice immunized with the conjugates of BSA with tetrasaccharide ligands related to fragments of CP of two pneumococcal serotypes under study. The antigen-binding ability of antibodies to tetrasaccharides related to one and two repeating units of CP from S. pneumoniae types 14 and 3, respectively, was higher than that of antibodies to BSA conjugated with oligosaccharides of other chemical structure in the reaction of biotinylated oligosaccharides and in the bacterial agglutination test with live cells on the glass slide but not in the inhibition of ELISA with bacterial CP. This research provided a rationale for the choice of tetrasaccharide ligands for the design of the third generation vaccines against S. pneumoniae types 3 and 14 and showed the validity of the ELISA protocol with biotinylated oligosaccharides as coating antigens for the assessment of the level of neoglycoconjugate-induced antibodies.

Protective Activity of a Glycoconjugate Based on a Synthetic Hexasaccharide Related to a Fragment of Capsular Streptococcus Pneumoniae Serotype 14 Polysaccharide Chain

We studied the effects of immunization with a conjugate of carrier protein and hexasaccharide ligand related to a fragment of capsular of Str. pneumoniae serotype 14 polysaccharide chain on activation of innate and adaptive immunity. It was found that two-fold immunization with the glycoconjugate adsorbed on aluminum hydroxide significantly increased the titer of IgG antibodies to capsular polysaccharide in the blood and protected 100% mice from infection with Str. pneumoniae serotype 14. Enhanced bactericidal activity of peripheral blood lymphocytes of mice was found 4 and 24 h after the first immunization with the immobilized glycoconjugate. Adsorption of the glycoconjugate on aluminum hydroxide resulted in modification of the immune processes at the stage of activation of innate immunity and subsequent strengthening of the adaptive immunity.

Syntheses of α- and β-Glycosyl Donors with a Disaccharide β-D-Gal-(1→3)-D-GalNAc Backbone

The synthesis of thioglycoside glycosyl donors with a disaccharide β-D-Gal-(1 → 3)-D-GalNAc backbone was studied using the glycosylation of a series of suitably protected 3-monohydroxy- and 3,4-dihydroxyderivatives of phenyl 2-azido-2-deoxy-1-thio-α- and 1-thio-β-D-galactopyranosides by galactosyl bromide, fluoride, and trichloroacetimidate. In the reaction with the monohydroxylated glycosyl acceptor, the process of intermolecular transfer of thiophenyl group from the glycosyl acceptor onto the cation formed from the molecule of glycosyl donor dominated. When glycosylating 3,4-diol under the same conditions, the product of the thiophenyl group transfer dominated or the undesired (1 → 4), rather than (1 → 3)-linked, disaccharide product formed. The aglycon transfer was excluded when 4-nitrophenylthio group was substituted for phenylthio group in the galactosyl acceptor molecule. This led to the target disaccharide, 4-nitrophenyl 2-azido-4,6-O-benzylidene-2-deoxy-3-O-(2,3,4,6-tetra-O-acetyl-β-D-galactopyranosyl)-1-thio-β-D-galactopyranoside, in 57% yield. This disaccharide product bears nonparticipating azido group in position 2 of galactosamine and can hence be used to form α-glycoside bond. Azido group and the aglycon nitro group were simultaneously reduced in this product and then trichloroacetylated, which led to the β-glycosyl donor, 4-trichloroacetamidophenyl 4,6-di-O-acetyl-2-deoxy-3-O-(2,3,4,6-tetra-O-acetyl-β-D-galactopyranosyl)-1-thio-2-trichloroacetamido-β-D-galactopyranoside, in 62% yield. The resulting glycosyl donor was used in the synthesis of tetrasaccharide asialo-GM1.

Reactivity of sugar trityl ethers in trityl—cyanoalkylidene condensation

The reactivity of a series of primary and secondary sugar trityl ethers in tritylcyanoalkylidene condensation has been determined. It was found that the stereoselectivity of glycosylation by this method does not directly depend on the reactivity of the trityl ethers. Based on the results obtained, a stepwise mechanism rather than a concerted mechanism has been chosen as the most probable.