Discovery of Oligosaccharide Antigens for Semi-Synthetic Glycoconjugate Vaccine Leads against Streptococcus suis Serotypes 2, 3, 9 and 14*

Abstract

Streptococcus suis bacteria are one of the most serious health problems for pigs and an emerging zoonotic agent in humans working in the swine industry. S. suis bacteria express capsular polysaccharides (CPS) a major bacterial virulence factor that define the serotypes. Oligosaccharides resembling the CPS of S. suis serotypes 2, 3, 9, and 14 have been synthesized, glycans related to serotypes 2 and 9 were placed on glycan array surfaces to screen blood from infected pigs. Lead antigens for the development of semi-synthetic S. suis serotypes 2 and 9 glycoconjugate veterinary vaccines were identified in this way.

Keywords: carbohydrates; glycans; immunology; oligosaccharides; total synthesis.

Figure 1

Structures of the most common S. suis CPS repeating units. Highlighted structures were the focus of the studies disclosed here.

Figure 2

Structure of S. suis serotype 2 CPS and related oligosaccharides to be prepared from building blocks 6–12.

Scheme 1

Synthesis of linear oligosaccharides 1, 2 and 3.

Scheme 2

Synthesis of branched oligosaccharides 4 and 5.

Figure 3

Glycan array analysis of S. suis serotype 2 oligosaccharides and native CPS. A) Printing pattern of microarray and binding pattern of rabbit serum to immobilized glycans. B) IgG antibody binding to glycans. A serum dilution of 1:100 was used. MFI, mean fluorescence in intensity (mean ±standard deviation); PB, printing buffer; CWPS, cell wall polysaccharide; CPS and synthetic glycans, see Figure S1.

Figure 4

Structure of S. suis serotype 3 CPS and synthetic oligosaccharide antigens resembling the S. suis serotype 3 CPS derived from building blocks 13, 41 and 42.

Scheme 3

Synthesis of oligosaccharides 34–40 related to S. suis serotype 3 CPS.

Figure 5

Structure of S. suis serotype 9 CPS and related oligosaccharides 58–65 to be prepared from building blocks 66–73.

Scheme 4

Synthesis of oligosaccharide fragments of S. suis serotype 9 CPS 58–65.

Figure 6

α Oligosaccharide antigens resembling S. suis serotype 9 CPS.

Scheme 5

Assembly of α oligosaccharides 87–93.

Figure 7

Glycan array analysis of S. suis serotype 9 oligosaccharides and native CPS. A) Printing pattern of microarray and binding of rabbit serum to immobilized glycans. B) IgG antibody binding to glycans. A serum dilution of 1:100 was used. MFI, mean fluorescence intensity (mean ±standard deviation); PB, printing buffer; CWPS, cell wall polysaccharide; CPS and synthetic glycans, see Figure S3.

Figure 8

Structure of S. suis serotype 14 CPS and retrosynthetic analysis of related oligosaccharides. (CMP=cytidine monophosphate.)

Scheme 6

Synthesis of S. suis serotype 14‐related oligosaccharides. Reagents and conditions: a) building block (5 equiv), TMSOTf, −30 °C (5 min)→−10 °C (40 min); b) Ac₂O, MsOH, DCM; c) piperidine, DMF; d) hν; overall yield (based on resin loading): 20 % for 120; 56 % for 121; e) H₂, Pd/C, THF/MeOH/AcOH; f) NaOMe, MeOH; 40 °C; 52 % over two steps for 111; 53 % over two steps for 112.