Higher Cytokine and Opsonizing Antibody Production Induced by Bovine Serum Albumin (BSA)-Conjugated Tetrasaccharide Related to Streptococcus pneumoniae Type 3 Capsular Polysaccharide.

Nadezhda B. Egorova,Anton E. Zaytsev,Nikolay E. Nifantiev,Nelli K. Akhmatova,Yury E. Tsvetkov,Elina A. Akhmatova,Natalya E. Yastrebova,Dmitriy V. Yashunsky,Ekaterina A. Kurbatova,Elena V. Sukhova

doi:10.3389/fimmu.2020.578019

Abstract

A number of studies have demonstrated the limited efficacy of S. pneumoniae type 3 capsular polysaccharide (CP) in the 13-valent pneumococcal conjugate vaccine against serotype 3 invasive pneumococcal diseases and carriage. Synthetic oligosaccharides (OSs) may provide an alternative to CPs for development of novel conjugated pneumococcal vaccines and diagnostic test systems. A comparative immunological study of di–, tri–, and tetra–bovine serum albumin (BSA) conjugates was performed. All oligosaccharides conjugated with biotin and immobilized on streptavidin-coated plates stimulated production of IL-1α, IL-2, IL-4, IL-5, IL-10, IFNγ, IL-17A, and TNFα, but not IL-6 and GM-CSF in monocultured mice splenocytes. The tetrasaccharide–biotin conjugate stimulated the highest levels of IL-4, IL-5, IL-10, and IFNγ, which regulate expression of specific immunoglobulin isotypes. The tetra–BSA conjugate adjuvanted with aluminum hydroxide elicited high levels of IgM, IgG1, IgG2a, and IgG2b antibodies (Abs). Anti-CP-induced Abs could only be measured using the biotinylated tetrasaccharide. The tetrasaccharide ligand possessed the highest binding capacity for anti-OS and antibacterial IgG Abs in immune sera. Sera to the tetra–BSA conjugate promoted greater phagocytosis of bacteria by neutrophils and monocytes than the CRM197-CP-antisera. Sera of mice immunized with the tetra–BSA conjugate exhibited the highest titer of anti-CP IgG1 Abs compared with sera of mice inoculated with the same doses of di– and tri–BSA conjugates. Upon intraperitoneal challenge with lethal doses of S. pneumoniae type 3, the tri– and tetra–BSA conjugates protected mice more significantly than the di–BSA conjugate. Therefore, it may be concluded that the tetrasaccharide ligand is an optimal candidate for development of a semi-synthetic vaccine against S. pneumoniae type 3 and diagnostic test systems.

Highlights

Gram-positive, encapsulated, opportunistic bacteria Streptococcus pneumoniae are a major cause of upper and lower respiratory tract infection, meningitis, bacteremia, and acute otitis media
We evaluated Th1/Th2/Th17 cytokine production by mononuclear cells of mice against biotinylated OSs immobilized on streptavidin-coated plates in vitro; OS conjugate-induced Abs isotypes; antigen-binding and opsonophagocytic capacity of glycoconjugate-induced Abs compared with Abs elicited by conjugated capsular polysaccharide (CP); and active protection of glycoconjugateimmunized mice upon challenge with S. pneumoniae serotype 3
All biotinylated OSs stimulated production of IL-1a, IL-2, IL-4, IL-5, IL-10, IFNg, IL-17A, and TNFa compared to the control (P < 0.05), but failed to induce production of IL-6 and GM-CSF (Figure 3)

Summary

Introduction

Gram-positive, encapsulated, opportunistic bacteria Streptococcus pneumoniae are a major cause of upper and lower respiratory tract infection, meningitis, bacteremia, and acute otitis media. The chemical structure of pneumococcal capsular polysaccharide (CP) determines the serotype of S. pneumoniae. Among more than 90 serotypes of pneumococci, approximately 20 serotypes, including S. pneumoniae type 3, are clinically significant, and their CPs are used to prepare polysaccharide and conjugate pneumococcal vaccines. Diseases caused by S. pneumoniae serotype 3 are associated with high risk of mortality in children and adults [3, 12,13,14,15,16,17,18,19], causing global concern. Greater capsule thickness and free CP (i.e., not covalently linked to peptidoglycan) released during growth may explain the high virulence of S. pneumoniae serotype 3 and low efficacy of CP as a component of pneumococcal vaccines [21]. The immunologically insufficient bacterial CP obtained from culture fluid [26] cannot be substituted with a structurally more reliable, chemically synthesized product due to the difficult nature of polysaccharide synthesis [27]

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