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Abstract

Pediatric vaccines targeting bacterial capsular polysaccharides are more effective for certain types of bugs than others, and the manufacturing process as well as immunodominance of different glycan epitopes (glycotopes) can lead to a mixed immune response that does not protect against disease. To directly identify glycotopes that induce a protective response, Schumann et al. combined antibody reverse engineering with automated glycan assembly using Streptococcus pneumoniae serotype 8 as a proof of concept. Promising glycotopes conjugated to a carrier protein induced protective antibodies in mice and were also immunogenic in rabbits. When combined with a commercially available pneumococcal vaccine, these glycoconjugates were able to boost the opsonophagocytic bacterial killing ability of sera from immunized rabbits. This approach leveraging semisynthetic glycoconjugates could lead to the design of more effective bacterial vaccines.Glycoconjugate vaccines based on capsular polysaccharides (CPSs) of pathogenic bacteria such as Streptococcus pneumoniae successfully protect from disease but suffer from incomplete coverage, are troublesome to manufacture from isolated CPSs, and lack efficacy against certain serotypes. Defined, synthetic oligosaccharides are an attractive alternative to isolated CPSs but require the identification of immunogenic and protective oligosaccharide antigens. We describe a medicinal chemistry strategy based on a combination of automated glycan assembly (AGA), glycan microarray–based monoclonal antibody (mAb) reverse engineering, and immunological evaluation in vivo to uncover a protective glycan epitope (glycotope) for S. pneumoniae serotype 8 (ST8). All four tetrasaccharide frameshifts of ST8 CPS were prepared by AGA and used in glycan microarray experiments to identify the glycotopes recognized by antibodies against ST8. One tetrasaccharide frameshift that was preferentially recognized by a protective, CPS-directed mAb was conjugated to the carrier protein CRM197. Immunization of mice with this semisynthetic glycoconjugate followed by generation and characterization of a protective mAb identified protective and nonprotective glycotopes. Immunization of rabbits with semisynthetic ST8 glycoconjugates containing protective glycotopes induced an antibacterial immune response. Coformulation of ST8 glycoconjugates with the marketed 13-valent glycoconjugate vaccine Prevnar 13 yielded a potent 14-valent S. pneumoniae vaccine. Our strategy presents a facile approach to develop efficient semisynthetic glycoconjugate vaccines.