Enhancing the immunogenicity of pneumococcal conjugate vaccines through site-specific glycosylation and presentation on synthetic vesicles

Northwestern University At Chicago, Evanston IL

Investigators

Abstract

PROJECT SUMMARY Streptococcus pneumoniae is a significant cause of morbidity and mortality in the United States and around the world. There are over 100 known serotypes of S. pneumoniae, and conjugate vaccines are an effective tool to protect the public from infection of covered serotypes. These vaccines work by conjugating capsular polysaccharide from unique S. pneumoniae serotypes to an immunogenic carrier protein, which induces a T cell-dependent memory immune response. However, serotype replacement, a process in which serotypes that are not covered by a vaccine increase in incidence due to the removal of competing serotypes, necessitates higher valency vaccines. The conventional, chemical technology used to synthesize pneumococcal conjugate vaccines has substantial limitations that prevent the synthesis of higher valency vaccines. This technology randomly conjugates glycan to carrier protein, which destroys T cell epitopes on the protein, reducing immunogenicity of the vaccine. Increasing valency using this conventional strategy also decreases immune responses against each serotype due to carrier suppression, or the decrease in anti-glycan immune response with higher amounts of carrier protein included per dose of vaccine. Therefore, there is a need to develop novel and more highly immunogenic vaccines with enhanced immunogenicity to increase vaccine valency and reduce carrier suppression. Protein glycan coupling technology is a novel method to enzymatically conjugate a carrier protein with pathogen glycan using an oligosaccharyltransferase. This technology enables site-specific glycosylation of carrier protein to avoid T cell epitope destruction. The central hypothesis of this proposal is that site-specific glycosylation of carrier protein using enzymatic conjugation can improve the immunogenicity of pneumococcal conjugate vaccines. Recent advances to protein glycan coupling technology have enabled its use in cell-free systems, which allows for the high-throughput synthesis of hundreds of unique glycoconjugate vaccine designs. This project will evaluate three distinct features – glycan location, density, and delivery – for synthesis of glycoconjugates via site-specific enzymatic glycosylation (Aim 1) and evaluate the immunogenicity of top candidates in mice (Aim 2). In Aim 1, glycosylation at every potential location throughout CRM197, an FDA-approved carrier protein, will be evaluated in high-throughput using cell-free systems. Sites away from T cell epitopes that can be glycosylated will be iteratively combined to increase glycan density, and glycoproteins will be attached to synthetic vesicles to improve antigen presentation. In Aim 2, multi-dose vaccine trials will be performed in mice with top vaccine candidates from Aim 1 and compared to a conventional vaccine. The immunogenicity of each vaccine will be evaluated by measuring antibody titers, opsonophagocytic activity, memory B cell production, and mouse survival after S. pneumoniae challenge. The results will inform conjugate vaccine design for S. pneumoniae and could be applied to develop novel vaccines against other pathogens.