Abstract
Current vaccination against Streptococcus pneumoniae uses vaccines based on capsular polysaccharides from selected serotypes and has led to nonvaccine serotype replacement disease. We have investigated an alternative serotype-independent approach, using multiple-antigen vaccines (MAV) prepared from S. pneumoniae TIGR4 lysates enriched for surface proteins by a chromatography step after culture under conditions that induce expression of heat shock proteins (Hsp; thought to be immune adjuvants). Proteomics and immunoblot analyses demonstrated that, compared to standard bacterial lysates, MAV was enriched with Hsps and contained several recognized protective protein antigens, including pneumococcal surface protein A (PspA) and pneumolysin (Ply). Vaccination of rodents with MAV induced robust antibody responses to multiple serotypes, including nonpneumococcal conjugate vaccine serotypes. Homologous and heterologous strains of S. pneumoniae were opsonized after incubation in sera from vaccinated rodents. In mouse models, active vaccination with MAV significantly protected against pneumonia, while passive transfer of rabbit serum from MAV-vaccinated rabbits significantly protected against sepsis caused by both homologous and heterologous S. pneumoniae strains. Direct comparison of MAV preparations made with or without the heat shock step showed no clear differences in protein antigen content and antigenicity, suggesting that the chromatography step rather than Hsp induction improved MAV antigenicity. Overall, these data suggest that the MAV approach may provide serotype-independent protection against S. pneumoniae.
Highlights
Current vaccination against Streptococcus pneumoniae uses vaccines based on capsular polysaccharides from selected serotypes and has led to nonvaccine serotype replacement disease
Heat shock was used to enrich for heat shock proteins (Hsp), and anion-exchange chromatography was used to enrich for negatively charged S. pneumoniae antigens (e.g., pneumococcal surface protein A (PspA) and Ply) (Fig. 1A)
Immunoblots determined which elution fractions contained the highest concentration of the Hsp60 and Hsp70 proteins and demonstrated a marked increase in the expression of both the Hsp60 and Hsp70 content in the multiple-antigen vaccine (MAV) compared to the bacterial heat-killed lysate (HKL) (Fig. 1B and C)
Summary
Current vaccination against Streptococcus pneumoniae uses vaccines based on capsular polysaccharides from selected serotypes and has led to nonvaccine serotype replacement disease. One method of enhancing immunogenicity is altering the preparation of the lysate to ensure increased representation of immunoprotective proteins This can be partially achieved using anion-exchange chromatography with a pH 8.0 buffer to preferentially capture several well-known S. pneumoniae antigens, all of which have a pI of 7.5 or lower, including PiuA, PiaA, PsaA, RrgA, RrgB, ClpP, PspA, and Ply. In addition, growth under stress conditions, such as high temperatures, to induce heat shock proteins (Hsps) could increase antigenicity [15], as Hsps facilitate the cross-presentation of peptides [16, 17] and act as natural adjuvants by stimulating macrophages and dendritic cells to cause cytokine secretion [18,19,20]. We present data on a multiple-antigen approach to a novel S. pneumoniae vaccine based on bacterial lysates that combines the advantages of a whole-cell approach with the potential additional benefit of an increased Hsp and surface antigen content in the vaccine preparation
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