Abstract
Nanoparticle systems are being explored for the display of carbohydrate antigens, characterized by multimeric presentation of glycan epitopes and special chemico-physical properties of nano-sized particles. Among them, outer membrane vesicles (OMVs) are receiving great attention, combining antigen presentation with the immunopotentiator effect of the Toll-like receptor agonists naturally present on these systems. In this context, we are testing Generalized Modules for Membrane Antigens (GMMA), OMVs naturally released from Gram-negative bacteria mutated to increase blebbing, as carrier for polysaccharides. Here, we investigated the impact of saccharide length, density, and attachment site on the immune response elicited by GMMA in animal models, using a variety of structurally diverse polysaccharides from different pathogens (i.e., Neisseria meningitidis serogroup A and C, Haemophilus influenzae type b, and streptococcus Group A Carbohydrate and Salmonella Typhi Vi). Anti-polysaccharide immune response was not affected by the number of saccharides per GMMA particle. However, lower saccharide loading can better preserve the immunogenicity of GMMA as antigen. In contrast, saccharide length needs to be optimized for each specific antigen. Interestingly, GMMA conjugates induced strong functional immune response even when the polysaccharides were linked to sugars on GMMA. We also verified that GMMA conjugates elicit a T-dependent humoral immune response to polysaccharides that is strictly dependent on the nature of the polysaccharide. The results obtained are important to design novel glycoconjugate vaccines using GMMA as carrier and support the development of multicomponent glycoconjugate vaccines where GMMA can play the dual role of carrier and antigen. In addition, this work provides significant insights into the mechanism of action of glycoconjugates.
Highlights
During the last years, nanoparticle systems have received increased interest for the display of carbohydrate antigens
Meningococcal serogroup A (MenA), meningococcal serogroup C (MenC), and H. influenzae type b (Hib) oligosaccharides were terminally activated with SIDEA linker [29] and randomly conjugated to lysines of Generalized Modules for Membrane Antigens (GMMA) surface proteins (Figure 1A)
Formation of saccharide– GMMA conjugates was verified by Western blotting analysis (Figure 2); and the amount of total saccharide and total protein were quantified by HPAEC-PAD and micro BCA, respectively
Summary
Nanoparticle systems have received increased interest for the display of carbohydrate antigens. Outer membrane vesicles (OMVs) combine antigen presentation with intrinsic adjuvant properties [5,6,7]. Outer membrane protein complex (OMPC) from Neisseria meningitidis has been used as carrier for Haemophilus influenzae type b conjugate vaccine [8, 9]. OMPC has been shown to possess TLR2-mediated adjuvant activity [10] and may contain TLR4 agonists such as lipopolysaccharides (LPS) since they derive from the outer membrane of Gram-negative bacteria. Streptococcus pneumoniae CPS14 capsule, for example, displayed on engineered E. coli OMVs induced IgG levels and efficacy in opsonophagocytic activity tests comparable with those induced by PCV13 [12]
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