Glycoconjugate vaccines are an important and successful countermeasure for control and treatment of infectious diseases. Elucidating the shortest portion of a polysaccharide able to bind to specific functional monoconal antibodies (glycotope) is key for optimal vaccine design. Historically, the determination of glycotopes has been hampered by technical challenges in obtaining well-defined oligosaccharides. New synthetic and depolymerization methods enbales the creation of glycans to study interactions with protective monoclonal antibodies. GBS is a leading cause of invasive infections in pregnant women, newborns, and elderly people, and the capsule is a major virulence factor targeted for vaccine development . GBS PSIII epitope has been historically considered the prototype of a conformational carbohydrate epitope . We have recently applied an integrated approach based on competitive ELISA/Surface Plasmon Resonance/Saturation Transfer NMR/X-ray crystallography to elucidate one epitope from the capsule of Group B Streptococcus type III, whose structure is composed of α-NeuAc-(2→3)-β-D-Gal-(1→4)-β-D-GlcNAc-(1→3)-β-D-Glc-(1→4)-β-D-Gal→, with branching at position 6 of GlcNAc. Using a panel of synthetic and semisynthetic structures, we found that the structural epitope consists of 5 disjoined sugar residues, which comprises a single repeating unit, and the GlcNAc moiety of the next consecutive repeat unit, where sialic acid is clearly engaged in antibody binding . Currently we are studying immunogenicity of vaccines based on short GBS PSIII fragments.
In addition, we are using the same approach to map the structural epitope of Neisseria meningitidis serogroup A (MenA), a Gram-negative encapsulated bacterium responsible for epidemic meningitis in the sub-Saharan region of Africa, termed meningitis belt . Despite multivalent and monovalent conjugate vaccines against MenA are now available, the structural minimal epitope of MenA polysaccharide is still unknown . MenA capsular polysaccharide (CPS) consists of (1→6) linked 2-acetamido-2-deoxy-α-D-mannopyranosyl phosphate repeating units with O-acetylation predominantly at position 3 , therefore the epitope is expected to be linear. Two parameters influence the immunogenicity of MenA conjugates: the oligomer length and the acetylation level [7,8]. Competitive ELISA revealed epitope optimization for oligomers longer than 6 repeating units. The hexamer was also sufficient to deplete the bactericidal activity of serum raised from vaccinated subjects, while inhibition of serum for shorter length oligomers was achieved at higher concentrations. Upon de-O-acetylation of MenA minimal epitope, the kinetics of dissociation increase 1000-fold and no hSBA inhibition was observed, revealing that these moieties play a pivotal role in recognition. STD-NMR effect was stronger for the O-acetyl group (100%) located at C3 followed H3/H4 and acetamide positioned at C2, indicating that these moieties are in close contact with the functional monoclonal antibody. High resolution X-ray crystal structure of a Fab in complex with a phosphosugar linked oligosaccharide revealed a trisaccharide linear epitope, where O-acetyl moieties are engaged in network of hydrogen bond. Efforts in understanding the structural basis of recognition of oligosaccharide epitopes are the foundations for future development of a MenA epitope optimal mimicry.
- Nuccitelli A et al. Ther. Adv. Vaccines 2015, 3, 76-90.
- Zou W et al. The Molecular Immunology of Complex Carbohydrates-2 2001, Ed. Albert M. Wu, Accademic/plenum Publishers, 473-84.
- Carboni F et al. Proceed. Natl. Acad. Sci. USA 2017, 114(19), 5017-22.
- Harrison LH et al. Vaccine 2009.,27 (Suppl. 2), B51-63.
- Trotter et al. Lancet Infect. Dis. 2017, 17(8), 867-72.
- Lemercinier X et al. Carbohydr. Res. 1996, 296, 83-96.
Berry DS et al. Infect. Immun. 2002. 70(7), 3707-13.
Costantino P et al. Vaccine. 1992, 10(10), 691-98.