title: Preparation, characterization, and immunogenicity of meningococcal immunotype L2 and L3,7,9 phosphoethanolamine group-containing oligosaccharide-protein conjugates.
abstract: A method was developed for the well-defined coupling of phosphoethanolamine group (PEA)- and carboxylic acid group-containing polysaccharides and oligosaccharides to proteins without the need for extensive modification of the carbohydrate antigens. The carboxylic acid group of the terminal 2-keto-3-deoxyoctulosonic acid moiety was utilized to introduce a thiol function in meningococcal immunotype L2 and L3,7,9 lipopolysaccharide-derived oligosaccharides. The thiol group-containing oligosaccharides were subsequently coupled to bromoacetylated proteins. Immunotype L2 and L3,7,9 PEA group-containing oligosaccharide-tetanus toxoid conjugates were prepared, and their immunogenicities were studied in rabbits. Both the immunotype L2 and immunotype L3,7,9 conjugates evoked high immunoglobulin G (IgG) antibody titers after the first booster injection. These conjugates also displayed an ability to induce long-lasting IgG antibody levels which could be detected until 9 months after one booster injection at week 3. The adjuvant Quil A enhanced the immune response to all the conjugates to a minor extent, which is in contrast with reported adjuvant effects of Quil A on these types of antigens in mice. A conjugate prepared from the dephosphorylated L3,7,9 oligosaccharides evoked a significantly lower IgG response than a similar PEA-containing conjugate, and enzyme-linked immunosorbent assay inhibition studies indicated a different epitope specificity. Furthermore, antisera elicited with the complete bacteria contained antibodies directed against PEA-containing epitopes, which stresses the importance of the presence of unmodified PEA groups in meningococcal lipopolysaccharide-derived oligosaccharide-protein conjugates. The procedure developed offers an elegant solution for the specific coupling of meningococcal PEA-containing oligosaccharides to proteins and may therefore be a very useful tool in the development of a vaccine against group B meningococci.
authors.affiliation: Eijkman-Winkler Laboratory of Medical Microbiology, Department of Experimental Microbiology, Academic Hospital, Utrecht University, The Netherlands.
authors.forename: A F
authors.initials: AF
authors.lastname: Verheul
authors.forename: A K
authors.initials: AK
authors.lastname: Braat
authors.forename: J M
authors.initials: JM
authors.lastname: Leenhouts
authors.forename: P
authors.initials: P
authors.lastname: Hoogerhout
authors.forename: J T
authors.initials: JT
authors.lastname: Poolman
authors.forename: H
authors.initials: H
authors.lastname: Snippe
authors.forename: J
authors.initials: J
authors.lastname: Verhoef
mesh_terms: D000818:Animals; D000907:Antibodies, Bacterial / Q000097:blood / Q000276:immunology; D000918:Antibody Specificity; D001428:Bacterial Vaccines / Q000737:chemistry / Q000276:immunology; D002240:Carbohydrate Sequence; D002264:Carboxylic Acids / Q000737:chemistry / Q000276:immunology; D002352:Carrier Proteins / Q000737:chemistry / Q000276:immunology; D004983:Ethanolamines / Q000737:chemistry* / Q000276:immunology; D007114:Immunization; D007074:Immunoglobulin G / Q000032:analysis; D008070:Lipopolysaccharides; D022401:Meningococcal Vaccines; D008969:Molecular Sequence Data; D009345:Neisseria meningitidis / Q000276:immunology*; D009844:Oligosaccharides / Q000737:chemistry* / Q000276:immunology; D010766:Phosphorylation; D011817:Rabbits; D012703:Serotyping; D013400:Sugar Acids / Q000737:chemistry; D013745:Tetanus Toxoid / Q000737:chemistry* / Q000276:immunology
chemical_list: D000907:Antibodies, Bacterial; D001428:Bacterial Vaccines; D002264:Carboxylic Acids; D002352:Carrier Proteins; D004983:Ethanolamines; D007074:Immunoglobulin G; D008070:Lipopolysaccharides; D022401:Meningococcal Vaccines; D009844:Oligosaccharides; D013400:Sugar Acids; D013745:Tetanus Toxoid; C002532:2-keto-3-deoxyoctonate; C005448:phosphorylethanolamine