Neisseria meningitidis is serologically classified into serogroups, serotypes and immunotypes based on capsular polysaccharides, outer-membrane proteins and lipooligosaccharides (LOSs), respectively. The LOS is one of the major surface antigens and can be divided into 12 immunotypes. Epidemiological studies have revealed that the L3 immunotype is the most common immunotype among serogroups B and C disease strains. We have been investigating the potential use of nontoxic LOS-based protein conjugates as vaccines for the prevention of group B disease. We selected an OP- mutant LOS instead of the L3 LOS to synthesize the LOS-based protein conjugate for two reasons. (1) Rabbit antisera to the OP- LOS cross-reacted with the L3 LOS and (2) unlike the L3 LOS, the OP- LOS does not contain the lacto-N-neotetraose (LNnT) blood group structure. Thus, there is no risk of inducing antibodies by the conjugate that may cross react with the LNnT-containing glycolipids (such as paragloboside) on human tissues. For the synthesis of the conjugate, the OP- LOS was first hydrolyzed in 1% acetic acid to separate the LOS oligosaccharide (OS) from the toxic lipid A. To create a linker in the carrier protein, glyceraldehyde was coupled to (-amino groups of lysine in meningococcal outer-membrane proteins (OMP) by reductive amination. Periodate oxidation of the glyceraldehyde linker on OMP generated new aldehyde functional groups. Finally, the amino group on the nontoxic OS was conjugated to the outer-membrane proteins (OMP) by a second reductive. The resulting conjugate was immunogenic in mice and induced IgG antibody to the native LOS. We are currently examining its immunogenicity in rabbits and improving the conjugation procedure. Antibodies to LPS (LOS) are elicited in convalescent sera from patients with gram-negative infection including Neisseria meningitidis. Detoxified LPS from different pathogens have been conjugated to proteins in many INDs as potential vaccines to induce antibodies to LPS in vaccine recipients for the prevention of diseases caused by these pathogens. Our project on the LOS-based oligosaccharide-protein conjugates provides us fundamental and current knowledge on conjugate vaccines for use in our regulatory activities.
