While most cases of gonorrhea are benign and asymptomatic, considerable morbidity results from severe or complicated salpingitis or pelvic inflammatory disease (PID). Approximately one-fourth of all women who have had acute salpingitis will experience one or more long-term sequelac, including infertility, ectopic pregnancy, and chronic pelvic pain. The sensitivity of various strains of Neisseria gonorrhoeae to killing by normal human serum (NHS) in vitro has been correlated with different disease syndromes and has been most closely related to differences in lipooligosaccharide (LOS) structure. While in vitro data have shown that gonococcal strains with different LOS structure differ in their ability to generate the complement-derived chemotactic peptide C5a and our preliminary data suggest that this may be related to differences in cleavage of C3b associated with different LOS structures, the mechanisms intrinsic to these differences remain to be elucidated. Furthermore, most gonococcal LOS has been shown to be sialylated in vivo. While surface-associated sialic acid has been shown in several bacterial systems to decrease activation of the alternative complement pathway, it is not yet clear what role sialylation of LOS plays in the pathogenesis of different gonococcal syndromes. Antibody directed against LOS influences neutrophil chemotaxis as well as phagocytosis and killing, but the mechanisms for these findings have not yet been clearly defined. This proposal will seek to define a training program to allow the candidate to acquire technical expertise in the fields of cell biology, biochemistry, immunochemistry, and molecular biology that will allow him to: 1) define the mechanisms by which different LOS structures (including sialylated LOS) alter the deposition and cleavage of complement components; 2) define the effects of different LOS structures (in the presence and absence of serum or anti-LOS monoclonal antibodies ([MAbs]) upon the phagocytosis and killing of gonococci; and 3) examine the effects of different LOS structures upon mediator release by mononuclear cells and complement receptor regulation of neutrophils. A major goal of this proposal is to investigate conserved and widely prevalent (as defined by MAb binding), in vivo expressed LOS epitopes for activity as bactericidal and opsonophagocytic epitopes. Should these epitopes prove to be promising targets, knowledge and expertise in molecular biological techniques acquired during Phase I will be used in Phase II to derive the genomic DNA sequences encoding the hypervariable regions of an anti-idiotypic MAb (raised against the primary anti-LOS MAb) that serves as an antigenic surrogate of the LOS epitope(s). These sequences will then be used to generate a candidate vaccine antigen, either in the form of a """"""""humanized"""""""" MAb or, alternatively, by coupling the derived protein to an immunogenic carrier molecule.
