A BROADLY CROSS-PROTECTIVE MONOCLONAL-ANTIBODY BINDING TO ESCHERICHIA-COLI AND SALMONELLA LIPOPOLYSACCHARIDES

During the last decade, episodes of sepsis have increased and Escheric hia coli has remained the most frequent clinical isolate. Lipopolysacc harides (LPS; endotoxin) are the major toxic and antigenic components of gram-negative bacteria and qualify as targets for therapeutic inter ventions. Molecules that neutralize the toxic effects of LPS are activ ely investigated. In this paper, we describe a murine monoclonal antib ody (MAb; WN1 222-5), broadly cross-reactive and cross-protective for smooth (S)-form and rough (R)-form LPS. As shown in enzyme-linked immu nosorbent assay and the passive hemolysis assay, WN1 222-5 binds to th e five known E. coli core chemotypes, to Salmonella core, and to S-for m LPS having these core structures. In immunoblots, it is shown to rea ct with both the nonsubstituted core LPS and with LPS carrying O-side chains, indicating the exposure of the epitope in both S-form and R-fo rm LPS. This MAb of the immunoglobulin G2a class is not lipid A reacti ve but binds to E. coli J5, an RcP+ mutant which carries an inner core structure common to many members of the family Enterobacteriaceae. Ph osphate groups present in the inner core contribute to the epitope but are not essential for the binding of WN1 222-5 to complete core LPS. Cross-reactivity for clinical bacterial isolates is broad. WN1 222-5 b inds to all E. coli clinical isolates tested so far (79 blood isolates , 80 urinary isolates, and 21 fecal isolates) and to some Citrobacter, Enterobacter, and Klebsiella isolates. This pattern of reactivity ind icates that its binding epitope is widespread among members of the Ent erobacteriaceae. WN1 222-5 exhibits biologically relevant activities. In vitro, it inhibits the Limulus amoebocyte lysate assay activity of S-form and R-form LPS in a dose-dependent manner and it neutralizes th e LPS-induced release of clinically relevant monokines (interleukin 6 and tumor necrosis factor). In vivo, WN1 222-5 blocks endotoxin-induce d pyrogenicity in rabbits and lethality in galactosamine-sensitized mi ce. The discovery of WN1 222-5 settles the long-lasting controversy ov er the existence of anti-core LPS MAbs with both cross-reactive and cr oss-protective activity, opening new possibilities for the immunothera py of sepsis caused by gram-negative bacteria.