Craig G. Wilde

Endotoxin-binding and -neutralizing properties of recombinant bactericidal/permeability-increasing protein and monoclonal antibodies HA-1A and E5.

ObjectiveTo compare the endotoxin-binding and -neutralizing properties of bactericidal/permeability-increasing protein, the human monoclonal antiendotoxin antibody HA-1A, and the murine antiendotoxin antibody E5. DesignProspective, randomized, placebo-controlled laboratory study. SettingBiotechnology company research laboratory. SubjectsFemale CD-1 mice. InterventionsRecombinant bactericidal/permeability-increasing protein, HA-1A, a human immunoglobulin M monoclonal antibody raised against Escherichia coli J5 (Rc) endotoxin, and E5, a murine immunoglobulin M monoclonal antibody raised against E. coli J5 endotoxin, were compared in the following assays: a) binding to rough lipopolysaccharide immobilized onto microtiter plates; b) inhibition of lipopolysaccharide activity in the limulus amebocyte lysate assay; c) inhibition of lipopolysaccharide-induced cytokine release in whole blood; and d) protection against lethal endotoxin challenge in CD-1 mice. Measurements and Main ResultsThe binding affinity of bactericidal/permeability-increasing protein for immobilized lipopolysaccharide is apparently greater than the binding affinity of HA-1A or E5. Bactericidal/permeability-increasing protein neutralized lipopolysaccharide activity in the chromogenic limulus amebocyte lysate assay, while neither monoclonal antibody inhibited lipopolysaccharide activity. Similarly, bactericidal/permeability-increasing proteinreduced lipopolysaccharide-mediated tumor necrosis factor production in human whole blood in vitro, whereas monoclonal antibodies had slight (HA-1A) or no (E5) effect on lipopolysaccharide activity in this system. Administration of bactericidal/permeability-increasing protein gave >90% protection against an LDg0 dose of endotoxin in CD-1 mice, while treatment with HA-1A or E5 did not improve survival rate. ConclusionsNeither monoclonal antibody was as effective as bactericidal/permeability-increasing protein at binding or neutralizing endotoxin in vitro or in vivo. The potent endotoxin-binding and -neutralizing properties of bactericidal/permeability-increasing protein indicate that it might be useful in the treatment of endotoxin-related disorders in humans. (Crit Care Med 1994; 22:559–565)

Regulation of the Response to Bacterial Lipopolysaccharide by Endogenous and Exogenous Lipopolysaccharide Binding Proteins

Bactericidal/permeability-increasing protein (BPI) is a natural constituent of human neutrophils. Recombinant BPI has been shown to bind to bacterial lipopolysaccharide (LPS), and to neutralize the ability of LPS to stimulate inflammatory cells in vitro and in vivo. BPI shares sequence homology and immunocrossreactivity with another endogenous LPS binding protein, lipopolysaccharide binding protein (LBP). Despite the homology, these proteins have opposite effects on LPS. LBP mediates cell activation by low, otherwise nonstimulatory concentrations, while BPI neutralizes LPS bioactivity. Exogenous LPS binding proteins in the form of monoclonal antibodies have been developed with the goal of generating antiendotoxin therapeutics to treat gram-negative sepsis and related syndromes. Here we show that LPS-binding and neutralizing properties of BPI compare favorably with two monoclonal antibodies tested, HA-1A and XMMEN-OE5. BPI also competes effectively with LBP for LPS. Thus, BPI may represent an endogenous LPS-regulatory molecule suitable for use as a potent antiendotoxin therapeutic.