title: The anti-lipid A monoclonal antibody E5 binds to rough gram-negative bacteria, fixes C3, and facilitates binding of bacterial immune complexes to both erythrocytes and monocytes.
abstract: Treatment of patients with septic shock using monoclonal antibodies (mAbs) to endotoxin is still controversial. Clinical trials of E5, one of the mAbs directed against the lipid A moiety of lipopolysaccharide (LPS), are currently in progress. The mechanisms of action of this, and other antibodies under clinical evaluation, are, however, poorly understood. In this study we examined in vitro the ways in which E5 interacted with Gram-negative bacteria, complement, erythrocytes and monocytes. By fluorescence-activated cell sorter (FACS) analysis we showed direct, dose-dependent binding of E5 to Escherichia coli (E. coli) and Salmonella minnesota (S. minnesota). Antibody binding to S. minnesota was enhanced by treatment with the beta-lactam antibiotic amoxycillin, but not by treatment with the aminoglycoside gentamicin. Immune complexes formed between E5 and both species of Gram-negative bacteria activated both classical and alternative complement pathways, but only in the case of S. minnesota did this facilitate binding to erythrocyte CR1 and monocyte CR3. Bacterial C3b and iC3b fixation by E5 was quantified using specific mAbs. These observations suggest that E5 may enhance bacterial clearance in several ways: (1) by facilitating direct complement fixation; (2) by facilitating the binding of opsonized bacteria to cells of the mononuclear phagocyte system; (3) by enabling bacteria to bind to erythrocyte CR1 (CD35), allowing safe carriage in the circulation to the fixed macrophages of the liver and spleen; (4) by acting synergistically with beta-lactam antibiotics.
authors.affiliation: Department of Medicine, Royal Postgraduate Medical School, Hammersmith Hospital, London, UK.
authors.forename: M A
authors.initials: MA
authors.lastname: Seelen
authors.forename: P
authors.initials: P
authors.lastname: Athanassiou
authors.forename: W A
authors.initials: WA
authors.lastname: Lynn
authors.forename: P
authors.initials: P
authors.lastname: Norsworthy
authors.forename: M J
authors.initials: MJ
authors.lastname: Walport
authors.forename: J
authors.initials: J
authors.lastname: Cohen
authors.forename: K A
authors.initials: KA
authors.lastname: Davies
mesh_terms: D000900:Anti-Bacterial Agents / Q000494:pharmacology; D000911:Antibodies, Monoclonal / Q000378:metabolism*; D000936:Antigen-Antibody Complex / Q000378:metabolism*; D003179:Complement C3b / Q000378:metabolism; D004912:Erythrocytes / Q000378:metabolism; D004926:Escherichia coli / Q000187:drug effects / Q000378:metabolism*; D005455:Fluorescent Antibody Technique; D006801:Humans; D007769:Lactams; D008050:Lipid A / Q000276:immunology*; D009000:Monocytes / Q000378:metabolism; D017463:Receptors, Complement 3b / Q000378:metabolism; D012475:Salmonella / Q000187:drug effects / Q000378:metabolism*
chemical_list: D000900:Anti-Bacterial Agents; D000911:Antibodies, Monoclonal; D000936:Antigen-Antibody Complex; D007769:Lactams; D008050:Lipid A; D017463:Receptors, Complement 3b; D003179:Complement C3b