Cross-reactivity of monoclonal antibodies and sera directed against lipid A and lipopolysaccharides

Abstract

The cross-reactive capacity of monoclonal and polyclonal lipid A antibodies was tested with rough and smooth lipopolysaccharides (LPS). The antibodies represented different specificities recognizing epitopes in the hydrophilic lipid A backbone. In none of the various assay systems applied did the antibodies react with complete rough or smooth-form LPS. Cross-reactions, in general, were only detected with the most rudimentary rough LPS tested, i.e. Re-LPS. A variety of reactivities with other LPS was shown not to be related to lipid A antibodies; such reactivities were present in rabbit sera as well as in crude ascites. These results underline the need for careful checks on the origin of reactivities observed. In addition, rabbit antisera raised with R- and S-LPS were screened for lipid A reactivity using synthetic lipid A and partial structures as antigens. No cross-reactivity of LPS antibodies with lipid A was detected in these sera.

Structural Basis for Antibody Recognition of Lipid A

“…However, S1-15 showed stronger binding to the B4P-conjugate confirming the weaker dependence on the recognition of the phosphate at the anomeric position (25). Binding data against synthetic lipid A and derivatives thereof have been reported earlier (15,25,29).…”

“…Although lipid A structure is relatively conserved among pathogenic species, none of the numerous reported antibodies claimed to be specific for lipid A have led to successful clinical implementation (13)(14)(15)25). Specific binding to lipid A was observed upon acid treatment of bacterial LPS, thereby liberating the lipid A fragment acting then as neoantigen, when embedded into erythrocytes or liposomes (25,26).…”

The Combining Sites of Anti-lipid A Antibodies Reveal a Widely Utilized Motif Specific for Negatively Charged Groups

“…Antibodies believed to be specific for lipid A were first observed during immunization with acid-treated bacterial LPS, where the liberated lipid A fragment can act as a neoantigen when embedded into erythrocytes or liposomes (30,32). Despite numerous reports of antibodies shown to be specific for lipid A, none have led to successful clinical implementation (17)(18)(19)30).Recently, the structure of antigen-binding fragments (Fabs) 4 from monoclonal antibodies (mAbs) A6 (IgG2b) (33) and S1-15 (IgG2b), also referred to as S1 (30), were determined both in complex with lipid A and in the unliganded form to high resolution (34). The structures provided a structural basis for the observed failure of anti-lipid A antibodies to bind intact LPS, as the free hydroxyl on the ␤-glucosamine C-6 attachment point for LPS inner core residues (35) was observed to be buried in the antibody-combining site.…”

“…Although antibodies specific for the various LPS components have been reported (15)(16)(17)(18)(19)(20)(21)(22)(23)(24), the structural variation in the core and O-polysaccharide regions together with the rapid onset of septic shock have hindered their introduction into clinical use (4,12,(25)(26)(27). To date, only the inner core binding mAb WN1 222-5 has been reported successful in neutralizing a wide range of Gram-negative bacteria, including Escherichia, Salmonella, Shigella, and Citrobacter (15,28,29).…”

“…Antibodies believed to be specific for lipid A were first observed during immunization with acid-treated bacterial LPS, where the liberated lipid A fragment can act as a neoantigen when embedded into erythrocytes or liposomes (30,32). Despite numerous reports of antibodies shown to be specific for lipid A, none have led to successful clinical implementation (17)(18)(19)30).…”