The production and characterisation of a chimaeric human IgE antibody, recognising the major mite allergen Der p 1, and its chimaeric human IgG1 anti-idiotype

Abstract

Background: Two mouse monoclonal antibodies have been described, namely: mAb 2C7 (IgG2bκ), which is directed against the major house dust mite allergen Der p 1, and mAb 2G10 (IgG1κ), which is an anti-idiotypic antibody raised against mAb 2C7. Given its broad IgE specificity, anti-idiotype mAb 2G10 could potentially have immunomodulatory applications. For example, a chimaeric human IgG version of mAb 2G10 could prove to be a useful molecule for binding to mast cell and basophil FceRI bound IgE, and in doing so co-ligating FceRI with FcγRIIB, which has been reported to have downregulatory effects. Aims: To produce a chimaeric human IgE version of mAb 2C7 (mAb 2C7huE) and a chimaeric human IgG1 version of its anti-idiotype mAb 2G10 (mAb 2G10huG1). Methods: The Vκ and VH regions of mAb 2C7 and its anti-idiotype mAb 2G10 were engineered into human constant regions of the IgE and IgG1 isotypes, respectively. Results: The production of chimaeric mAb 2C7huE and its anti-idiotype mAb 2G10huG1 confirmed that the respective mouse antibody V regions were successfully engineered into human constant regions and still retained the specificity of the original murine V regions. Conclusion: The newly constructed chimaeric antibodies will be useful to investigate the downregulation of IgE mediated hypersensitivity by the crosslinking of FceRI with FcγRIIB.

Generation of Human Monoclonal Allergen-Specific IgE and IgG Antibodies from Synthetic Antibody Libraries

“…Chimerization of animal-derived antibodies by addition of desired human constant regions can open alternatives but is very time-consuming and is not specific for allergenic epitopes. Nonetheless, the recloning of murine hybridomas to chimeric IgE has been reported (31 ). Indeed, we used a murine antibody fragment with specificity for Gal d4 to express recombinant human Fc antibodies.…”

An immunoglobulin E‐reactive chimeric human immunoglobulin G1 anti‐idiotype inhibits basophil degranulation through cross‐linking of FcɛRI with FcγRIIb

“…The chimerization of the original mouse 2G10 antibody, an anti-idiotype with broad human IgE specificity, to form an antibody with a human IgG1 constant region [15] was driven by the expectation that such a construct may prove to be therapeutically useful. Thus, a human version of 2G10 will not only bind to FceRI-bound IgE, but should also bind to the inhibitory receptor FcgRIIb, thereby crosslinking FceRI with FcgRIIb and in doing so inhibit mast cell and basophil activation [28].…”

“…Chimeric human IgG1 2G10 (k light chain) was engineered and characterized as reported previously [15]. The original mouse IgG1 (k light chain) version of 2G10 was prepared and fully characterized as described elsewhere [17,18].…”

“…Antibody 2G10, an anti-idiotype raised against 2C7, binds IgE regardless of its allergen specificity and is therefore thought to recognize a common IgE framework (Fw) region, rather than complementarity-determining regions (CDRs) [17,18]. Owing to its ability to bind to IgE, via its Fab regions, and FcgRIIb, via its Fc region, 2G10 has the potential to cross-link FceRI-bound IgE with FcgRIIb and in doing so could down-regulate cellular triggering [15]. Here, we demonstrate, using cell surface expression of CD63 [19] and intracellular IL-4 detection, that human 2G10 inhibits anti-IgE and allergen-driven basophil activation.…”

“…In a previous work, we described the production of two allergy-related monoclonal antibodies by conventional mouse immunization and chimerization to form antibodies with human IgE (2C7) and IgG1 (2G10) constant regions [15]. Antibody 2C7 is directed against the major mite allergen Der p 1 and is representative of a significant component of the human IgE anti-Der p 1 specificity [16].…”