Function characterization of a glyco-engineered anti-EGFR monoclonal antibody cetuximab in vitro

Aim： To evaluate the biochemical features and activities of a glyco-engineered form of the anti-human epidermal growth factor receptor monoclonal antibody （EGFR mAb） cetuximab in vitro. Methods： The genes encoding the Chinese hamster bisecting glycosylation enzyme （GnTIII） and anti-human EGFR mAb were cloned and coexpressed in CHO DG44 cells. The bisecting-glycosylated recombinant EGFR mAb （bisec-EGFR mAb） produced by these cells was characterized with regard to its glycan profile, antiproliferative activity, Fc receptor binding affinity and cell lysis capability. The content of galactose-α-1,3-galactose （α-Gal） in the bisec-EGFR mAb was measured using HPAEC-PAD. Results： The bisec-EGFR mAb had a higher content of bisecting N-acetylglucosamine residues. Compared to the wild type EGFR mAb, the bisec-EGFR mAb exhibited 3-fold higher cell lysJs capability in the antibody-dependent cellular cytotoxJcJty assay, and 1.36- fold higher antiproliferative activity against the human epidermoid carcinoma line A431. Furthermore, the bisec-EGFR mAb had a higher binding affinity for human FcyRla and FcyRIIla-158F than the wild type EGFR mAb. Moreover, α-Gal, which was responsible for cetuximab-induced hypersensitivity reactions, was not detected in the bisec-EGFR mAb. Conclusion： The glyco-engineered EGFR mAb with more bisecting modifications and lower α-Gal content than the approved therapeutic antibody Erbitux shows improved functionality in vitro, and requires in vivo validations.