Enhancing activity of FcαRI-bispecific antibodies using glycoengineering.

Macrophages and natural killer (NK) cells can effectively kill tumor cells in the presence of anti-cancer IgG monoclonal antibodies (mAbs), but neutrophils are less effective. We previously showed that IgG1 bispecific antibodies (BsAb), which target the IgA Fc receptor (FcαRI, CD89) and a tumor associated antigen induce effective neutrophil recruitment and tumor cell killing in vivo. Here we investigated if the efficacy of an anti-EGFR (CetuximAb)/FcαRI-bispecific antibody could be further improved by implementing glycoengineering of the IgG-Fc, aimed at increasing FcγRIIIa/b binding and/or complement activity. Fc afucosylation was introduced to enhance antibody-dependent cellular cytotoxicity (ADCC) by FcγRIIIa on NK/macrophages, which can also reduce neutrophil-mediated ADCC through their GPI-linked FcγRIIIb. Fc galactylation was found to enhance antibody hexamerization and thereby complement dependent cytotoxicity (CDC). Low fucosylated BsAbs moderately increased NK cell-mediated tumor cell killing, but did not affect neutrophil-mediated tumor cell killing nor phagocytosis by macrophages. Glycoengineering of these EGFR-specific BsAb, which normally are devoid of CDC-activity, did not enable their complement activities. In conclusion, glycoengineered FcαRI BsAbs increased ADCC by NK cells but had little effect on neutrophil or macrophage mediated tumor killing.