Monoclonal antibodies (mAbs) are an important class of therapeutic glycoproteins widely used for the treatment of cancer, inflammation, and infectious diseases. Recent studies have demonstrated that the structures of Fc glycans can dictate the immunological functions, e.g., pro-inflammatory (activation) vs. anti-inflammatory activity, of a given antibody. Therefore, controlling glycosylation of a therapeutic monoclonal antibody to a desired glycosylation status is critical for maximizing its therapeutic efficacy. However, most monoclonal antibodies are produced as mixtures of glycoforms that are not optimal for efficacy, and preparation of homogeneously glycosylated mAbs has been a challenging task. In this presentation, we describe a chemoenzymatic method that enables site-specific glycosylation remodeling of both Fc and Fab glycans in a monoclonal antibody. Using cetuximab, an mAb that carries both Fab and Fc glycans, as an example, we have shown that a remarkable site-specific glycosylation remodeling of both Fc and Fab glycans can be achieved by taking advantage of the substrate specificity of the deglycosylation and transglycosylation enzymes. The glycoengineered rituximab, with a fully sialylated glycan at the Fab domain, and a non-fucosylated N-glycan at the Fc domain, showed comparable antigen binding affinity as the commercial mAb, but its antibody-dependent cellular cytotoxicity was significantly enhanced. The scope and limitation of the chemoenzymatic approach to glycoengineering of mAbs will be discussed.

Site-specific chemoenzymatic glycoengineering of therapeutic antibodies

249th National Meeting (2015)

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