BioEngineering

Jason Hodoniczky

Modification of monoclonal IgG Fc N-glycans in vitro for increased recombinant product bioactivity

Binding of the Fc region of IgG molecules to Fc? receptors triggers important effector functions in the immune system, linking the humoral and the cellular aspects of the immune response. High affinity (Fc?RI) and low affinity (Fc?RII and Fc?RIII) IgG receptors mediate a number of responses such as endocytosis, phagocytosis, antibody-dependent cell-mediated cytotoxicity (ADCC) and cytokine release, depending on the Fc?R isoform and the cell type on which it is expressed. It is now known that the N-glycans at Asn297 in the Fc domain of an IgG molecule are absolutely required for FcR binding and thus biological activity. Moreover, changes in Fc N-glycan structure can play a significant role in modulating bioactivity. With respect to the latter, a range of differently sialylated and galactosylated antibodies have been shown to differ in their biological activities in a variety of studies. However, two specific N-glycan modifications; a bisecting GlcNAc conferred by N-acetylglucosaminyl transferase III (GnT-III) and a core fucose catalysed by ?1,6-fucosyltransferase, have recently been shown to be of crucial significance for IgG biological activity. These modifications have been shown to be particularly important for antibody therapeutics whose mode of action depends on effector functions such as ADCC. One recent study, deriving from IDEC Pharmaceuticals, has showed that recombinant IgG1 antibodies produced in CHO cells co-expressing recombinant GnT-III were almost entirely substituted with a bisecting GlcNAc. These Mab’s exhibited significantly elevated ADCC activity mediated via an increased affinity for Fc?RIII. Similarly in an unpublished study by RL Shields et al, core fucose deficient IgG1 molecules also showed enhanced ADCC activity.

The potential significance of the above experimental observations are the development of Mab therapeutics with increased specific bioactivity. This will substantially reduce dose and thus require smaller scale manufacturing capacity and/or increase throughput. This can be achieved by production of Mab’s with elevated bisecting GlcNAc and decreased fucose content.

Therefore, in collaboration with Neose Technologies the major aim of the project is to develop a process for in vitro substitution of recombinant monoclonal antibody N-glycans at Asn297 with a bisecting GlcNAc and without a core fucose.

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Personal

Having spent my schooling years in the northern suburbs of Perth, I continued my education at the University of Western Australia. I completed my Bachelor of Science with majors in Chemistry and Microbiology in 1999 and continued at UWA the following year, achieving First Class Honours in Chemistry. My project was supervised by Jackie Wilce and was entitled “The Intracellular Delivery of Therapeutic Molecules Using Carrier Peptides.” I continued working in the Wilce lab until being employed as a Peptide Chemist at Xenome Ltd in 2001 here in Brisbane. Currently I am completing my PhD at the University of QLD, having commenced June 2002.