(597e) Evaluation and Characterization of Medium Factors Impacting the Fc Glycan Structure of Recombinant Antibodies Using Chemostat Cultures

Glycan structures attached to the CH2 domain of recombinant antibody produced by Chinese Hamster Ovary (CHO) cells have been shown to affect antibody effector function, clearance, and therapeutic efficacy. Hence, glycosylation heterogeneity on the Fc moiety is an important critical product attribute for commercially-manufactured antibodies.  During process development, multiple medium and process factors have been demonstrated to affect Fc glycan structures. However, these factors tend to be cell line specific and vary in their magnitude of effect. To develop a consistent Fc glycoform for a therapeutic antibody, a robust system to study and evaluate the impact of various factors on Fc glycosylation is necessary. A steady-state chemostat culture features constant specific cell growth rate, high viability, prolonged cell culture duration, and stable metabolite profiles, therefore, making it an ideal experimental tool for studying product quality of recombinant antibodies. In this study, a CHO chemostat producing recombinant antibody was operated for several months with high viability, and used to study the effects of various media and supplements on Fc glycosylation, including high mannose content and galactosylation percentage. NMR and mass spectroscopy were also used to characterize cell culture processes to correlate and elucidate the mechanisms of Fc glycan modulation. The results indicate that culture medium profoundly affects Fc glycan profile during antibody production. Individual supplements that impact protein glycosylation were also evaluated in the chemostat and their impact on Fc glycosyaltion were assessed. From this study, we demonstrate that chemostat culture can be a useful tool to complement cell culture process development by determining Fc product quality of a therapeutic antibody under varying cell culture medium and supplement conditions.