Modulation of high mannose levels in N ‐linked glycosylation through cell culture process conditions to increase antibody‐dependent cell‐mediated cytotoxicity activity for an antibody biosimilar

Abstract The regulatory approval of a biosimilar product is contingent on the favorable comparability of its safety and efficacy to that of the innovator product. As such, it is important to match the critical quality attributes of the biosimilar product to that of the innovator product. The N ‐glycosylation profile of a monoclonal antibody (mAb) can influence effector function activities such as antibody‐dependent cell‐mediated cytotoxicity (ADCC) and complement‐dependent cytotoxicity. In this study, we describe efforts to modulate the high‐mannose (HM) levels of a biosimilar mAb produced in a Chinese hamster ovary cell fed‐batch process. Because the HM level of the mAb was observed to impact ADCC activity, it was desirable to match it to the innovator mAb's levels. Several cell culture process related factors known to modulate the HM content of N ‐glycosylation were investigated, including osmolality, ammonium chloride (NH 4 Cl) addition, glutamine concentration, monensin addition, and the addition of alternate sugars and amino sugars to the feed medium. The process conditions evaluated varied in impact on HM levels, process performance and product quality. One condition, the addition of alternate sugars and amino sugars to feed medium, was identified as the preferred method for increasing HM levels with minimal disruptions to process performance or other product quality attributes. Interestingly, a secondary interaction between sugar and amino sugar supplemented feeds and osmolality was observed during process scale‐up. These studies demonstrate sugar and amino sugar concentrations and osmolality are critical variables to evaluate to match HM content in biosimilar and their innovator mAbs.