Sialic acid levels of therapeutic glycoprotein play an important role in plasma half-life. An undesirable decrease of sialic acid content was observed when we increased Fc-fusion protein productivity fourfold in a GS-CHO cell line by bioprocess optimization. We investigated the potential mechanism for the sialic acid content reduction. We found that limited nucleotide sugar precursor and the extracellular sialidase were not responsible for the reduction of the sialic acid content after titer improvement. Oligosaccharide analysis revealed that the lack of protein galactosylation was the potential cause for the reduction of sialic acid content. Thus we validated this notion by evaluated galactose supplementation in 2 L bioreactors. Cell culture performance was not impacted by addition of up to 40 mM galactose except for the glucose consumption rate. Addition of 20 mM galactose to the bioreactor resulted in the increase of 44 % for total sialic acid content and 20.3 % for sialylated glycans. These data were further validated when the process was run on 200 L scaled bioreactor. These data together show that the galactosylation plays an apparent role in sialylation in our current system.

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Long-term safety and efficacy of biosimilars are approved by using abbreviated methods and clinical studies may not always adequate to ensure comparability of biosimilar mAbs comparing to its reference product. Therefore, the analytical strategy of the physicochemical comparison of a biosimilar to its reference product becomes an important data to indicate clinical similarity in safety and efficacy. FDA recommended that demonstration of biosimilarity between reference and biosimilar versions is based upon data derived from analytical studies to show "high similarity" to the reference product not withstanding minor differences in clinically inactive components. Therefore, the physicochemical analytical comparison between biosimilar and its reference product is the primary consideration during biosimilar development. In these review, the approach for physicochemical characterization, biological activity and impurities assessment were reviewed.

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