Maintaining high sialic acid content on glycoproteins intended for therapeutic uses is important for the biotechnology industry because desialyated glycoproteins exhibit reduced activity, stability, serum half-life, and immunogenicity. In addition, variably sialyated glycoproteins complicate purification and formulation of the glycoprotein product. During recombinant protein expression in Chinese Hamster Ovary (CHO) cells, extracellular sialidases are responsible for cleaving the O-glycosidic bond between sialic acid and other sugars. This enzymatic cleavage can be prevented by preparation of carbon or sulfur glycoside competitive inhibitors. We aim to synthesize and screen a library of carbon and sulfur glycosidic sialidase inhibitors bound to a water compatible resin such as PEGA. Hydrophobic interactions of the resin should increase binding affinity to the sialidase. Attachment of a resin will simplify removal of the inhibitor from the glycoprotein product, prevent uptake into the cell during protein expression, and allow for possible multivalent interactions.

Existing inhibitors, commonly used in treatment of influenza, are not cost effective for large scale operations, therefore we will utilize inexpensive metabolically engineered sialic acid as starting material to synthesize resin bound glycosides. Inhibitors will be screened against bacterial sialidases in vitro and Ki values will be determined. The best inhibitors will be tested against authentic CHO cell sialidases for evaluation of application in CHO cell protein expression cultures. Implementation of this on an industrial scale could significantly decrease cost of production as well as improve the protein product giving pharmacokinetic profile with reduced side effects.