The effect of differential glycosylation on human recombinant protein secretion, function and clearance

This project aims to investigate the effect of glycosylation on the secretion, physical properties, and functional biological and physiological properties of human recombinant proteins. In preferred expression systems, such as Chinese hamster ovary (CHO) cells, only about half of the genes involved in human glycan synthesis are expressed, leading to significant differences between CHO protein products and endogenous human glycoproteins, particularly in fucose, N-acetylglucosamine and sialic acid composition [1]. This differential glycosylation can affect secretion efficiency, solubility, functional activity, immuno-compatibility and in vivo circulatory half-life [2]. Human erythropoietin (EPO) comprises 165 amino acids with four glycosylation sites. The presence of glycosylation is essential for receptor binding and removal of sialic acid sugars results in rapid serum clearance and diminished in vivo efficacy. Recombinant EPO has been produced for decades (by CHO cells), and is approved for treating chronic anaemia and the side-effects of cancer therapy. The CHO-derived products contain branched and sialylated glycosylation but differ from endogenous EPO in several respects. While high-resolution crystal structures of EPO exist, e.g., bound to its receptor [3], they universally lack crucial information on glycosylation. Using our substantial experience in CHO expression and bioprocessing, and technologies for accurate 3D-shape analysis of glycosylation from extensive computer simulations [4], models will be produced for glycosylated endogenous EPO and recombinant EPO expressed by CHO cells. Algorithms will then be used to make predictions of aggregation propensity, solubility and secretion [5] in order to correlate glycosylation with CHO expression efficiency, and drive the design of host cells with specific glycosylation potential (to be experimentally tested in clonal and genome-engineered CHO cell variants). This research will help understand how glycosylation influences human recombinant protein production and efficacy, which, by extension to antibodies and novel format molecules has potential for substantial academic and industrial impact.

Contact for further Information:
[email protected]