Engineering Biopharmaceuticals - An overall increasing proportion of future product approvals will be engineered in some way, either directly or indirectly. - BioPharm International

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Engineering Biopharmaceuticals
An overall increasing proportion of future product approvals will be engineered in some way, either directly or indirectly.


BioPharm International
Volume 20, Issue 11

One avenue of current research relates to engineering the glycosylation pathways in alternative potential production systems with a view to humanize the glycosylation patterns characteristic of the proteins they produce. Such engineering is not an insubstantial task, because the exact glycosylation pathways present may not be completely characterized, and the glycosylation reactions themselves occur in a specific predetermined sequence spanning both the ER and Golgi. This necessitates the targeting of human glycosylation reactions to specific organelles.

Despite the difficulties, substantial progress has been recorded, particularly in the case of yeast-based systems. For example, engineered strains of Pichia pastoris have been developed which undertake specific human glycosylation reactions, and there are yeast cell lines now in existence that can actually produce more uniform glycoform profiles than do currently used mammalian cell lines.5,6

Engineering the PTM capacity of a producer cell line to tailor the PTM characteristics of therapeutic proteins produced can also be done for other reasons, e.g., in order to optimize a particular biological activity of the target protein, as exemplified by some recent antibody research.

ANTIBODY PTM ENGINEERING


Figure 1. The structure of IgG highlighting the oligosaccharide component. Refer to text for detail. Reproduced from reference 7.
Overall, the oligosaccharide component of immunoglobulin G (IgG) accounts only for 2–3% of its mass. Intact antibodies display a characteristic oligosaccharide side chain attached by asparagine 297, present in the antibody Fc region (Figure 1).7 This oligosaccharide plays an indirect but vital role in triggering antibody effector functions, such as antibody dependent cell-mediated cytotoxicity (ADCC), which is believed to be a primary mechanism by which several therapeutic antibodies, e.g., Herceptin (Genentech, South San Francisco, CA), bring about their therapeutic effect. Recent results illustrate that remodeling of the antibody glycocomponent can have a substantial effect on the exact profile and strength of effector functions triggered. It has been demonstrated that the removal of fucose residues from the antibody sugar side chain can increase ADCC activity8 and an engineered CHO cell line has been developed in which the fucosylating enzyme has been knocked out. This facilitates the production of completely defucosylated IgG which displays up to a 100-fold increase in ADCC activity.

CONCLUSIONS

An understanding of protein structure–function relationship coupled with the continued development and refinement of molecular techniques allowing the alteration of protein amino acid sequence or PTM complement provides increasing scope for the engineering of biopharmaceuticals to optimize their therapeutic usefulness. Therefore, an overall increasing proportion of future approvals will be engineered in some way, either directly by site directed mutagenesis, or indirectly via the development of engineered producer cell lines capable of producing therapeutic proteins displaying an optimized PTM profile.

Gary Walsh, PhD, is an associate professor in the Industrial Biochemistry Program at the University of Limerick, Limerick City, Ireland, +353.61.202664,

REFERENCES

1. Walsh, G. Second-generation biopharmaceuticals. Eur. J Pharm. Biopharm. 2004;58:185–196

2. Walsh, G. and Jefferis, R. Post-translational modifications in the context of therapeutic proteins. Nature Biotechnol. 2006;24:1241–1252

3. Kobata, A. Structure and function of the sugar chains of glycoproteins. Eur. J Biochem. 1992;209:483–501

4. Dwek, R., Butters, T. Platt, F., Zitzmann, N. Targeting glycosylation as a therapeutic approach. Nature Rev. Drug Discov. 2002;1:65–75

5. Wildt, S. and Gerngross, T. The humanization of N-glycosylation pathways in yeast. Nat. Rev. Microbiol. 2005;3:119–128.


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