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Optimisation of the Cellular Metabolism of Glycosylation for Recombinant Proteins Produced by Mammalian Cell Systems

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Abstract

Many biopharmaceuticals are now produced as secreted glycoproteins from mammalian cell culture. The glycosylation profile of these proteins is essential to ensure structural stability and biological and clinical activity. However, the ability to control the glycosylation is limited by our understanding of the parameters that affect the heterogeneity of added glycan structures. It is clear that the glycosylation process is affected by a number of factors including the 3-dimensional structure of the protein, the enzyme repertoire of the host cell, the transit time in the Golgi and the availability of intracellular sugar-nucleotide donors. From a process development perspective there are many culture parameters that can be controlled to enable a consistent glycosylation profile to emerge from each batch culture. A further, but more difficult goal is to control the culture conditions to enable the enrichment of specific glycoforms identified with desirable biological activities. The purpose of this paper is to discuss the cellular metabolism associated with protein glycosylation and review the attempts to manipulate, control or engineer this metabolism to allow the expression of human glycosylation profiles in producer lines such as genetically engineered Chinese hamster ovary (CHO) cells.

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Abbreviations

ADCC:

antibody-mediated cytotoxicity

BHK:

baby hamster kidney (cells)

C2GnT:

core 2 GlcNAC transferase (UDP-GlcNAc: Galβ1,3GalNAc-R β1,6-N-acetyl glucosaminyl transferase)

CHO:

Chinese hamster ovary (cells)

CMP:

cytidine monophosphate

DO:

dissolved oxygen

EPO:

erythropoietin

ER:

endoplasmic reticulum

FT:

fucosyl transferase

G0:

agalactosylated glycans

G1:

monogalactosylated glycans

G2:

digalactosylated glycans

GalNAc:

N-acetyl galactosamine

GDM:

GDP mannose 4,6 dehydratase

GDP:

guanosine diphosphate

GFAT:

glutamine: fructose 6-phosphate amidotransferase

GlcNAc:

N-acetyl glucosamine

GnT:

N-acetyl glucosaminyl transferase

HIV:

human immunodeficiency virus

IFN:

interferon

IgG:

immunoglobulin

LAMP:

lysosomal membrane glycoprotein

ManNAc:

N-acetyl mannosamine

mPL-I:

mouse placental lactogen I

NANA:

N-acetyl-neuraminic acid

NGNA:

N-glycolyl-neuraminic acid

OST:

oligosaccharyltransferase

ST:

sialyl transferase

ST3Gal1:

sialyl transferase 3 (CMP-sialic acid: Galβ1,3GalNAc2,3 sialyl transferase)

TIMP:

tissue inhibitors of metalloproteinases

t-PA:

tissue plasminogen activator

UDP:

uridine diphosphate

UTP:

uridine triphosphate

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  1. Department of Microbiology, University of Manitoba, R3T 2N2, Winnipeg, Manitoba, Canada

    M. Butler

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Butler, M. Optimisation of the Cellular Metabolism of Glycosylation for Recombinant Proteins Produced by Mammalian Cell Systems. Cytotechnology 50, 57–76 (2006). https://doi.org/10.1007/s10616-005-4537-x

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