Engineering Chinese hamster ovary cells to maximize sialic acid content of recombinant glycoproteins.
We have engineered two Chinese hamster ovary cell lines secreting different recombinant glycoproteins to express high levels of human beta1,4-galactosyltransferase (GT, E.C. 2.4.1.38) and/or alpha2, 3-sialyltransferase (ST, E.C. 2.4.99.6). N-linked oligosaccharide structures synthesized by cells overexpressing the glycosyltransferases showed greater homogeneity compared with control cell lines. When GT was overexpressed, oligosaccharides terminating with GlcNAc were significantly reduced compared with controls, whereas overexpression of ST resulted in sialylation of >/=90% of available branches. As expected, GT overexpression resulted in reduction of oligosaccharides terminating with GlcNAc, whereas overexpression of ST resulted in sialylation of >/=90% of available branches. The more highly sialylated glycoproteins had a significantly longer mean residence time in a rabbit model of pharmacokinetics. These experiments demonstrate the feasibility of genetically engineering cell lines to produce therapeutics with desired glycosylation patterns.
Weikert
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S
Department of Cell Culture and Fermentation Research and Development, Genentech, Inc., South San Francisco, CA 94080, USA.
Papac
D
D
Briggs
J
J
Cowfer
D
D
Tom
S
S
Gawlitzek
M
M
Lofgren
J
J
Mehta
S
S
Chisholm
V
V
Modi
N
N
Eppler
S
S
Carroll
K
K
Chamow
S
S
Peers
D
D
Berman
P
P
Krummen
L
L
D000818:Animals; D016466:CHO Cells; D006224:Cricetinae; D005700:Galactosyltransferases / Q000378:metabolism; D005818:Genetic Engineering; D006023:Glycoproteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D019158:N-Acetylneuraminic Acid / Q000737:chemistry* / Q000378:metabolism*; D011817:Rabbits; D011994:Recombinant Proteins / Q000737:chemistry* / Q000378:metabolism; D012799:Sialyltransferases / Q000378:metabolism
D006023:Glycoproteins; D011994:Recombinant Proteins; D005700:Galactosyltransferases; D012799:Sialyltransferases; D019158:N-Acetylneuraminic Acid