Engineering Chinese hamster ovary cells to maximize effector function of produced antibodies using FUT8 siRNA.
We explored the possibility of converting established antibody-producing cells to cells producing high antibody-dependent cellular cytotoxicity (ADCC) antibodies. The conversion was made by constitutive expression of small interfering RNA (siRNA) against alpha1,6 fucosyltransferase (FUT8). We found two effective siRNAs, which reduce FUT8 mRNA expression to 20% when introduced into Chinese hamster ovary (CHO)/DG44 cells. Selection for Lens culinaris agglutinin (LCA)-resistant clones after introduction of the FUT8 siRNA expression plasmids yields clones producing highly defucosylated (approximately 60%) antibody with over 100-fold higher ADCC compared to antibody produced by the parental cells (approximately 10% defucosylated). Moreover, the selected clones remain stable, producing defucosylated antibody even in serum-free fed-batch culture. Our results demonstrate that constitutive FUT8 siRNA expression can control the oligosaccharide structure of recombinant antibody produced by CHO cells to yield antibodies with dramatically enhanced ADCC.
Mori
Katsuhiro
K
Tokyo Research Laboratories, Kyowa Hakko Kogyo Co., Ltd., 3-6-6 Asahi-machi, Machida-shi, Tokyo 194-8533, Japan.
Kuni-Kamochi
Reiko
R
Yamane-Ohnuki
Naoko
N
Wakitani
Masako
M
Yamano
Kazuya
K
Imai
Harue
H
Kanda
Yutaka
Y
Niwa
Rinpei
R
Iida
Shigeru
S
Uchida
Kazuhisa
K
Shitara
Kenya
K
Satoh
Mitsuo
M
D000818:Animals; D000911:Antibodies, Monoclonal / Q000235:genetics* / Q000378:metabolism*; D016466:CHO Cells; D003001:Cloning, Molecular / Q000379:methods*; D006224:Cricetinae; D003412:Cricetulus; D005647:Fucosyltransferases / Q000235:genetics* / Q000378:metabolism*; D020868:Gene Silencing / Q000502:physiology; D024861:Genetic Enhancement / Q000379:methods*; D015202:Protein Engineering / Q000379:methods*; D034741:RNA, Small Interfering / Q000235:genetics
D000911:Antibodies, Monoclonal; D034741:RNA, Small Interfering; D005647:Fucosyltransferases; C020559:Glycoprotein 6-alpha-L-fucosyltransferase