title: CHO-gmt5, a novel CHO glycosylation mutant for producing afucosylated and asialylated recombinant antibodies.
abstract: Engineered zinc-finger nucleases (ZFNs) are powerful tools for creating double-stranded-breaks (DSBs) in genomic DNA in a site-specific manner. These DSBs generated by ZFNs can be repaired by homology-directed repair or nonhomologous end joining, in which the latter can be exploited to generate insertion or deletion mutants. Based on published literature, we designed a pair of zinc-finger nucleases and inactivated the GDP-fucose transporter gene (Slc35c1) in a previously reported CHO mutant that has a dysfunctional CMP-sialic acid transporter gene (Slc35a1). The resulting mutant cell line, CHO-gmt5, lacks functional GDP-fucose transporter and CMP-sialic acid transporter. As a result, these cells can only produce asialylated and afucosylated glycoproteins. It is now widely recognized that removal of the core fucose from the N-glycans attached to Asn(297) of human IgG1 significantly enhances its binding to its receptor, FcγRIIIa, and thereby dramatically improves antibody-dependent cellular cytotoxicity (ADCC). Recent reports showed that removal of sialic acid from IgG1 also enhances ADCC. Therefore, CHO-gmt5 may represent a more advantageous cell line for the production of recombinant antibodies with enhanced ADCC. These cells show comparable growth rate to wild type CHO-K1 cells and uncompromised transfection efficiency, which make them desirable for use as a production line.
authors.affiliation: Bioprocessing Technology Institute, Agency for Science, Technology and Research (A*STAR), Singapore.
authors.forename: Ryan
authors.initials: R
authors.lastname: Haryadi
authors.forename: Peiqing
authors.initials: P
authors.lastname: Zhang
authors.forename: Kah Fai
authors.initials: KF
authors.lastname: Chan
authors.forename: Zhiwei
authors.initials: Z
authors.lastname: Song
mesh_terms: D000818:Animals; D000906:Antibodies / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D016466:CHO Cells; D006224:Cricetinae; D005643:Fucose / Q000737:chemistry*; D006023:Glycoproteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D006031:Glycosylation; D009004:Monosaccharide Transport Proteins / Q000235:genetics / Q000378:metabolism; D019158:N-Acetylneuraminic Acid / Q000737:chemistry*; D033704:Nucleotide Transport Proteins / Q000235:genetics / Q000378:metabolism; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*
chemical_list: D000906:Antibodies; D006023:Glycoproteins; D009004:Monosaccharide Transport Proteins; D033704:Nucleotide Transport Proteins; D011994:Recombinant Proteins; C491081:SLC35A1 protein, human; C426650:SLC35C1 protein, human; D005643:Fucose; D019158:N-Acetylneuraminic Acid