title: Glycan structure and serum half-life of recombinant CTLA4Ig, an immunosuppressive agent, expressed in suspension-cultured rice cells with coexpression of human β1,4-galactosyltransferase and human CTLA4Ig.
abstract: Human cytotoxic T-lymphocyte antigen 4-immunoglobulin (hCTLA4Ig) is an immunosuppressive therapeutic, and recently produced rice cell-derived hCTLA4Ig (hCTLA4Ig(P)) reportedly exhibits in vitro immunosuppressive activities equivalent to those of Chinese hamster ovary cell-derived hCTLA4Ig (hCTLA4Ig(M)). However, limitations of hCTLA4Ig(P) include shortened in vivo half-life as well as the presence of nonhuman N-glycans containing (β1-2)-xylose and α1,3-fucose, which cause immunogenic reactions in humans. In the present study, human β1,4-galactose-extended hCTLA4Ig(P) (hCTLA4Ig(P)-Gal) was expressed through the coexpression of human β1,4-galactosyltransferase (hGalT) and hCTLA4Ig in an attempt to overcome these unfavorable effects. The results indicated that both encoding hGalT and hCTLA4Ig were successfully coexpressed, and the analysis of N-glycan and its relative abundance in purified hCTLA4Ig(P)-Gal indicated that not only were the two glycans containing (β1-4)-galactose newly extended, but also glycans containing both β1,2-xylose and α1,3-fucose were markedly reduced and high-mannose-type glycans were increased compared to those of hCTLA4Ig(P), respectively. Unlike hCTLA4Ig(P), hCTLA4Ig(P)-Gal was effective as an acceptor via (β1-4)-galactose for in vitro sialylation. Additionally, the serum half-life of intravenously injected hCTLA4Ig(P)-Gal in Sprague-Dawley rats was 1.9 times longer than that of hCTLA4Ig(P), and the clearance pattern of hCTLA4Ig(P)-Gal was close to that for hCTLA4Ig(M). These results indicate that the coexpression with hGalT and hCTLA4Ig(P) is useful for both reducing glycan immunogens and increasing in vivo stability. This is the first report of hCTLA4Ig as an effective therapeutics candidate in glycoengineered rice cells.
authors.affiliation: Boryung Central Research Institute, Boryung Pharmaceutical Co. Ltd., 1122-3, Shingil-dong, Danwon-gu, Ansan-si, Kyungki-do, 425-839, Republic of Korea.
authors.forename: Seung Hoon
authors.initials: SH
authors.lastname: Kang
authors.forename: Hahn Sun
authors.initials: HS
authors.lastname: Jung
authors.forename: Song Jae
authors.initials: SJ
authors.lastname: Lee
authors.forename: Cheon Ik
authors.initials: CI
authors.lastname: Park
authors.forename: Sang Min
authors.initials: SM
authors.lastname: Lim
authors.forename: Heajin
authors.initials: H
authors.lastname: Park
authors.forename: Byung Sun
authors.initials: BS
authors.lastname: Kim
authors.forename: Kwang Heum
authors.initials: KH
authors.lastname: Na
authors.forename: Gyeong Jin
authors.initials: GJ
authors.lastname: Han
authors.forename: Jae Woo
authors.initials: JW
authors.lastname: Bae
authors.forename: Hyun Joo
authors.initials: HJ
authors.lastname: Park
authors.forename: Keuk Chan
authors.initials: KC
authors.lastname: Bang
authors.forename: Byung Tae
authors.initials: BT
authors.lastname: Park
authors.forename: Hye Seong
authors.initials: HS
authors.lastname: Hwang
authors.forename: In-Soo
authors.initials: IS
authors.lastname: Jung
authors.forename: Jae Il
authors.initials: JI
authors.lastname: Kim
authors.forename: Doo Byung
authors.initials: DB
authors.lastname: Oh
authors.forename: Dong Il
authors.initials: DI
authors.lastname: Kim
authors.forename: Hirokazu
authors.initials: H
authors.lastname: Yagi
authors.forename: Koichi
authors.initials: K
authors.lastname: Kato
authors.forename: Dae Kyong
authors.initials: DK
authors.lastname: Kim
authors.forename: Ha Hyung
authors.initials: HH
authors.lastname: Kim
mesh_terms: D000069594:Abatacept / Q000097:blood / Q000737:chemistry*; D000818:Animals; D016466:CHO Cells; D002240:Carbohydrate Sequence; D018929:Cell Culture Techniques / Q000379:methods; D003412:Cricetulus; D005700:Galactosyltransferases / Q000235:genetics* / Q000378:metabolism; D006207:Half-Life; D006801:Humans; D007166:Immunosuppressive Agents / Q000097:blood / Q000493:pharmacokinetics*; D008297:Male; D008969:Molecular Sequence Data; D012275:Oryza / Q000166:cytology / Q000235:genetics*; D030821:Plants, Genetically Modified; D011134:Polysaccharides / Q000737:chemistry*; D017207:Rats, Sprague-Dawley; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism
chemical_list: D007166:Immunosuppressive Agents; D011134:Polysaccharides; D011993:Recombinant Fusion Proteins; D000069594:Abatacept; D005700:Galactosyltransferases; C493310:beta1,4-galactosyltransferase, human