title: Recombinant glycoproteins that inhibit complement activation and also bind the selectin adhesion molecules.
abstract: Soluble human complement receptor type 1 (sCR1, TP10) has been expressed in Chinese hamster ovary (CHO) DUKX-B11 cells and shown to inhibit the classical and alternative complement pathways in vitro and in vivo. A truncated version of sCR1 lacking the long homologous repeat-A domain (LHR-A) containing the C4b binding site has similarly been expressed and designated sCR1[desLHR-A]. sCR1[desLHR-A] was shown to be a selective inhibitor of the alternative complement pathway in vitro and to function in vivo. In this study, sCR1 and sCR1[desLHR-A] were expressed in CHO LEC11 cells with an active alpha(1,3)-fucosyltransferase, which makes possible the biosynthesis of the sialyl-Lewisx (sLex) tetrasaccharide (NeuNAcalpha2-3Galbeta1-4(Fucalpha1-3)GlcNAc) during post-translational glycosylation. The resulting glycoproteins, designated sCR1sLex and sCR1[desLHR-A]sLex, respectively, retained the complement regulatory activities of their DUKX B11 counterparts, which lack alpha(1-3)-fucose. Carbohydrate analysis of purified sCR1sLex and sCR1[desLHR-A]sLex indicated an average incorporation of 10 and 8 mol of sLex/mol of glycoprotein, respectively. sLex is a carbohydrate ligand for the selectin adhesion molecules. sCR1sLex was shown to specifically bind CHO cells expressing cell surface E-selectin. sCR1[desLHR-A]sLex inhibited the binding of the monocytic cell line U937 to human aortic endothelial cells, which had been activated with tumor necrosis factor-alpha to up-regulate the expression of E-selectin. sCR1sLex inhibited the binding of U937 cells to surface-adsorbed P-selectin-IgG. sCR1sLex and sCR1[desLHR-A]sLex have thus demonstrated both complement regulatory activity and the capacity to bind selectins and to inhibit selectin-mediated cell adhesion in vitro.
authors: Avant Immunotherapeutics, Inc., Needham, Massachusetts 02494-2725, USA.
authors: C W
authors: CW
authors: Rittershaus
authors: L J
authors: LJ
authors: Thomas
authors: D P
authors: DP
authors: Miller
authors: M D
authors: MD
authors: Picard
authors: K M
authors: KM
authors: Geoghegan-Barek
authors: S M
authors: SM
authors: Scesney
authors: L D
authors: LD
authors: Henry
authors: A C
authors: AC
authors: Sen
authors: A M
authors: AM
authors: Bertino
authors: G
authors: G
authors: Hannig
authors: H
authors: H
authors: Adari
authors: R A
authors: RA
authors: Mealey
authors: M L
authors: ML
authors: Gosselin
authors: M
authors: M
authors: Couto
authors: E G
authors: EG
authors: Hayman
authors: J L
authors: JL
authors: Levin
authors: V N
authors: VN
authors: Reinhold
authors: H C
authors: HC
authors: Marsh
mesh_terms: D000818:Animals; D015153:Blotting, Western; D016466:CHO Cells; D002448:Cell Adhesion; D003167:Complement Activation / Q000187:drug effects*; D006224:Cricetinae; D004586:Electrophoresis / Q000379:methods; D005434:Flow Cytometry; D006023:Glycoproteins / Q000737:chemistry / Q000494:pharmacology*; D006801:Humans; D013058:Mass Spectrometry; D009005:Monosaccharides / Q000032:analysis; D009844:Oligosaccharides / Q000032:analysis; D011485:Protein Binding; D011869:Radioligand Assay; D011994:Recombinant Proteins / Q000737:chemistry / Q000494:pharmacology; D019087:Selectins / Q000378:metabolism*; D020298:U937 Cells; D015854:Up-Regulation
chemical_list: D006023:Glycoproteins; D009005:Monosaccharides; D009844:Oligosaccharides; D011994:Recombinant Proteins; D019087:Selectins