Recombinant Glycoproteins That Inhibit Complement Activation and Also Bind the Selectin Adhesion Molecules

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 α(1,3)-fucosyltransferase, which makes possible the biosynthesis of the sialyl-Lewis x (sLe x ) tetrasaccharide (NeuNAcα2–3Galβ1–4(Fucα1–3)GlcNAc) during post-translational glycosylation. The resulting glycoproteins, designated sCR1sLe x and sCR1[desLHR-A]sLe x , respectively, retained the complement regulatory activities of their DUKX B11 counterparts, which lack α(1–3)-fucose. Carbohydrate analysis of purified sCR1sLe x and sCR1[desLHR-A]sLe x indicated an average incorporation of 10 and 8 mol of sLe x /mol of glycoprotein, respectively. sLe x is a carbohydrate ligand for the selectin adhesion molecules. sCR1sLe x was shown to specifically bind CHO cells expressing cell surface E-selectin. sCR1[desLHR-A]sLe x inhibited the binding of the monocytic cell line U937 to human aortic endothelial cells, which had been activated with tumor necrosis factor-α to up-regulate the expression of E-selectin. sCR1sLe x inhibited the binding of U937 cells to surface-adsorbed P-selectin-IgG. sCR1sLe x and sCR1[desLHR-A]sLe x have thus demonstrated both complement regulatory activity and the capacity to bind selectins and to inhibit selectin-mediated cell adhesion in vitro .