Synthesis of a new nanomolar saccharide inhibitor of lymphocyte adhesion: Different polylactosamine backbones present multiple sialyl Lewis x determinants to L-selectin in high-affinity mode

Abstract

Lymphocyte infiltration is a hallmark of acute rejections in solid organ transplants, such as cardiac allograft. We have previously shown that lymphocyte extravasation to cardiac grafts undergoing rejection is largely due to interactions between lymphocyte L-selectin and its sialyl Lewis x (sLex) decorated ligands. Our previous work demonstrated further that an enzymatically synthetized tetravalent sLex glycan of a branched polylactosamine backbone is a highly efficient inhibitor of L-selectin-dependent lymphocyte adhesion to graft endothelium. To improve the availability of multivalent sLex glycans for anti-inflammatory indications, we now report enzymatic synthesis of another tetravalent sLex glycan that can be potentially produced on a large scale, and show that even the new saccharide is a nanomolar inhibitor of L-selectin-dependent lymphocyte adhesion. The novel antagonist is sLexβ1-3'(sLexβ1-6')LacNAcpβ1-3' (sLexβ1-6')LacNAcβ1-3'(sLexβ1-6')LacNAc (8) (where LacNAc is the disaccharide Galβ1-4GlcNAc and sLex is the tetrasaccharide Neu5Aα2-3Galβ1-4(Fucα-3)G1cNAc). Its five-step synthesis was started from the octameric polylactosamine LacNAcβ1-3'(GlcNAcβ1-6')LacNAcβ1-3'(GlcNAcβ1-6')LacNAc (3), which in turn is accessible in one step from the hexasaccharide LacNBcβ1-3'LacNAcβ1-3'LacNAc. Importantly, the hexasaccharide primer has been synthesized chemically (Alais and Veyrieres, Tetrahedron Lett., 24, 5223, 1983), Hence, our data outline a route to glycan 8, consisting of a combination of chemical and enzymatic methods of oligosaccharide synthesis. In addition, our data show that polylactosamine backbones are able to present multiple sialyl Lewis x determinants to L-selectin in high-affinity mode, without a requirement for uniqueness in the backbone structure.