ABSTRACT
The major glycosphingolipid in pig vascular endothelium is the ceramide pentasaccharide Gal alpha(1 --> 3)Gal beta(1 --> 4)GlcNAc beta(1 --> 3)Gal beta(1 --> 4)Glc beta(1 --> 0)Cer (1), which binds specifically to human anti-Gal antibody and is involved in the hyperacute rejection response in xenotransplantation from pig to man. The synthesis of 1 and its methyl glycoside 2 is described.
Subject(s)
Antigens, Heterophile/biosynthesis , Glycosphingolipids/chemical synthesis , Graft Rejection/immunology , Oligosaccharides/chemical synthesis , Transplantation, Heterologous/immunology , Animals , Antibodies, Heterophile/immunology , Antigens, Heterophile/adverse effects , Antigens, Heterophile/chemistry , Carbohydrate Conformation , Carbohydrate Sequence , Endothelium, Vascular/chemistry , Galactose/immunology , Glycosphingolipids/immunology , Humans , Molecular Sequence Data , Swine , Transplantation, Heterologous/adverse effectsSubject(s)
Antibodies, Heterophile/blood , Disaccharides/immunology , Graft Rejection/prevention & control , Heart Transplantation/immunology , Transplantation, Heterologous/immunology , Acute Disease , Animals , Animals, Genetically Modified , Antigens, CD/genetics , Antigens, CD/immunology , CD55 Antigens/genetics , CD55 Antigens/immunology , Complement C3/analysis , Complement C4/analysis , Endothelium, Vascular/immunology , Graft Rejection/immunology , Humans , Immunoglobulin M/analysis , Papio , SwineABSTRACT
An early step of the inflammatory response-the rolling of leukocytes on activated endothelial cells-is mediated by selectin/carbohydrate interactions. The tetrasaccharide sialyl Lewis(x) (sLe(x)) 1 is a ligand for E-, P-, and L-selectin and, therefore, serves as a lead structure to develop analogues which allow the control of acute and chronic inflammation. Here we describe the efficient synthesis (10 linear steps) of the potent sLe(x) mimetic 2. Compared to sLe(x), compound 2 showed a 30-fold improved affinity in a static, cell-free E-selectin-ligand binding assay (IC(50) = 36 microM). These data were confirmed by a marked inhibition in an in vitro cell-cell rolling assay which simulates in vivo conditions (IC(50) approximately 40 microM). The assays are predictive for the in vivo efficacy of test compounds as indicated by a marked inhibitory effect of 2 in a thioglycollate induced peritonitis model of acute inflammation in mice (ED(50) approximately 15 mg/kg).
Subject(s)
E-Selectin/metabolism , Oligosaccharides/chemical synthesis , Acute Disease , Animals , Carbohydrate Sequence , Ligands , Mice , Molecular Mimicry , Molecular Sequence Data , Oligosaccharides/chemistry , Oligosaccharides/pharmacology , Peritonitis/chemically induced , Peritonitis/drug therapy , Sialyl Lewis X Antigen , Structure-Activity Relationship , ThioglycolatesABSTRACT
The synthesis of the ethoxycarbonyloctanyl glycoside of the Lewis-Y (Ley) tetrasaccharide, a part of complex glycosphingolipids, was based on the trichloroacetimidate method for glycoside synthesis. The regioselective introduction of protective groups and the high yield of epimers of the azidonitration reaction applied to O-[2,3,4-tri-O-acetyl-6-O-[dimethyl-(2,3-dimethyl-2-butyl)silyl]-beta-D- galactopyranosyl]-(1-->4)-3-O-acetyl-1,5-anhydro-6-O-[dimet hyl-(2,3- dimethyl-2-butyl)silyl]-D-arabino-hex-1-enitol led to a lactosamine acceptor molecule. The double specific introduction of an alpha-L-fucosyl group in high yield gave a protected Ley-tetrasaccharide. After activation to give the alpha-trichloroacetimidate, specific beta-glycosylation with 8-ethoxycarbonyl octanol under SN 2 condition, followed by cleavage of all protective groups led to the tetrasaccharide, alpha-L-Fucp-(1-->2)-beta-D-galp-(1-->4)-[alpha-L-fucp-(1--> 3)]-beta-D- GlcpNAcO(CH2)8CO2Et in high yield.
Subject(s)
Antigens, Neoplasm/chemistry , Lewis Blood Group Antigens/chemistry , Oligosaccharides/chemical synthesis , Carbohydrate Conformation , Carbohydrate Sequence , Carbohydrates , Glycosphingolipids/chemistry , Indicators and Reagents , Magnetic Resonance Spectroscopy , Molecular Sequence Data , Structure-Activity RelationshipABSTRACT
The synthesis of the methyl alpha- and beta-N-dansyl-D-galactosaminides is described using methyl alpha,beta-2-azido-2-deoxy-D-galactopyranoside as starting material. This was reduced to the corresponding methyl alpha,beta-2-amino-2-deoxy-D-galactopyranoside and then treated with dansyl chloride to yield a mixture of methyl alpha,beta-N-dansyl-D-galactosaminides which was separated into individual anomeric forms by flash chromatography on silica gel. Methyl alpha-N-dansyl-D-galactosaminide was used as a fluorescent indicator ligand in continuous substitution titrations to determine the association constants of nonchromophoric carbohydrates with the N-acetyl-D-galactosamine specific lectin from Erythrina corallodendron.
Subject(s)
Acetylgalactosamine/metabolism , Dansyl Compounds/chemical synthesis , Galactosamine/analogs & derivatives , Lectins/metabolism , Galactosamine/chemical synthesis , Molecular Probes , Molecular Structure , Protein Binding , Sensitivity and Specificity , Spectrometry, FluorescenceABSTRACT
Mucin-type O-glycopeptides containing the beta-D-Galp-(1----3)-D-GalpNAc disaccharide core unit, which is also the T-antigenic determinant, were synthesized from D-galactose, 2-azido-2-deoxy-D-galactose, 2-azido-2-deoxylactose, and L-serine precursors by applying the trichloroacetimidate method. Thus, beta-D-Galp-(1----3)-alpha-D-GalpNAc-(1----3)-Ser (1) and beta-D-Galp-(1----4)-beta-D-GlcpNAac-(1----6)-[beta-D-Galp-(1----3 )]-alpha-D-GalpNAc-(1----3)-Ser (2) were obtained. A protected precursor of 2 having free OH groups at C-4 and C-6 of the inner sugar unit is a valuable intermediate for the synthesis of further O-glycopeptides of this core-unit type.
