A versatile approach has been developed for the multiple labeling of oligonucleotides. First, three linkers as a H-phosphonate monoester derivative were condensed on a solid-supported T12 to introduce H-phosphonate diester linkages which were oxidized in the presence of propargylamine. Second, three galactosyl azide derivatives were conjugated to the solid-supported three-alkyne-modified T12 by a 1,3-cycloaddition so-called "click chemistry" in the presence of Cu(I) assisted by microwaves.

A family of seven topologically isomeric calix[4]arene glycoconjugates was prepared through the synthesis of a series of alkyne-derivatised calix[4]arene precursors that are suitable for the attachment of sugar moieties by microwave-assisted copper(I)-catalysed azide-alkyne cycloaddition (CuAAC). The glycoconjugates thus synthesised comprised one mono-functionalised derivative, two 1,2- or 1,3-divalent regioisomers, one trivalent and three tetravalent topoisomers in the cone, partial cone or 1,3-alternate conformations. The designed glycoconjugates were evaluated as ligands for the galactose-binding lectin PA-IL from the opportunistic bacterium Pseudomonas aeruginosa, a major causative agent of lung infections in cystic fibrosis patients. Binding affinities were determined by isothermal titration calorimetry (ITC), and the interaction with the lectin was shown to be strongly dependant on both the valence and the topology. Whereas the trivalent conjugate displayed enhanced affinity when compared to a monosaccharide model, the tetravalent conjugates are to-date the highest-affinity ligands measured by ITC. The topologies presenting carbohydrates on both faces of calixarene are the most potent ones with dissociation constants of approximately 200 nM. Molecular modelling suggests that such a multivalent molecule can efficiently chelate two of the binding sites of the tetrameric lectin; this explains the 800-fold increase of affinity achieved by the tetravalent molecule. Surface plasmon resonance (SPR) experiments confirmed that this glycoconjugate is the strongest inhibitor for binding of PA-IL to galactosylated surfaces for potential applications as an anti-adhesive agent.

This paper is dedicated to Dr. 0. E. (Ted) Edwards J.-P. PRALY and R. U. LEMIEUX. Can. J. Chem. 65, 213 (1987). A novel application of I3c nuclear magnetic resonance provided the effects of solvent polarity and hydrogen-bond formation on the conformational equilibria for a range of 2-substituted tetrahydropyrans and the results are interpreted in terms of how solvent affects the competition between the endo-and exo-anomeric effects in determining the magnitude of the anomeric effect. In accord with the generally accepted origin of the endo-and exo-anomeric effects (anti-periplanar n-a* interaction of the oxygen lone-pair orbital with the antibonding orbital of the adjacent C-0 bond), the exo-anomeric effect for the a anomer is expected to be weaker because charge delocalization from the glycosidic oxygen to anomeric center is in competition with delocalization from the ring-oxygen atom. The effects of solvent on the relative magnitudes of the endo-and exo-anomeric effects are then considered to arise from the formation of specific complexes with the solvent, and the exo-anomeric effect of a P-glycoside is more strongly influenced. It is contended that hydrogen bonding of solvent to the ring oxygen increases the exo-anomeric effects. For this reason water is particularly effective for the strengthening of the exo-anomeric effect and, thereby, the conformational rigidity of glycosides. Experimental evidence is presented that indicates that the anomeric hydroxyl groups of free sugars dissolved in water tend to prefer the equatorial orientation because these provide stronger hydrogen bonds as proton donors to water.

The design of multivalent glycoclusters requires the conjugation of biologically relevant carbohydrate epitopes functionalized with linker arms to multivalent core scaffolds. The multigram-scale syntheses of three structurally modified triethyleneglycol analogues that incorporate amide moiety(ies) and/or a phenyl ring offer convenient access to a series of carbohydrate probes with different water solubilities and rigidities. Evaluation of flexibility and determination of preferred conformations were performed by conformational analysis. Conjugation of the azido-functionalized carbohydrates with tetra-propargylated core scaffolds afforded a library of 18 tetravalent glycoclusters, in high yields, by Cu(I)-catalyzed azide-alkyne cycloaddition (CuAAC). The compounds were evaluated for their ability to bind to PA-IL (the LecA lectin from the opportunistic pathogen Pseudomonas aeruginosa). Biochemical evaluation through inhibition of hemagglutination assays (HIA), enzyme-linked lectin assays (ELLA), surface plasmon resonance (SPR), and isothermal titration microcalorimetry (ITC) revealed improved and unprecedented affinities for one of the monovalent probes (K(d)=5.8 μM) and also for a number of the tetravalent compounds that provide several new nanomolar ligands for this tetrameric lectin.

This Review summarizes close to 500 primary publications and surveys published since 2000 about the syntheses and diverse bioactivities of C-glycopyranosyl (het)arenes. A classification of the preparative routes to these synthetic targets according to methodologies and compound categories is provided. Several of these compounds, regardless of their natural or synthetic origin, display antidiabetic properties due to enzyme inhibition (glycogen phosphorylase, protein tyrosine phosphatase 1B) or by inhibiting renal sodium-dependent glucose cotransporter 2 (SGLT2). The latter class of synthetic inhibitors, very recently approved as antihyperglycemic drugs, opens new perspectives in the pharmacological treatment of type 2 diabetes. Various compounds with the C-glycopyranosyl (het)arene motif were subjected to biological studies displaying among others antioxidant, antiviral, antibiotic, antiadhesive, cytotoxic, and glycoenzyme inhibitory effects.

The design of multivalent glycoconjugates has been developed over the past decades to obtain high-affinity ligands for lectin receptors. While multivalency frequently increases the affinity of a ligand for its lectin through the so-called "glycoside cluster effect", the binding profiles towards different lectins have been much less investigated. We have designed a series of multivalent galactosylated glycoconjugates and studied their binding properties towards two lectins, from plant and bacterial origins, to determine their potential selectivity. The synthesis was achieved through copper(I)-catalysed azide-alkyne cycloaddition (CuAAC) under microwave activation between propargylated multivalent scaffolds and an azido-functionalised carbohydrate derivative. The interactions of two galactose-binding lectins from Pseudomonas aeruginosa (PA-IL) and Erythrina cristagalli (ECA) with the synthesized glycoclusters were studied by hemagglutination inhibition assays (HIA), surface plasmon resonance (SPR) and isothermal titration microcalorimetry (ITC). The results obtained illustrate the influence of the scaffold's geometry on the affinity towards the lectin and also on the relative potency in comparison with a monovalent galactoside reference probe.

Carbohydrates and glycoconjugates play a major role in key biological events such as cell-cell recognition, pathogenesis, and inflammation. [1,2] As a consequence, there is a need to understand the structural parameters governing the recognition of carbohydrates by their receptors. This knowledge will be of use for both fundamental research and potential applications in diagnostics or therapeutics. However, research in this field is slowed by the wide diversity of carbohydrate structures and by the minute amounts of materials available for experimentation. The design of sensitive and highthroughput technologies for the characterization of oligosaccharide/protein interactions [3] is therefore emerging as an attractive tool for chemists and biochemists. Available techniques such as isothermal calorimetry, enzyme-linked lectin assay, and even crystallographic studies provide data on carbohydrate/protein interactions, but they are often limited by the amount of available material.Carbohydrate microarray technology [4][5][6][7][8][9][10][11][12][13][14][15][16] is a promising approach for probing carbohydrate/protein interactions, and it permits the simultaneous screening of a number of biological interactions with only minute amounts of material. A large family of carbohydrate derivatives has been designed for immobilization on surfaces by various means. [5][6][7][8][9][10][11][12][13][14] However, this technology has various limitations. Relative surface densities of bound ligands are often not assessed. A careful optimization of the orientation and the distance separating the carbohydrate probe from the surface is often required.The interactions of oligosaccharides with lectins are usually weak (mm range) and can be enhanced using the "cluster effect" with multivalent ligands. [17][18][19][20] In the latter case, the distance between the residues should be optimized for binding. [21][22][23] Finally, the syntheses of functionalized oligosaccharide ligands are labor intensive.We report herein an original approach for the surface immobilization of oligosaccharides using glycoconjugate molecules that present a DNA sequence for anchoring onto DNA chips through hybridization. This approach has been used in the field of protein microarrays, [24,25] but to our knowledge this is the first time that such a strategy has been reported in the field of glycoarrays.Several syntheses of glycoconjugated oligonucleotides have been reported, but none are suitable for introducing different carbohydrate moieties. [26][27][28] We designed a conjugate that incorporates carbohydrate residue(s) for interacting with a lectin, an oligonucleotide sequence for anchoring on the surface, and a fluorescent tag at the 5'-end for the determination of relative surface densities (Figure 1). These moieties were assembled through a combination of automated oligonucleotide synthesis, and amidative oxidation and 1,3-dipolar cycloaddition ("click" chemistry) performed on a solid support (Scheme 1).[29] We introduced either one or three saccharide residue...

Multivalency is playing a major role in biological processes and particularly in lectin-carbohydrate interactions. The design of high-affinity ligands of lectins should provide molecules capable of interfering with these biological processes and potentially inhibit bacterial or viral infections. Azide-alkyne "click" chemistry was applied to the synthesis of dodecavalent fullerene-based glycoclusters. The conjugation could be efficiently performed from alkyne or azide functions on either partners (i.e. hexakis-fullerene adduct or glycoside). PA-IL is a bacterial lectin from the opportunistic pathogen Pseudomonas aeruginosa and is involved in the recognition of glycoconjugates on human tissues. The glycoclusters obtained were evaluated as ligands of PA-IL and for their potential for competing with its binding to glycosylated surfaces. The affinities measured by hemagglutination inhibition assay (HIA), enzyme-linked lectin assay (ELLA), and surface plasmon resonance (SPR) displayed a significant "glycoside cluster effect" with up to a 12,000-fold increase in binding when comparing a monovalent carbohydrate reference probe with a dodecavalent fullerene-based glycocluster, albeit with some differences depending on the analytical technique.