Shang-Cheng Hung

Regioselective one-pot protection of carbohydrates

Carbohydrates are involved in a wide range of biological processes. These structurally diverse compounds are more complex than other biological polymers, and are often present as heterogeneous mixtures in nature. The chemical synthesis of carbohydrates is one way to obtain pure oligosaccharides, but it is hampered by difficulties associated with the regioselective protection of polyhydroxyls and challenges related to the stereoselective assembly of glycosidic linkages. Here we describe a combinatorial, and highly-regioselective, method that can be used to protect individual hydroxy groups of a monosaccharide. This approach can be used to install an orthogonal protecting group pattern in a single reaction vessel (a ‘one-pot’ reaction), which removes the need to carry out the time-consuming isolation and purification of intermediates. Hundreds of building blocks have been efficiently prepared starting from d-glucose, and the iterative coupling of these building blocks enabled us to assemble β-1,6-glucans and a library of oligosaccharides based on the influenza-virus-binding trisaccharide.

Toward Automated Oligosaccharide Synthesis

Carbohydrates have been shown to play important roles in biological processes. The pace of development in carbohydrate research is, however, relatively slow due to the problems associated with the complexity of carbohydrate structures and the lack of general synthetic methods and tools available for the study of this class of biomolecules. Recent advances in synthesis have demonstrated that many of these problems can be circumvented. In this Review, we describe the methods developed to tackle the problems of carbohydrate-mediated biological processes, with particular focus on the issue related to the development of the automated synthesis of oligosaccharides. Further applications of carbohydrate microarrays and vaccines to human diseases are also highlighted.

Synthesis of 3-O-sulfonated heparan sulfate octasaccharides that inhibit the herpes simplex virus type 1 host–cell interaction

Cell surface carbohydrates play significant roles in a number of biologically important processes. Heparan sulfate, for instance, is a ubiquitously distributed polysulfated polysaccharide that is involved, among other things, in the initial step of herpes simplex virus type 1 (HSV-1) infection. The virus interacts with cell-surface heparan sulfate to facilitate host-cell attachment and entry. 3-O-Sulfonated heparan sulfate has been found to function as an HSV-1 entry receptor. Achieving a complete understanding of these interactions requires the chemical synthesis of such oligosaccharides, but this remains challenging. Here, we present a convenient approach for the synthesis of two irregular 3-O-sulfonated heparan sulfate octasaccharides, making use of a key disaccharide intermediate to acquire different building blocks for the oligosaccharide chain assembly. Despite substantial structural differences, the prepared 3-O-sulfonated sugars blocked viral infection in a dosage-dependent manner with remarkable similarity to one another.

Regioselective one-pot protection of glucose

Detailed protocols for the regioselective protection of individual hydroxyls in monosaccharide units are described here. This expedient methodology incorporates up to seven reaction sequences, obviating the necessity to carry out intermittent tedious work-ups and time-consuming purifications. Using this TMSOTf-catalyzed one-pot protocol, the 2,3,4,6-tetra-O-trimethylsilylated hexopyranosides bearing an anomeric group could be transformed into a whole set of differentially protected 2-alcohols, 3-alcohols, 4-alcohols, 6-alcohols and fully protected monosaccharides in high yields. These tailor-made glycosyl donors and acceptors can then be used for stereoselective one-pot glycosylation for oligosaccharide synthesis. The total time for the preparation of a purified protected sugar unit ranges between 1 and 2 d. This process would otherwise take 1–2 weeks.

Sc(OTf)3-catalyzed acetolysis of 1,6-anhydro-β-hexopyranoses and solvent-free per-acetylation of hexoses

Acetolysis of 1,6-anhydro-β-hexopyranoses and solvent-free per-acetylation of hexoses with acetic anhydride at room temperature in excellent yields employing 0.5 mol% scandium(III) trifluoromethanesulfonate as an extremely efficient catalyst are, respectively, described here.

α-Glycosylation by d-Glucosamine-Derived Donors: Synthesis of Heparosan and Heparin Analogues That Interact with Mycobacterial Heparin-Binding Hemagglutinin

Numerous biomolecules possess α-D-glucosamine as structural component. However, chemical glycosylations aimed at this backbone are usually not easily attained without generating the unwanted β-isomer. We report herein a versatile approach in affording full α-stereoselectivity built upon a carefully selected set of orthogonal protecting groups on a D-glucosaminyl donor. The excellent stereoselectivity provided by the protecting group combination was found independent of leaving groups and activators. With the trichloroacetimidate as the optimum donor leaving group, core skeletons of glycosylphosphatidyl inositol anchors, heparosan, heparan sulfate, and heparin were efficiently assembled. The orthogonal protecting groups were successfully manipulated to further carry out the total syntheses of heparosan tri- and pentasaccharides and heparin di-, tetra-, hexa-, and octasaccharide analogues. Using the heparin analogues, heparin-binding hemagglutinin, a virulence factor of Mycobacterium tuberculosis, was found to bind at least six sugar units with the interaction notably being entropically driven.

Divergent synthesis of 48 heparan sulfate-based disaccharides and probing the specific sugar-fibroblast growth factor-1 interaction.

Several biological processes involve glycans, yet understanding their ligand specificities is impeded by their inherent diversity and difficult acquisition. Generating broad synthetic sugar libraries for bioevaluations is a powerful tool in unraveling glycan structural information. In the case of the widely distributed heparan sulfate (HS), however, the 48 theoretical possibilities for its repeating disaccharide call for synthetic approaches that should minimize the effort in an undoubtedly huge undertaking. Here we employed a divergent strategy to afford all 48 HS-based disaccharides from just two orthogonally protected disaccharide precursors. Different combinations and sequence of transformation steps were applied with many downstream intermediates leading up to multiple target products. With the full disaccharide library in hand, affinity screening with fibroblast growth factor-1 (FGF-1) revealed that four of the synthetic sugars bind to FGF-1. The molecular details of the interaction were further clarified through X-ray analysis of the sugar-protein cocrystals. The capability of comprehensive sugar libraries in providing key insights in glycan-ligand interaction is, thus, highlighted.

Characterization of Molecular Interactions between Eosinophil Cationic Protein and Heparin

Eosinophil cationic protein (ECP) is currently used as a biomarker for airway inflammation. It is a heparin-binding ribonuclease released by activated eosinophils. Its cytotoxicity toward cancer cell lines is blocked by heparin. The objective of this study was to locate the heparin binding site of ECP by site-directed mutagenesis and construction of a synthetic peptide derived from this region. Synthetic heparin with ≥5 monosaccharide units showed strong inhibition of ECP binding to the cell surface. Analysis of ECP mt1 (R34A/W35A/R36A/K38A) showed that these charged and aromatic residues were involved in ECP binding to heparin and the cell surface. A potential binding motif is located in the loop L3 region between helix α2 and strand β1, outside the RNA binding domain. The synthetic peptide derived from the loop L3 region displayed strong pentasaccharide binding affinity and blocked ECP binding to cells. In addition, ECP mt1 showed reduced cytotoxicity. Thus, the tight interaction between ECP and heparin acts as the primary step for ECP endocytosis. These results provide new insights into the structure and function of ECP for anti-asthma therapy.

Acyl and Silyl Group Effects in Reactivity-Based One-Pot Glycosylation: Synthesis of Embryonic Stem Cell Surface Carbohydrates Lc4 and IV2Fuc-Lc4

Relative reactivity evaluations showed the graded arming of toluenyl thioglucosides by variously positioned silyl groups but not by their acyl counterparts. These findings were applied in reactivity-based one-pot assembly of linker-attached Lc(4) and IV(2)Fuc-Lc(4), which are components of human embryonic stem cell surface. The sugar-galectin-1 binding was also examined.

Regioselective One-Pot Protection of D-Glucosamine

A highly regioselective one-pot transformation of 2-azido-2-deoxy-1,3,4,6-tetra-O-trimethylsilyl-d-glucopyranose via sequential additions of various reagents was systematically studied, yielding the fully protected derivatives and the 1-, 3-, 4-, as well as 6-alcohols, respectively.