•Various neoglycosphingolipids were synthesized in a one-step reaction.•The neoglycosphingolipids were made in good yields with high chemoselectivities.•Most of the synthesized compounds could upregulate MMP-9 expression in B16 cells.•These neoglycosphingolipids have potential for use as anti-tumor agents.Various neoglycosphingolipids were efficiently synthesized in a one-step reaction by the coupling of free sugars with an N-alkylaminooxy-functionalized ceramide analogue. The bioactivity studies demonstrated that most of these compounds could upregulate the expression of matrix metalloproteinase-9 (MMP-9, extracellular matrix proteins associated with tumor migration) in murine melanoma B16 cells in a similar manner to the natural ganglioside monosialodihexosylganglioside (GM3), which highlights the potential use of these neoglycosphingolipids as inhibitors of tumor migration.Download high-res image (155KB)Download full-size image

Zanamivir and oseltamivir are principal influenza antiviral drugs that target viral neuraminidase (NA), but resistant viruses containing mutant NAs with diminished drug affinity are increasingly emerging. Using the structural knowledge of both drug-binding sites and their spatial arrangement on the homotetrameric NA, we have developed a tetravalent zanamivir (TZ) molecule that exhibited marked increases in NA binding affinity, inhibition of NA enzyme activity, and in vitro plus in vivo antiviral efficacy over zanamivir. TZ functioned against both human seasonal H3N2 and avian H7N9 viruses, including drug-resistant mutants. Crystal structure of a resistant N9 NA in complex with TZ explained the function, which showed that four zanamivir residues simultaneously bound to all four monomers of NA. The design method of TZ described in this study may be useful to develop drugs or ligands that target proteins with multiple binding sites. The potent anti-influenza activity of TZ makes it attractive for further development.

•The site-specific conjugation of PEG to a peptide drug enfuvirtide is accomplished.•The conjugate shows similar anti-HIV activity and action mechanism to enfuvirtide.•The conjugate has better aqueous solubility and longer half-life than enfuvirtide.•The conjugate inhibits the most common HIV strains circulating in China.Enfuvirtide (ENF) is a clinically used peptide drug for the treatment of HIV infections, but its poor pharmacokinetic profile (T1/2 = 1.5 h in rats) and low aqueous solubility make the therapy expensive and inconvenience. In this study, we present a simple and practical strategy to address these problems by conjugating ENF with polyethylene glycol (PEG). Site-specific attachment of a 2 kDa PEG at the N-terminus of ENF resulted in an ENF−PEG (EP) conjugate with high solubility (≥3 mg/mL) and long half-life in rats (T1/2 = 16.1 h). This conjugate showed similar antiviral activity to ENF against various primary HIV-1 isolates (EC50 = 6–91 nM). Mechanistic studies suggested the sources of the antiviral potency. The conjugate bound to a functional domain of the HIV gp41 protein in a helical conformation with high affinity (Kd = 307 nM), thereby inhibiting the gp41-mediated fusion of viral and host-cell membranes. As PEG conjugation has advanced many bioactive proteins and peptides into clinical applications, the EP conjugate described here represents a potential new treatment for HIV infections that may address the unmet medical needs associated with the current ENF therapy.

Many peptide-based therapeutics have short circulatory half-lives. We report here that the pharmacokinetics of an anti-HIV peptide drug enfuvirtide (ENF) can be dramatically improved by a chemical glycosylation approach. A set of glycosylated ENFs with varying glycosylation sites and glycan structures were synthesized. Among these, a sialic acid-introduced peptide (SL-ENF) demonstrated a 15-fold extended half-life in rats relative to ENF (T1/2: 23.1 vs 1.5 h), and its antiviral potency was comparable to that of ENF (EC50: 2 vs 3 nM). SL-ENF bound to a functional fragment of the HIV fusogenic protein gp41 and formed complexes with high affinity and α-helicity, revealing the mechanism behind its potent antiviral activity. Because it is widely accepted in biology that glycosylation protects proteins from denaturation and proteases, our approach may be useful for the development of novel protein and peptide drugs with enhanced pharmaceutical properties.

An ionic liquid-supported synthetic method for the construction of glycopeptides in high yields is reported. This method avoids the use of large excesses of reagents and chromatographic purification and, therefore, represents a useful addition to existing approaches for the ionic liquid-supported synthesis of oligosaccharides and peptides.

•A chitosan–sialyloligosaccharides (CS) complex has been prepared and characterized.•The CS complex was formed via ionic adsorption between chitosan and sialyloligosaccharides.•The CS complex was prepared on a gram-scale from the natural source by a one-step dialysis process.•Potent inhibition of influenza virus infection by the CS complex was demonstrated.With the recent emergence of drug-resistant influenza viruses, effective means of preventing and treating these contagious pathogens have become imperative. The binding receptors of influenza virus are sialyloligosaccharides (SOS), which are present on the surfaces of host cells, and are therefore attractive targets for antiviral development. We report the preparation and identification of a novel influenza virus entry inhibitor, designated chitosan–SOS complex (CS complex). The CS complex was formed through noncovalent adsorption between cationic chitosan and anionic SOS, the latter derived from bovine colostrum. The preparation was accomplished in gram quantities from chitosan and bovine colostrum oligosaccharides by a one-step dialysis process. The inhibitory activity of the complex against influenza virus infection was determined by cytotoxicity inhibition assay (IC50 = 42 μM). This simple preparation, combined with efficient anti-infective activity and the rich natural availability of chitosan and SOS, highlights the potential of the CS complex as a safe, practical agent for influenza prevention and control.

Recent cases of human infection with avian influenza H5N1 and H7N9 viruses underscore an urgent need for techniques that can rapidly assess their potential threat to the humans. Determination of the receptor-binding property of influenza virus is crucial to direct viral control and prevention measures. Current methods to perform this analysis are dependent on immunoanalytical strategies that use unstable biological components and complex procedures. We have developed a facile colorimetric assay to determine the interaction of the viral hemagglutinin (HA) protein with host glycan receptors using glycan-functionalized gold nanoparticles (gGNPs). This method is based on the color and absorbance changes of gold probes when the solution is simply mixed with HAs or intact viruses. The resulting sensitivity and selectivity has enabled HA/virus binding to various glycan structures to be differentiated visually and rapidly. Using this system, we have screened, in parallel, the receptor specificity of eight representative human and avian viral HAs and three whole viruses including an emerging H7N9 strain. Our results reveal the detailed receptor-binding profiles of H7N9 virus and its HA and show that they effectively bind to human-type receptors. This gGNP-based assay represents a strategy that would be helpful for developing simple and sensitive systems to probe glycan-mediated biological processes.Keywords: glycan; gold nanoparticle; influenza virus; receptor specificity; sialic acid

Shiga toxin (Stx)-producing Escherichia coli (STEC) causes diarrhea and colitis in humans that can develop into a life-threatening hemolytic uremic syndrome (HUS). Developing efficient means of controlling STEC diseases, for which no drugs or vaccines are currently available, remains a high priority. We report here the construction and development of chitosan conjugates bearing the Stx ligand trisaccharide globotriose to demonstrate their potential as STEC disease treatment agents. The synthesis was accomplished by grafting a globotriose derivative containing an aldehyde-functionalized aglycone to chitosan amino groups. The obtained globotriose–chitosan conjugate bound with high affinity to Stx and efficiently neutralized its toxicity on Vero cells. Moreover, Stx levels in the gut of infected mice receiving oral doses of the conjugate were greatly diminished, enabling the mice to resist a fatal STEC challenge. The conjugate appears to function as a Stx adsorbent in the gut, preventing toxin entry into the bloodstream and consequent development of HUS. As such, the conjugate could act as a novel agent against STEC disease.

This paper describes the facile synthesis and bioevaluations of a novel conjugated polymer having tetrasaccharide SLeX branch and chitosan backbone (SLeX–chitosan conjugate). The synthesis was achieved by a convenient chemoenzymatic approach using an aldehyde-functionalized N-acetylglucosamine to branch it onto chitosan amino groups followed by three enzymatic reactions to further append galactose, sialic acid, and fucose residue to the branch. Surface plasmon resonance (SPR) study of SLeX–chitosan conjugate revealed a high affinity binding with E-selectin (Kd = 920 nM) and a potent inhibitory effect on the binding of E-selectin with SLeX–BSA (IC50 = 240 μM). By using biocompatible chitosan as the scaffold for presenting SLeX ligands, as well as the concise route tailored for the conjugate syntheses, the present study provides a practical method to explore safe and efficient anti-inflammatory agents.

GlmM and GlmU are key enzymes in the biosynthesis of UDP-N-acetyl-d-glucosamine (UDP-GlcNAc), an essential precursor of peptidoglycan and the rhamnose–GlcNAc linker region in the mycobacterial cell wall. These enzymes are involved in the conversion of two important precursors of UDP-GlcNAc, glucosamine-6-phosphate (GlcN-6-P) and glucosamine-1-phosphate (GlcN-1-P). GlmM converts GlcN-6-P to GlcN-1-P, GlmU is a bifunctional enzyme, whereby GlmU converts GlcN-1-P to GlcNAc-1-P and then catalyzes the formation of UDP-GlcNAc from GlcNAc-1-P and uridine triphosphate. In the present study, methyl 2-amino-2-deoxyl-α-d-glucopyranoside 6-phosphate (1α), methyl 2-amino-2-deoxyl-β-d-glucopyranoside 6-phosphate (1β), two analogs of GlcN-6-P, were synthesized as GlmM inhibitors; 2-azido-2-deoxy-α-d-glucopyranosyl phosphate (2) and 2-amino-2,3-dideoxy-3-fluoro-α-d-glucopyranosyl phosphate (3), analogs of GlcN-1-P, were synthesized firstly as GlmU inhibitors. Compounds 1α, 1β, 2, and 3 as possible inhibitors of mycobacterial GlmM and GlmU are reported herein. Compound 3 showed promising inhibitory activities against GlmU, whereas 1α, 1β and 2 were inactive against GlmM and GlmU even at high concentrations.

The high transmissibility and genetic variability of the influenza virus have made the design of effective approaches to control the infection particularly challenging. The virus surface hemagglutinin (HA) protein is responsible for the viral attachment to the host cell surface via the binding with its glycoligands, such as sialyllactose (SL), and thereby is an attractive target for antiviral designs. Herein we present the facile construction and development of two SL-incorporated chitosan-based materials, either as a water-soluble polymer or as a functional fiber, to demonstrate their abilities for viral adhesion inhibition and decontamination. The syntheses were accomplished by grafting a lactoside bearing an aldehyde-functionalized aglycone to the amino groups of chitosan or chitosan fiber followed by the enzymatic sialylation with sialyltransferase. The obtained water-soluble SL–chitosan conjugate bound HA with high affinity and inhibited effectively the viral attachment to host erythrocytes. Moreover, the SL-functionalized chitosan fiber efficiently removed the virus from an aqueous medium. The results collectively demonstrate that these potential new materials may function as the virus adsorbents for prevention and control of influenza. Importantly, these materials represent an appealing approach for presenting a protein ligand on a chitosan backbone, which is a versatile molecular platform for biofunctionalization and, thereby, can be used for not only antiviral designs, but also extensive medical development such as diagnosis and drug delivery.

The aim of this work is to construct a safe and effective drug candidate against Streptococcus suis infection. A panel of chitosan-based polymer conjugates with branched galabiose (Galα1−4Gal) side chains was synthesized as inhibitors of S. suis adhesion. The synthesis was achieved by using an aldehyde-functionalized galabiose derivative to graft it onto chitosan amino groups. Structural compositions of the conjugates were verified by 1H NMR spectroscopy and CHN elemental analyses. Potent inhibitory activities of the conjugates against S. suis adhesion to human erythrocytes were determined at low nanomolar concentration by HAI assay. An SPR study revealed a high affinity binding (Kd = 39.6 nM) of the conjugate with BSI-B4 lectin. By using biocompatible chitosan as the scaffold for presenting S. suis-specific galabiose units, as well as the concise route tailored for the conjugate syntheses, the present study provides a practical way for explorations of new anti-S. suis therapies.

•A simple method for the detection of influenza viruses is reported.•The method uses glycan-functionalized gold nanoparticles as self-reporting probes.•Fourteen viruses were differentiated in a single-step colorimetric procedure.•The method has potential for the development of rapid sensors for clinical use.Accurate diagnosis of influenza viruses is difficult and generally requires a complex process because of viral diversity and rapid mutability. In this study, we report a simple and rapid strategy for the detection and differentiation of influenza viruses using glycan-functionalized gold nanoparticles (gGNPs). This method is based on the aggregation of gGNP probes on the viral surface, which is mediated by the specific binding of the virus to the glycans. Using a set of gGNPs bearing different glycan structures, fourteen influenza virus strains, including the major subtypes currently circulating in human and avian populations, were readily differentiated from each other and from a human respiratory syncytial virus in a single-step colorimetric procedure. The results presented here demonstrate the potential of this gGNP-based system in the development of convenient and portable sensors for the clinical diagnosis and surveillance of influenza viruses.