Carbohydrates themed issue

Glycoscience was for a long time the Cinderella field of science, as carbohydrates were the least exploited of the three major classes of biomolecules: proteins, nucleic acids, and carbohydrates. Thus, despite the important roles that carbohydrates play in numerous biological recognition events, the molecular details of these recognition processes are generally not well understood, and consequently the rate of development of carbohydrate-based therapeutics has been relatively slow. This slow rate of development was further hindered by the lack of practical synthetic and analytical methods available for carbohydrate research and the problems associated with some undesirable properties of carbohydrates as drug candidates.

However, recent advances in the field of glycoscience have demonstrated that many of these problems can be circumvented with the development of new analytical and synthetic methods and new concepts for carbohydrate research. New developments in glycobiology research have tremendously increased our understanding of numerous carbohydrate-mediated biological processes and provided new targets for therapeutic discovery. Parallel progress in carbohydrate chemistry has created new tools for the practical synthesis of highly diverse and chemically complicated oligosaccharide structures that were otherwise almost impossible to access. Maybe the best highlights of this success were the Chemistry Wolf Prizes that have been awarded to Prof. Samuel Danishefsky (1996), to the late Prof. Raymond U. Lemieux (1999), and most recently to Prof. Chi-Huey Wong (2014) for their fundamental and seminal contributions to the study and synthesis of complex oligosaccharides and glycoproteins.

Thus, if a history of twentieth century science is ever written, one of the highlights may well be a chapter on the creation and development of glycoscience. It is not easy to find an area of scientific work that encompasses so many elements; highlighting just a few of them: great complexity and variety; challenge verging on impossibility; demand for both mental and manipulative rigor, as well as for dedication, persistence, and hard work; never-ending frontiers for discovery and never-ending advances in sophistication; unlimited opportunities for intellectual excitement; strong coupling not only with all areas of chemistry and biology, but also with medicine and therapeutic discovery; finally, great relevance to human well-being, health, and education.

This themed issue on Carbohydrates, comprised of eight review articles and 18 original research manuscripts, highlights recent advances in glycoscience in its broad sense to drug discovery. It does not focus on the chemistry or chemical biology only, but rather on the synergy between carbohydrate-based medicinal chemistry, chemical biology and biophysics as a key facet to carbohydrate-based drug discovery. The themes covered include bacterial adhesion-based therapeutics and various detection methods, neoglycosylation, new trends in aminoglycoside (AGs) research, targeting bacterial cell wall biosynthesis as a strategy for development of new antibiotics, targeting enzymes utilizing various carbohydrates for the discovery of novel therapeutics, and new trends in the discovery and development of carbohydrate-based vaccines.

Pera and Pieters (DOI: 10.1039/c3md00346a) review bacterial adhesion-based therapeutics and detection methods. Being an important first step towards bacterial infection, the bacterial adhesion process to host cells provides an attractive target for the discovery of new therapeutics. The possible benefits of such therapeutics include (i) activity against drug-resistant bacteria and (ii) reduced potential for generating new resistance. Recent progresses on the development of new carbohydrate-based adhesion inhibitors of the human pathogens uropathogenic Escherichia coli and Pseudomonas aeruginosa as well as the pig and zoonotic pathogen Streptococcus suis are highlighted. The recognition process responsible for adhesion has also been used for the detection and removal of bacteria by large multivalent molecules or nanoparticles. As a very nice complement to this review, Ernst and co-workers (DOI: 10.1039/c3md00093e) present an original research manuscript on a new class of potent FimH antagonists, the branched α-D-mannopyranosides, for the potential treatment of infections caused by uropathogenic E. coli.

In continuation of the concept of bacterial adhesion-based therapeutics, Barbier and co-workers (DOI: 10.1039/c4md00097h) report on the development of new types of glycopolymers as multivalent systems for lectin recognition. Using living anionic ring-opening polymerization, oligomers with single or geminal sugar substituents located on every third carbon alongside the macromolecular scaffold were obtained under mild conditions. The resulting glycopolymers have shown efficient binding potency towards concanavalin A. This proof-of-concept strategy is now used by this team to prepare glycopolymers with anti-adhesive properties towards bacterial lectins.

Another drug discovery approach, namely ‘neoglycosylation’, is reviewed by Goff and Thorson (DOI: 10.1039/c4md00117f). Focusing on the highly versatile chemoselective alkoxyamine-based ‘neoglycosylation’ reaction, the fundamentals of neoglycosylation and the subsequent development of a ‘neoglycorandomization’ platform to afford differentially-glycosylated libraries of plant-based and microbial-based natural products as well as small molecule-based drugs for drug discovery applications are discussed. It is apparent from this review that there is an exciting possibility that this approach can yield improvement of in vitro properties (solubility, potency) and, in some cases, alter the fundamental mechanism of action of a parent compound. Indeed, several in vivo studies described indicate neoglycosides to be stable when delivered via IP or IV injection and to display improved properties (PK, PD, and efficacy).

Surface plasmon resonance (SPR) analysers have emerged as powerful tools for the characterization of label-free biomolecular interactions, enabling the dynamics of complex formation and dissociation to be monitored in real time. A wide variety of sensor chips are currently available for many applications. Brogioni and Berti (DOI: 10.1039/c4md00088a) review the use of SPR technology for the characterization of bacterial polysaccharide antigens. Using selected examples, an overview of significant applications of conventional flow SPR to investigate the specific interactions of bacterial polysaccharide antigens is presented.

A series of four review articles describe AGs as potent molecular scaffolds with broad medicinal applications. Garneau-Tsodikova and co-workers (DOI: 10.1039/c4md00163j) thoroughly review the new trends observed in the use of AGs in the past decade, along with the current understanding of their mechanisms of action in various bacterial and eukaryotic cellular processes. A well-defined discussion is provided on AGs' antibacterial mode of action, along with a series of novel applications of AGs investigated during the last decade such as riboswitch binders, oncogenic microRNAs (miRNAs) targeting molecules, antileishmanial compounds, antifungal agents, amphiphiles, and as potential treatment of genetic diseases arising from premature termination codons (PTCs). An in-depth discussion towards alleviation of AG-induced ototoxicity is presented by highlighting the solely non-toxic AG apramycin and the prospective clinical candidate antibiotic plazomicin.

Chang and Takemoto (DOI: 10.1039/c4md00078a) review a particular area of AGs' perspective use by focusing on antifungal amphiphilic AGs. The discovery of amphiphilic kanamycins that are antifungal, but not antibacterial, and that inhibit the growth of fungi by perturbation of plasma membrane functions, is discussed. In addition, these new derivatives have been found to exhibit low toxicities against plant and mammalian cells, and as such offer new options for fighting fungal pathogens and are examples of reviving old drugs to confront new therapeutic challenges. Cationic amphiphiles with a different angle of therapeutic use are reviewed by Herzog and Fridman (DOI: 10.1039/c4md00012a). In this review, recent years' progress in the development of cationic amphiphiles with diverse chemical structures, including amphiphilic AGs, that target the bacterial membrane bilayer as a strategy for the development of antibiotics, is presented. Excellent discussions on the synthesis, antimicrobial activity, membrane selectivity and modes of action aspects are provided. By using isothermal titration calorimetry (ITC) and diffusion NMR studies, Schmidt-Lassen and Lindhorst (DOI: 10.1039/c4md00122b) nicely complement these studies on amphiphilic AGs by exploring the significance of the anomeric configuration and epimerization at the sugar ring of six octyl glycoside micelles.

In their review article, Shalev and Baasov (DOI: 10.1039/c4md00081a) specifically focus on recent progress in the redesign of AGs for improved PTC suppression activity and reduced toxicity. By covering recent achievements in the redesign strategies of AGs and the mechanisms of AG-induced ototoxicity, along with the structural research on the action of AGs in bacterial and eukaryotic cells, the authors also point to the necessity to further understand the molecular mechanisms by which AGs exert their biological activity in eukaryotic cells for further rational drug design.

One approach towards developing potent ribosome-targeting antibacterial agents with diminished toxicity lies in the high selectivity of these compounds towards bacterial versus eukaryotic, cytosolic, and mitochondrial ribosomes. The validity of this approach has recently been demonstrated by Böttger, Vasella, and co-workers for the natural AG antibiotic apramycin and for the 4′-ether and 4′,6′-benzylydene derivatives of paromomycin. Based on these excellent observations, Crich, Böttger, Vasella and co-workers (DOI: 10.1039/c4md00119b) describe in this issue the generation of a library of 4′-O-glycopyranosyl derivatives of paromomycin. While all new derivatives were less active than paromomycin itself, differences in activity were observed between the gluco-, manno-, and galactopyranosyl series and between individual anomers. This work suggests that further tuning on structure–activity–toxicity is required to achieve the desired analogues with high specificity and selectivity towards bacterial versus cytosolic and mitochondrial ribosomes.

Deoxyfluorination of AG antibiotics is another approach that has been previously employed to lower the toxicity of AG derivatives relative to their parent compounds. The observed lower toxicity was correlated with a diminution in the basicity of the amine group adjacent to the fluorine atom. Taking advantage of this property, Hanessian and co-workers (DOI: 10.1039/c4md00072b) explore the impact of deoxyfluorination of AGs at the 4′-position. Using various synthetic methodologies, some impressive synthetic observations were made, which finally led to the construction of the desired 4′-deoxy-4′-fluoro-neamine and 4′-deoxy-4′-fluoro-4′-epi-neamine.

Some AGs have also been found to target the HIV-1 transactivation response (TAR) RNA region of the HIV virus. By generating neomycin dimers with linkers varying in functionality and length, Arya and co-workers (DOI: 10.1039/c4md00165f) demonstrated that AG dimers could distinguish between the conformational differences of mutant TAR RNA structures.

Reid, Basu, and co-workers (DOI: 10.1039/c4md00127c) describe an interesting example towards the development of new antibiotics by targeting bacterial cell wall N-acetylglucosaminidases, representative enzymes of autolysins that act on the glycan backbone of bacterial peptidoglycan. A library of triazole derivatives of N-acetylglucosamine (GlcNAc) was generated and their ability to inhibit bacterial growth was examined. Interestingly, two compounds that show bacteriostatic activity against Gram-positive Bacillus were identified. This proof-of-concept report elegantly illustrates the potential of the approach for further research towards the discovery of novel antibiotics that target bacterial cell wall N-acetylglucosaminidases.

Another example of targeting bacterial cell wall biosynthesis as a strategy towards the development of new antibiotics is described by Li and Lowary (DOI: 10.1039/c4md00067f). In this work, a bifunctional galactofuranosyltransferase, GlfT2, one of two essential enzymes for mycobacterial cell wall biosynthesis, was chosen as a target of new antibiotics. Since glycoconjugates containing furanose residues are absent in humans, the glycosyltransferases involved in arabinogalactan biosynthesis are of interest as targets for new antibacterial agents. The authors took advantage of the fact that a sulfonium ion can mimic the putative oxocarbenium ion-like transition state of the enzyme-catalyzed reaction and synthesized a series of sulfonium ion compounds that contain moieties that mimic both the galactofuranose and uridine diphosphate domains of the donor substrate, uridine diphosphate galactofuranose, as potential inhibitors of GlfT2. Some of the compounds displayed promising inhibitory activity against the target enzyme.

Vidal and co-workers (DOI: 10.1039/c4md00063c) explore the inhibition of human O-GlcNAc transferase (OGT) by using substrate analogues in which the pyrophosphate moiety is replaced by pyridine tether. A series of such derivatives was prepared and their activity was evaluated against human OGT with complementary docking, binding, and cellular assays. Some of these structures showed moderate in vitro inhibition of the target OGT enzyme. Continuing on glycosyltransferases, Wagner and co-workers (DOI: 10.1039/c4md00077c) report on the development of an operationally simple, colorimetric assay protocol for the identification and evaluation of glycosyltransferase inhibitors. The authors took advantage of the previously reported work by Wu and co-workers who demonstrated that the formation of the nucleoside diphosphate during the glycosyltransferase reaction can be monitored with a phosphatase/malachite green detection system at 620 nm, and optimized it to establish a highly robust, inexpensive, and rapid assay protocol that can be used in a microplate format for high-throughput screening (HTS) campaigns.

Rodriguez and Kohler (DOI: 10.1039/c4md00164h) report on an important drug target, the mammalian O-GlcNAc hydrolase (OGA), known to remove O-GlcNAc from serine and threonine residues on intracellular glycoproteins. The authors provide insight into the active site of OGA by demonstrating that a C215A mutant of the enzyme is capable of removing an alkyl diazirine-modified O-GlcNAc (O-GlcNDAz) hydrolytically stable to the parent OGA. Cairo (DOI: 10.1039/c4md00089g) also reviews an important human enzyme, the neuramidase (hNEU), involved in several signaling pathways implicated in diabetes and cancer. A concise summary of the substrate specificity and the known inhibitors of the hNEU isoenzymes as well as advances towards development of isoenzyme-specific inhibitors are presented.

Barron and Murphy (DOI: 10.1039/c4md00074a) explore iminosugar scaffolds as potential inhibitors for the binding of somatostatin-14 to human recombinant somatostatin receptors (hSSTRs). For this purpose, a series of 1-deoxynojirimycin derivatives were synthesized and one of them was found to selectively bind to hSSTR5 over hSSTR4. The authors also used the ligand lipophilicity efficiency calculations and concluded that partial protection or leaving the hydroxyl groups of the iminosugar free could be important for drug discovery research based on sugar scaffolds.

The mezzettiasides are made up of a unique class of partially acetylated oligosaccharides, isolated from the stem bark of Mezzettia leptopoda, which has been long used in the folk medicinal tradition of the Malaysian island of Borneo. O'Doherty, Beuning, and co-workers (DOI: 10.1039/c4md00095a) report excellent data on the synthesis of ten members of the mezzettiaside family of natural products along with the synthesis of four new analogues and their evaluation for antibacterial and anticancer activity. Ten mezzettiaside analogues were synthesized in sufficient quantities for biological evaluation and SAR studies by using highly divergent de novo asymmetric synthesis. These efforts led to the discovery of previously unreported antibacterial activities for the mezzettiaside class of acylated oligo-rhamnoside natural products.

α-N-Acetylgalactosamine (α-GalNAc) residues are commonly found as constituents of mucin glycoproteins, and they are critical antigenic components of blood group A and of the sensory nerve structures of humans. Enzymatic removal of the antigenic α-GalNAc residue from blood group A cells has been studied as a potential route for the production of universal red blood cells (RBCs) for transfusion. Bennet and co-workers (DOI: 10.1039/c4md00104d) describe the cloning, expression, purification, and characterization of a recombinant glycoside hydrolase family 109 α-N-acetylgalactosaminidase from the pathogenic bacteria E. meningosepticum. Detailed kinetic analysis of the enzyme-catalyzed reaction led the authors to the conclusion that NAD⁺-mediated oxidation followed by an α,β-elimination gives a Michael acceptor intermediate, which then undergoes hydration along with proton and hydride transfer to generate 2-acetamido-2-deoxy-α-D-galactose as the reaction product.

Carbohydrates play relevant roles in cell–matrix interactions that are still far from being fully comprehended. In order to gain insight into cell responses to the grafted carbohydrates, Cipolla and co-workers (DOI: 10.1039/c4md00056k) describe the study of chemically modified collagen matrices exposing galactose moieties and their preliminary in vitro evaluation with selected cell lines. As a whole, the results presented demonstrate that the neoglycosylated collagen can be recognized as a preferential substrate for the growth of cells of the skeletal system. Prospectively, this chemical modification could be used to implement cell colonization of collagen-based scaffolds for tissue engineering approaches.

Liskamp and co-workers (DOI: 10.1039/c4md00013g) explore synthetic antifreeze glycopeptide-based polymers as potential cryoprotectants of sensitive tissues and cells. The target structures were synthesized from an azide/alkyne glycopeptide building block by partial reduction of the azide and subsequent copper catalyzed azide alkyne cycloaddition polymerization to obtain linear oligomers. To compare the activity with native antifreeze glycoproteins (AFGPs), a linear dodecapeptide, which had a comparable length to AFGP-8, the AFGP with the lowest molecular mass found in nature, was synthesized. While CD spectroscopy showed that the triazole-based compound possessed a similar secondary structure to that of the related amide-based carbon-linked AFGP tetramer based on AFGP-8, its ice recrystallization inhibition activity was only modest compared with AFGP-8 and a previously described carbon-linked AFGP analogue.

In addition to small molecule drugs, vaccines are powerful tools for disease prevention and therapy. Recent discovery of zwitterionic polysaccharides (ZPSs), a new class of bacterial polysaccharides characterized by an alternating zwitterionic charge motif on adjacent monosaccharides enabling them to stimulate T- and B-cell immune responses, has promoted extensive research endeavors towards the development of entirely carbohydrate-based vaccines. Towards these ends, Andreana and co-workers (DOI: 10.1039/c4md00038b) describe rigorous work towards the elucidation of structural features of an entirely carbohydrate cancer vaccine construct, a zwitterionic polysaccharide PS A1 from anaerobe Bacteroides fragilis, using CD and fluorescent labeling techniques. The results gleaned from this study argue that altering the structure of PS A1, without chemically modifying the electrostatic charge character, does not alter immune response/recognition in mice. These findings have important implications for the design of entirely carbohydrate-based vaccine constructs.

Finally, Huang and co-workers (DOI: 10.1039/c4md00103f) present an impressive study towards the development of carbohydrate antigen delivery system by employing water-soluble copolymers as potential anticancer vaccines. To overcome the poor immunogenicity of tumor associated carbohydrate antigens, a fully synthetic glycopolymer vaccine incorporating multiple Tn antigens and Th cell peptide epitopes was prepared, which elicited significant and long lasting anti-Tn IgG antibody titers. The antibodies recognized Tn antigens on tumor cells. These proof-of-principle experiments nicely illustrate the potential of further optimization of the glycopolymer construct to enhance the humoral responses against the tumor associated carbohydrate antigens.

We thank the Editors and the Publisher of MedChemComm for cooperation and continuous support. We also express our deepest thanks to all the authors for their efforts and valuable contributions to this special themed issue.

Sylvie Garneau-Tsodikova

Timor Baasov

Guest Editors

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