Elsevier

Biochemical Pharmacology

Volume 65, Issue 5, 1 March 2003, Pages 699-707
Biochemical Pharmacology

Commentary
“Multivalent” saccharides: development of new approaches for inhibiting the effects of glycosphingolipid-binding pathogens

https://doi.org/10.1016/S0006-2952(02)01553-8Get rights and content

Abstract

A number of diseases are initiated by the adherence of viruses, bacteria, or bacterial toxins to cell surface carbohydrates, a number of which are components of glycosphingolipids (GSLs). Studies of the binding of lectins indicated that many adhered weakly to monomeric carbohydrate ligands. The seminal observation that lectins adhered more strongly to a ligand with multiple carbohydrate binding sites initiated a plethora of studies designed to identify effective “multivalent” carbohydrate ligands for pathogens expressing multiple carbohydrate-binding sites. In addition to more completely defining ligand specificity of the carbohydrate-binding pathogen, identification of “multivalent” carbohydrate ligands has led to studies of their efficacy as pathogen inhibitors. This commentary focuses on pathogens that recognize the carbohydrate portion of GSLs. Because many GSL-binding pathogens have been shown to bind “multivalent” saccharides, approaches for identifying and preparing them as well as methods for characterizing their effectiveness as ligands are reviewed. Also discussed are areas of promise that should be investigated and pitfalls that might be encountered in the development of “multivalent” saccharides as pharmacologic agents.

Introduction

Three problems in the treatment of infectious diseases are: (a) the emergence of antibiotic-resistant strains of bacteria, (b) the existence of bacteria that secrete toxins, the activity of which is not affected by antibiotics, and (c) the lack of effective anti-viral agents. The first step in infection of a cell by viruses, bacteria, or bacterial toxins is their adherence to ligands on the cell surface. Over the past several decades, a number of pathogens have been shown to adhere to cell surface GSLs, and these interactions are the focus of this commentary. Table 1 contains examples of pathogens that adhere to GSLs, and Table 2 shows the saccharide portion of the GSLs discussed. So, how were the GSL ligands listed for the pathogens shown in Table 1 identified?

Section snippets

Identification of potential GSL ligands

A simple method for determining whether a GSL is needed for a pathogen to act upon its target cell is to determine the effect of inhibition of GSL synthesis on its ability to act upon cells that are normally susceptible to it. Treatment of cells with fumonisin B1, an inhibitor of sphingosine N-acyltransferase [1], blocks the synthesis of all GSLs. In contrast, inhibitors of ceramide:UDP glucosyltransferase, such as N-butyldeoxygalactonojirimycin [2], d,l-threo

Identification of the need for “multivalent” saccharides for optimum binding by a pathogen

Why should a “multivalent” saccharide be needed to optimize binding by a pathogen whose natural ligand is a GSL? The answer was provided, in part, by the observation that GSLs are found in clusters in lipid rafts on the cell surface [12]. The concept of multiple carbohydrate binding sites providing the basis for strong adherence to carbohydrate receptors evolved from studies of lectin binding. Those studies indicated that even though binding of a lectin to a single saccharide might be weak, its

Carriers for the preparation of “multivalent” saccharides

A number of proteins, e.g. bovine serum albumin, α-amylase, and lysozyme [22], have been used for the preparation of “multivalent” saccharides. Dendrimers, hyperbranched polymers having a known structure, were discovered in the early 1990s (e.g. Ref. [23]). These have provided researchers with more defined carriers for “multivalent” saccharide preparation. A number of different types of divergent dendrimers (dendrimers prepared outwardly from a central core) have been synthesized and used for

Synthesis of “multivalent” saccharides

To facilitate the linkage of saccharides and to retain the internal sugar in its cyclic conformation, a number of reactive groups have been added to, or left on the C(1) position of that sugar. Examples include the addition of phenylisothiocyanate [31], 3-mercaptopropionic acid [32], squaric acid diester [33], or allyl alcohol [28], and retention of a portion of the ceramide during formation of a “ceramide” acid [34]. Reductive amination [35] has also been used to link saccharides through their

Characterization of a “multivalent” ligand as a potential inhibitor of pathogen adherence

The efficacy of a “multivalent” ligand as a potential inhibitor of the binding of a pathogen is usually determined by measuring the affinity of the pathogen for it or by determining its ic50. Surface plasmon resonance (SPR), which optically measures changes in mass concentration of molecules at a biospecific interface, can be used to obtain real time measurements of binding affinities. Advantages of this approach are that measurements can be made using unlabeled protein, and only small amounts

Promising results

Rhesus monkeys are susceptible to colonization by Helicobacter pylori and can be experimentally colonized using strains of human origin. Over time, they develop symptoms similar to those seen in people [42]. Using monkeys colonized with H. pylori for a year or more, Mysore et al.[43] were able to cure 2 of 6 monkeys using oral administration of 3′-sialyllactose. Interestingly, bovine serum albumin carrying ∼20 3′-sialyllactose residues was found to have an ic50 that was about a thousand-fold

Problems that need to be addressed

From the foregoing discussion, it can be seen that a number of questions need to be addressed when designing a “multivalent” saccharide inhibitor. First and foremost is the problem of “multivalent” receptors. The molecules that mediate adherence of bacteria and viruses to their target cells are present in multiple copies on their surfaces, and toxins often express multiple binding sites. For a monovalent ligand to inhibit their adherence, it is necessary to have the ligand present at a

Conclusions

The use of “multivalent” saccharides to inhibit adherence of pathogens to their target cells is an area of research that has come into its own. This is due, in part, to advances in structure analysis and computer modeling that help enormously in designing effective “multivalent” ligands able to antagonize pathogen–host interactions (e.g. Refs. [29], [37]). The development of automated approaches to the chemical synthesis of oligosaccharides, coupled with the use of bioengineered bacteria to

Acknowledgements

Many thanks to Dr. Herbert C. Yohe, Dartmouth College, and Dr. Stanley J. Naides, this University, for their helpful comments about this manuscript. This work was supported, in part, by Grant NS40231 from the National Institutes of Health.

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