Vijayanand Chandrasekaran

Inhibition of bacterial adhesion to live human cells: Activity and cytotoxicity of synthetic mannosides

Bacterial adhesion to glycosylated surfaces is a key issue in human health and disease. Inhibition of bacterial adhesion by suitable carbohydrates could lead to an anti‐adhesion therapy as a novel approach against bacterial infections. A selection of five α‐mannosides has been evaluated as inhibitors of bacterial adhesion to the polysaccharide mannan, as well as to the surface of live human HT‐29 cells. Cell toxicity studies were performed to identify the therapeutic window for a potential in vivo‐application of the tested carbohydrates. A previously published mannosidic squaric acid diamide was shown to be exceptionally effective as inhibitor of the bacterial lectin FimH.

Neutrophil uptake of nitrogen-bisphosphonates leads to the suppression of human peripheral blood γδ T cells

Nitrogen-bisphosphonates (n-BP), such as zoledronate, are the main class of drugs used for the prevention of osteoporotic fractures and the management of cancer-associated bone disease. However, long-term or high-dose use has been associated with certain adverse drug effects, such as osteonecrosis of the jaw and the loss of peripheral of blood Vγ9Vδ2 T cells, which appear to be linked to drug-induced immune dysfunction. In this report we show that neutrophils present in human peripheral blood readily take up zoledronate, and this phenomenon is associated with the potent immune suppression of human peripheral blood Vγ9Vδ2 T cells. Furthermore, we found this zoledronate-mediated inhibition by neutrophils could be overcome to fully reconstitute Vγ9Vδ2 T cell proliferation by concomitantly targeting neutrophil-derived hydrogen peroxide, serine proteases, and arginase I activity. These findings will enable the development of targeted strategies to mitigate some of the adverse effects of n-BP treatment on immune homeostasis and to improve the success of immunotherapy trials based on harnessing the anticancer potential of peripheral blood γδ T cells in the context of n-BP treatment.

Switching of Bacterial Adhesion to a Glycosylated Surface by Reversible Reorientation of the Carbohydrate Ligand

The surface recognition in many biological systems is guided by the interaction of carbohydrate-specific proteins (lectins) with carbohydrate epitopes (ligands) located within the unordered glycoconjugate layer (glycocalyx) of cells. Thus, for recognition, the respective ligand has to reorient for a successful matching event. Herein, we present for the first time a model system, in which only the orientation of the ligand is altered in a controlled manner without changing the recognition quality of the ligand itself. The key for this orientational control is the embedding into an interfacial system and the use of a photoswitchable mechanical joint, such as azobenzene.

Sweet switches: azobenzene glycoconjugates synthesized by click chemistry

Azobenzene glycoconjugates can be switched between two isomeric states, E and Z, to change the spatial orientation of the conjugated carbohydrate ligands. Mono-, di- and trivalent azobenzene glycoconjugates were synthesized using click chemistry and their photochromic properties determined. Multivalency effects were observed in photoisomerisation.

Synthesis and testing of the first azobenzene mannobioside as photoswitchable ligand for the bacterial lectin FimH

Summary In order to allow spatial and temporal control of carbohydrate-specific bacterial adhesion, it has become our goal to synthesise azobenzene mannosides as photoswitchable inhibitors of type 1 fimbriae-mediated adhesion of E. coli. An azobenzene mannobioside 2 was prepared and its photochromic properties were investigated. The E→Z isomerisation was found to be highly effective, yielding a long-lived (Z)-isomer. Both isomers, E and Z, show excellent water solubility and were tested as inhibitors of mannoside-specific bacterial adhesion in solution. Their inhibitory potency was found to be equal and almost two orders of magnitude higher than that of the standard inhibitor methyl mannoside. These findings could be rationalised on the basis of computer-aided docking studies. The properties of the new azobenzene mannobioside have qualified this glycoside to be eventually employed on solid support, in order to fabricate photoswitchable adhesive surfaces.

Synthesis and Surface-Spectroscopic Characterization of Photoisomerizable glyco-SAMs on Au(111)

Photoisomerizable glyco-SAMs (self-assembled monolayers), utilizing synthetic azobenzene glycoside derivatives were fabricated. The ultimate goal of this project is to assay the influence of the 3D arrangement of sugar ligands on cell adhesion, and eventually make cell adhesion photoswitchable. However, it is a prerequisite for any biological study on the spatial conditions of carbohydrate recognition, that photoisomerization of the surface molecules can be verified. Here, we employed IRRAS and XPS to spectroscopically characterize glyco-SAMs. In particular and unprecedented to date, we prove reversible E→Z→E isomerization of azobenzene glycoside-terminated SAMs.

Synthesis and Photochromic Properties of Configurationally Varied Azobenzene Glycosides

Spatial orientation of carbohydrates is a meaningful parameter in carbohydrate recognition processes. To vary orientation of sugars with temporal and spatial resolution, photosensitive glycoconjugates with favorable photochromic properties appear to be opportune. Here, a series of azobenzene glycosides were synthesized, employing glycoside synthesis and Mills reaction, to allow “switching” of carbohydrate orientation by reversible E/Z isomerization of the azobenzene N=N double bond. Their photochromic properties were tested and effects of azobenzene substitution as well as the effect of anomeric configuration and the orientation of the sugars 2-hydroxy group were evaluated.

Synthesis of Bifunctional Azobenzene Glycoconjugates for Cysteine‐Based Photosensitive Cross‐Linking with Bioactive Peptides

Azobenzene linker molecules can be utilized to control peptide/protein function when they are ligated to appropriately spaced amino acid side chains of the peptide. This is because the photochemical E/Z isomerization of the azobenzene N=N double bond allows to switch peptide conformation between folded and unfolded. In this context, we have introduced carbohydrate-functionalized azobenzene derivatives in order to advance the biocompatible properties of azobenzene peptide linkers. Chloroacetamide-functionalized and O-allylated carbohydrate derivatives were synthesized and conjugated with azobenzene to achieve new bifunctional cross-linkers, in order to allow ligation to cysteine side chains by nucleophilic substitution or thiol-ene reaction, respectively. The photochromic properties of the new linker glycoconjugates were determined and first ligation reactions performed.

Novel synthesis of fluorochrome-coupled zoledronate with preserved functional activity on gamma/delta T cells and tumor cells

In addition to their effects on bone resorption, nitrogen-containing bisphosphonates (N-BP) selectively activate γδ T cells, an innate-like immune cell population with potent anti-tumor activity. N-BP stimulate γδ T cells through induction of intracellular accumulation of mevalonate pathway-derived pyrophosphates, which are strong and selective antigens for human γδ T cells. The most potent among several classes of N-BP is zoledronate (ZOL). To study the uptake of ZOL and its immunological consequences in the γδ T cell/tumor cell interplay, we have synthesized a novel fluorescently labeled ZOL derivative termed FluorZOL by covalently coupling ZOL to carboxyfluorescein succinidimyl ester. Here we describe in detail the synthesis of FluorZOL and we further show that FluorZOL is functionally fully active as revealed by the selective expansion of γδ T cells and the enhancement of tumor cell lysis by γδ T cells.