Using insights from funnel metadynamics, a molecular dynamics protocol that provides a detailed representation of protein-ligand interactions, we investigated how a single heavy-atom modification can enhance the activity of an initial hit against human peroxiredoxin 5. This improvement was validated by NMR experiments and enzyme-inhibition assays. Our results illustrate how molecular dynamics simulations offer a rational framework for designing ligands with improved properties, starting from low affinity but selective hits with minimal structural modifications.

Using insights from funnel metadynamics, a molecular dynamics protocol that provides a detailed representation of protein-ligand interactions, we investigated how a single heavy-atom modification can enhance the activity of an initial hit against human peroxiredoxin 5. This improvement was validated by NMR experiments and enzyme-inhibition assays. Our results illustrate how molecular dynamics simulations offer a rational framework for designing ligands with improved properties, starting from low affinity but selective hits with minimal structural modifications.

Carbohydrate-protein interactions are important in cell-cell communication, signal transduction, cancer, or infection. Chemists have designed glycosylated multivalent systems to mimic these recognition phenomena and produce potent ligands of lectins with therapeutic applications. Dynamic combinatorial chemistry (DCC) provides access to libraries of glycosylated macrocycles equilibrating through reversible covalent bonds. This strategy can be applied to the rapid and efficient identification of multivalent glycoclusters by introducing a protein into the equilibrating library. This strategy allowed the identification of the best ligands for more than one lectin in a single experimental set up by using two simple 1,4-dithiophenol building blocks. Selection of the best binder by each lectin (ConA, LecA, and LecB) was accompanied by the amplification of glyco-dyn[3]arenes and glyco-dyn[4]arenes. These macrocycles could be synthesized, isolated, and displayed nanomolar dissociation constants. Furthermore, while no toxicity could be detected against human cells or bacteria, their anti-adhesive properties against Pseudomonas aeruginosa were confirmed through a virulence assay on human cells. Altogether, extremely simple 1,4-dithiophenol building blocks provided access to a large diversity of glycoconjugates that could be selected by a lectin in a simple experimental set up to identify glycoconjugates with potential anti-infectious applications, thus speeding up the discovery of potential new antibacterial treatments.

Carbohydrate-protein interactions are of prime importance in cell-cell communication, signal transduction, cancer, bacterial or viral infection. Chemists have designed multivalent systems to mimick these recognition phenomena and provide potent ligands of these proteins with foreseen therapeutic applications. Dynamic combinatorial chemistry provides access to a library of chemical species in equilibrium through reversible covalent bonds. This strategy can be readily applied to the rapid and efficient identification of multivalent glycoclusters by introducing a protein into the equilibrating library for the selection of the fittest glycocluster for this protein. 1,4-Dithiophenols conjugated to monosaccharides were equilibrated into dynamic combinatorial libraries providing a diverse mixture of glycoclusters. Selection of the best ligand for different lectins (ConA, LecA and LecB from Pseudomonas aeruginosa) could increase the concentration of glyco-dyn[3]arenes and glyco-dyn[4]arenes. A key aspect of this strategy is that multiplexing can be readily achieved by using two building blocks (galactosylated and fucosylated 1,4-dithiophenols) to interrogate several lectins at once in a single experiment. These macrocyclic glycoclusters could be synthesized, isolated, then evaluated as ligands of the lectins and displayed nanomolar dissociation constants. Furthermore, while no toxicity could be detected against human cells or bacteria, their evaluation as anti-adhesive agents could be confirmed through a virulence assay on human A549 lung epithelial cells.

Tetraphenylethylene (TPE) is fluorescent through aggregation induced emission (AIE) in water. Herein, TPE was used as the core of glycoclusters that target the bacterial lectins LecA and LecB of Pseudomonas aeruginosa. Synthesis of these TPE-based glycoclusters was accomplished by using azide-alkyne "click" chemistry. The AIE properties of the resulting glycoclusters could be readily verified, but imaging could not be pursued due to the overlap of the fluorescence signals from cells and bacteria. Nonetheless, the glycoclusters displayed nanomolar affinities toward LecA and LecB. Further evaluation in a cell-based anti-adhesive assay highlighted a limited decrease in adhesion (20%) for the fucosylated glycocluster. This confirmed that these TPE-based glycoclusters are indeed LecA and LecB high-affinity ligands. Nevertheless, the hypotheses involving their application in imaging or anti-adhesive therapy could not be verified.

The synthesis of eight perylenediimide-based glycoclusters was readily performed from hexa-and tetra-propargylated cores through azide-alkyne "click" conjugation. Variations in the carbohydrate epitope (Glc, Gal, Man, Fuc) and the linker arm provided molecular diversity. Interactions with LecA and LecB, two proteins involved in the adhesion of Pseudomonas aeruginosa to host tissues, were evaluated by micro-calorimetry (ITC). In both cases high affinities were obtained with K-d values in the nanomolar range. Further evaluation of their anti-adhesive properties using cultured epithelial cells demonstrated their potent anti-adhesive activities against Pseudomonas aeruginosa with only 30-40% residual adhesion observed. The fluorescence properties of the PDI core were then investigated by confocal microscopy on cell-bacteria cultures. However, the red fluorescence signal of the PDI-based glycocluster was too weak to provide significant data. The present study provides another type of anti-adhesive glycocluster against bacterial infection with a large aromatic PDI core.