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Lectins are involved in a wide range of carbohydrate mediated recognition processes. Therefore, the availability of highly performant fluorescent tools tailored for lectin targeting and able to efficiently track events related to such key targets is in high demand. We report here on the synthesis of the glyco-BODIPYs 1 and 2, based on the efficient combination of a Heck-like cross coupling and a Knoevenagel condensation, which revealed efficient in addressing lectins. In particular, glyco-BODIPY 1 has two glycosidase stable C-mannose residues, which act as DC-SIGN (dendritic cell-specific intercellular adhesion molecule-3-grabbing non-integrin) targeting modules. By using live-cell fluorescence microscopy, we proved that BODIPY-mannose 1 was efficiently taken up by immune cells expressing DC-SIGN receptors. Super-resolution stimulated emission depletion (STED) microscopy further revealed that the internalized 1 localized in membranes of endosomes, proving that 1 is a reliable tool also in STED applications. Of note, glyco-BODIPY 1 contains an aryl-azido group, which allows further functionalization of the glycoprobe with bioactive molecules, thus paving the way for the use of 1 for tracking lectin-mediated cell internalization in diverse biological settings., (Copyright © 2021 Elsevier Inc. All rights reserved.)

Study of interaction of mannose-based ligands with receptor DC-SIGN using high resolution NMR in combination with molecular modelling showed that four α-d-mannoside ligands interact with the binding site predominantly through the mannose moiety. The other two aromatic groups that are bound to α-d-mannose through a glycerol linker demonstrate interaction that can be related to their substitution pattern. Ligand with naphthyl and meta-substituted phenyl ring exhibited the most favourable binding characteristics. In addition to the predicted hydrophobic interactions of aromatic moieties our results propose new contacts of substituted phenyl moiety in the more polar area of the flat binding site of DC-SIGN and thus offer new possibilities in further designing of novel, more potent DC-SIGN antagonists.

Five docking tools, namely AutoDock, FRED, CDOCKER, FlexX and GOLD, have been critically examined, with the aim of selecting those most appropriate for use as docking tools for docking molecules to the lectin dendritic cell-specific intercellular adhesion molecule-3-grabbing non-integrin (DC-SIGN). This lectin has been selected for its rather non-druggable binding site, which enables complex interactions that guide the binding of the core monosaccharide. Since optimal orientation is crucial for forming coordination bonds, it was important to assess whether the selected docking tools could reproduce the optimal binding conformation for several oligosaccharides that are known to bind DC-SIGN. Our results show that even widely used docking programs have certain limitations when faced with a rather shallow and featureless binding site, as is the case of DC-SIGN. The FRED docking software (OpenEye Scientific Software, Inc.) was found to score as the best tool for docking ligands to DC-SIGN. The performance of FRED was further assessed on another lectin, Langerin. We have demonstrated that this validated docking protocol could be used for docking to other lectins similar to DC-SIGN.

Dendritic cell-specific, intercellular adhesion molecule-3-grabbing non-integrin (DC-SIGN) is a C-type lectin expressed specifically on dendritic cells. It is a primary site for recognition and binding of various pathogens and thus a promising therapeutic target for inhibition of pathogen entry and subsequent prevention of immune defense cell infection. We report the design and synthesis of d-mannose-based DC-SIGN antagonists bearing diaryl substituted 1,3-diaminopropanol or glycerol moieties incorporated to target the hydrophobic groove of the receptor. The designed glycomimetics were evaluated by in vitro assay of the isolated DC-SIGN extracellular domain for their ability to compete with HIV-1 gp120 for binding to the DC-SIGN carbohydrate recognition domain. Compounds 14d and 14e, that display IC50 values of 40 μM and 50 μM, are among the most potent monovalent DC-SIGN antagonists reported. The antagonistic effect of all the synthesized compounds was further evaluated by a one-point in vitro assay that measures DC adhesion. Compounds 14d, 14e, 18d and 18e were shown to act as functional antagonists of DC-SIGN-mediated DC adhesion. The binding mode of 14d was also studied by molecular docking and molecular dynamics simulation, which revealed flexibility of 14d in the binding site and provides a basis for further optimization., (Copyright © 2014 Elsevier Masson SAS. All rights reserved.)

DC-SIGN is a C-type lectin selectively expressed by certain types of DCs, including monocyte-derived DCs. Many reports have described the impact of DC-SIGN engagement with concomitant TLR signaling in tailoring of the DC maturation process, but so far, none has addressed the importance of DC-SIGN engagement during their differentiation from blood progenitors. We therefore examined the role of DC-SIGN engagement limited to the stage of IL-4-guided differentiation of DCs from human peripheral blood monocytes but not during maturation. We used two different anti-DC-SIGN antibodies with reported DC-SIGN-engaging activities. In cultures with DC-SIGN ligands, the resulting iDCs displayed abrogated expression of differentiation markers CD1a and DC-SIGN. Without further DC-SIGN activation, such DCs matured with low CD80/CD86 and high ILT3 expression, along with the appearance of macrophage marker CD14. Additionally, treated DCs indicated a tolerogenic potential by possessing a low, allostimulatory capacity and inducing naïve, allogeneic CD4(+) T cells to produce low levels of IFN-γ. Upon activation, IL-10 production was greatly increased by such DCs; however, the use of IL-10-blocking antibodies could not completely reverse alternative DC activation. This suggests an alternative activation response that is a result of a different elementary state of DCs generated with concomitant ligation of DC-SIGN. During differentiation, IL-4-induced pSTAT6 was reduced by DC-SIGN ligands. Furthermore, during LPS-induced maturation, treated DCs displayed lowered activation levels of p38 MAPK, STAT1, as well as STAT6, compared with controls. Collectively, evidence is presented confirming a crucial role for DC-SIGN signaling in DC generation from monocytes.

In this article, we describe the design, synthesis and activity evaluation of glycomimetic DC-SIGN antagonists, that use a mannose residue to anchor to the protein carbohydrate recognition domain (CRD). The molecules were designed from the structure of the known pseudo-mannobioside antagonist 1, by including additional hydrophobic groups, which were expected to engage lipophilic areas of DC-SIGN CRD. The results demonstrate that the synthesized compounds potently inhibit DC-SIGN-mediated adhesion to mannan coated plates. Additionally, in silico docking studies were performed to rationalize the results and to suggest further optimization.

We report a new dendritic cell adhesion assay, using either immature or mature dendritic cells, for identifying functional dendritic cell-specific ICAM-3-grabbing nonintegrin (DC-SIGN) inhibitors. Because immature dendritic cells are responsible for pathogen binding and invasion, this in vitro assay provides an important link between in vitro results and pathogen-based in vivo assays. Furthermore, this assay does not require laborious expression, refolding, and purification of DC-SIGN carbohydrate recognition domain or extracellular domain as receptor-based assays. The assay power evaluated with Z and Z' parameters enables screening of compound libraries and determination of IC(50) values in the first stage of DC-SIGN inhibitor development., (Copyright 2010 Elsevier Inc. All rights reserved.)

The dendritic cell-specific intercellular adhesion molecule-3-grabbing non-integrin (DC-SIGN) is a type II C-type lectin whose expression is restricted to the most potent antigen-presenting cells (APCs), the dendritic cells (DCs). In recent years, DC-SIGN has gained an exponential increase in attention because of its involvement in multiple aspects of immune function. Besides being an adhesion molecule, particularly in binding ICAM-2 and ICAM-3, it is also crucial in recognizing several endogenous and exogenous antigens. Additionally, the intracellular domain of DC-SIGN includes molecular motifs, which enable the activation of signal transduction pathways involving Raf-1 and subsequent modulation of DC-maturation status, through direct modification of nuclear factor Nf-kappaB in DCs. Upon DC-SIGN engagement by mannose- or fucose-containing oligosaccharides, the latter leads to a tailored Toll-like receptor signalling, resulting in an altered DC-cytokine profile and skewing of Th1/Th2 responses. In this article, we will discuss recent advances on a broad perspective concerning DC-SIGN structure, signalling and immune function., (Copyright 2010 Elsevier Inc. All rights reserved.)

The dendritic cell-specific intercellular adhesion molecule (ICAM) 3-grabbing nonintegrin (DC-SIGN) is a C-type lectin that appears to perform several different functions. Besides mediating adhesion between dendritic cells and T lymphocytes, DC-SIGN recognizes several pathogens some of which, including HIV, appear to exploit it to invade host organisms. The intriguing diversity of the roles attributed to DC-SIGN and their therapeutic implications have stimulated the search for new ligands that could be used as biological probes and possibly as lead compounds for drug development. The natural ligands of DC-SIGN consist of mannose oligosaccharides or fucose-containing Lewis-type determinants. Using the known 3D structure of the Lewis-x trisaccharide, we have identified some monovalent alpha-fucosylamides that bind to DC-SIGN with inhibitory constants 0.4-0.5 mM, as determined by SPR, and have characterized their interaction with the protein by STD NMR spectroscopy. This work establishes for the first time alpha-fucosylamides as functional mimics of chemically and enzymatically unstable alpha-fucosides and describes interesting candidates for the preparation of multivalent systems able to block the receptor DC-SIGN with high affinity and with potential biomedical applications.