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Background:
Galectin-3 (Gal-3) is a promising target in cancer therapy with a high therapeutic potential due to its abundant localization within the tumor tissue and its involvement in tumor development and proliferation. Potential clinical application of Gal-3-targeted inhibitors is often complicated by their insufficient selectivity or low bioco...
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... 36 In the last few years, various multivalent carbohydrate-based or carbohydrate-decorated materials have been designed. 37,38 Many types of scaffolds have been used for the multivalent presentation, such as polymers, 30,39 calixarenes, 31,40 serum albumins, 23,41,42 peptides, 29,43,44 or dendrimers. 45−47 Dendrimers (DDMs) are regular, highly branched spherical macromolecules prepared by repetition of individual synthetic steps. ...
... We previously found that a triazole next to the carbohydrate unit may have a positive effect on the affinity, especially to Gal-3. 29,39 The interactions of Lac-CS-DDMs with human galectins (Gal-1, Gal-3, Gal-8, and Gal-9) were thoroughly studied using two complementary methods: (i) competitive enzyme-linked immunosorbent assay (ELISA), which assessed the ligand inhibitory potency, and (ii) dynamic light scattering (DLS), which correlated the Lac-CS-DDM-galectin aggregation behavior with an increased affinity. 29 Si spectra were referenced to external standard hexamethyldisilane (−19.87 ppm). ...
... The series A of Lac-CS-DDMs featuring a directly bound lactose showed a lower overall inhibitory potency than the series B with the triazole group adjacent to the glycan. This is in line with our previous studies where we found a considerable effect of the carbohydrate linker on the affinity to galectins�the direct N-triazole linker exhibited the best result with Gal-3 compared to the O-ethyltriazole and thiourea linker in a series of glycopolymers 39 and partially also in the study with glycoclusters, where, however, this effect was not apparent with Gal-1. 29 In our present study, the positive impact of the N-triazole linker adjacent to the carbohydrate moiety is evident for all galectin types, including tandem-repeat galectins. ...
Galectins, the glycan binding proteins, and their respective carbohydrate ligands represent a unique fundamental regulatory network modulating a plethora of biological processes. The advances in galectin-targeted therapy must be based on a deep understanding of the mechanism of ligand-protein recognition. Carbosilane dendrimers, the well-defined and finely tunable nanoscaffolds with low toxicity, are promising for multivalent carbohydrate ligand presentation to target galectin receptors. The study discloses a synthetic method for two types of lactose-functionalized carbosilane glycodendrimers (Lac-CS-DDMs). Furthermore, we report their outstanding, dendritic effect-driven affinity to tandem-type galectins, especially Gal-9. In the enzyme-linked immunosorbent assay, the affinity of the third-generation multivalent dendritic ligand bearing 32 lactose units to Gal-9 reached nanomolar values (IC50 = 970 nM), being a 1400-fold more effective inhibitor than monovalent lactose for this protein. This demonstrates a game-changing impact of multivalent presentation on the inhibitory effect of a ligand as simple as lactose. Moreover, using DLS hydrodynamic diameter measurements, we correlated the increased affinity of the glycodendrimer ligands to Gal-3 and Gal-8 but especially to Gal-9 with the formation of relatively uniform and stable galectin/Lac-CS-DDM aggregates.
... There was no positive contribution of the thiourea linker in this case, as the deprotected lactosyl amine showed IC 50 = 2.8 mM (see also the legend in Table 2), which was practically the same value as that of the free lactose (2.5 mM). This was also in accordance with our previous results with other galectins and a variety of linkers and carriers [32,36,37]. Hence, the considerable avidity increase should be attributed to the multivalency effect, possibly based on statistical rebinding, which may result in stabilized ligand-lectin crosslinked complexes [38]. ...
Galectins are carbohydrate-binding lectins that modulate the proliferation, apoptosis, adhesion, or migration of cells by cross-linking glycans on cell membranes or extracellular matrix components. Galectin-4 (Gal-4) is a tandem-repeat-type galectin expressed mainly in the epithelial cells of the gastrointestinal tract. It consists of an N- and a C-terminal carbohydrate-binding domain (CRD), each with distinct binding affinities, interconnected with a peptide linker. Compared to other more abundant galectins, the knowledge of the pathophysiology of Gal-4 is sparse. Its altered expression in tumor tissue is associated with, for example, colon, colorectal, and liver cancers, and it increases in tumor progression, and metastasis. There is also very limited information on the preferences of Gal-4 for its carbohydrate ligands, particularly with respect to Gal-4 subunits. Similarly, there is virtually no information on the interaction of Gal-4 with multivalent ligands. This work shows the expression and purification of Gal-4 and its subunits and presents a structure-affinity relationship study with a library of oligosaccharide ligands. Furthermore, the influence of multivalency is demonstrated in the interaction with a model lactosyl-decorated synthetic glycoconjugate. The present data may be used in biomedical research for the design of efficient ligands of Gal-4 with diagnostic or therapeutic potential.
... The literature has thus far reported on statistical copolymers of HPMA and APMA monomers, which were synthesized by free radical [29][30][31] or reversible addition − fragmentation chain transfer (RAFT) polymerization [20,21,[32][33][34]. The amino groups of the side chains of APMA monomer units are used for complexation with DNA and RNA [20,21,32] or have been used for covalent attachment of drugs and various targeting and biofunctional moieties [29,31,32,34]. ...
... The literature has thus far reported on statistical copolymers of HPMA and APMA monomers, which were synthesized by free radical [29][30][31] or reversible addition − fragmentation chain transfer (RAFT) polymerization [20,21,[32][33][34]. The amino groups of the side chains of APMA monomer units are used for complexation with DNA and RNA [20,21,32] or have been used for covalent attachment of drugs and various targeting and biofunctional moieties [29,31,32,34]. In our study, the p(HPMA-b-APMA) diblock copolymers were prepared by RAFT polymerization, which allows precise control over the molecular weight and polydispersity of the synthesized polymers [35]. ...
... Utilization of the functional diversity of galectins for medical and biotechnological applications of galectins has developed a considerable interest in the scientific community in the last few years. Due to the overexpression of galectins in cancer, many studies focus on the development of tailor-made galectin targeting and inhibiting agents [13][14][15]. In the medical field, galectin biosensors are developed as a diagnostic tool for the early-stage diagnosis of cancer or heart failure [16][17][18]. ...
... HGal-3 and HGal-8N were purified well with almost no galectin protein in the flow-through and wash fractions ( Figure S10A, lanes 5-8 and 9-12). As described above, the presence of HGal-8C after IMAC was detected by Western blot ( Figure S9) but not after application to fetuin-Sepharose ( Figure S10A, lane [13][14][15][16]. The fusion proteins HSDsRedMGal-1C2S and HSeGFPGal-8C did not bind to the fetuin-resin. ...
Galectins are β-galactosyl-binding proteins that fulfill essential physiological functions. In the biotechnological field, galectins are versatile tools, such as in the development of biomaterial coatings or the early-stage diagnosis of cancer diseases. Recently, we introduced galectin-1 (Gal-1) and galectin-3 (Gal-3) as fusion proteins of a His6-tag, a SNAP-tag, and a fluorescent protein. We characterized their binding in ELISA-type assays and their application in cell-surface binding. In the present study, we have constructed further fusion proteins of galectins with fluorescent protein color code. The fusion proteins of Gal-1, Gal-3, and Gal-8 were purified by affinity chromatography. For this, we have prepared glycoprotein affinity resins based on asialofetuin (ASF) and fetuin and combined this in a two-step purification with Immobilized Metal Affinity chromatography (IMAC) to get pure and active galectins. Purified galectin fractions were analyzed by size-exclusion chromatography. The binding characteristics to ASF of solely His6-tagged galectins and galectin fusion proteins were compared. As an example, we demonstrate a 1.6–3-fold increase in binding efficiency for HSYGal-3 (His6-SNAP-yellow fluorescent protein-Gal-3) compared to the HGal-3 (His6-Gal-3). Our results reveal an apparent higher binding efficiency for galectin SNAP-tag fusion proteins compared to His6-tagged galectins, which are independent of the purification mode. This is also demonstrated by the binding of galectin fusion proteins to extracellular glycoconjugates laminin, fibronectin, and collagen IV. Our results indicate the probable involvement of the SNAP-tag in apparently higher binding signals, which we discuss in this study.
... Utilization of the functional diversity of galectins for medical and biotechnological applications of galectins has developed a considerable interest in the scientific community in the last few years. Due to the overexpression of galectins in cancer, many studies focus on the development of tailor-made galectin targeting and inhibiting agents [13][14][15]. In the medical field, galectin biosensors are developed as a diagnostic tool for the early-stage diagnosis of cancer or heart failure [16][17][18]. ...
... HGal-3 and HGal-8N were purified well with almost no galectin protein in the flow-through and wash fractions (Fig. S10A, lanes 5-8 and 9-12). As described above, the presence of HGal-8C after IMAC was detected by Western blot (Fig. S9) but not after application to fetuin-Sepharose (Fig. S10A, lane [13][14][15][16]. The fusion proteins HSDsRedMGal-1C2S and HSeGFPGal-8C did not bind to the fetuin-resin. ...
Galectins are β-galactosyl-binding proteins that fulfill essential physiological functions. Recently, we introduced galectin-1 (Gal-1) and galectin-3 (Gal-3) as fusion proteins of a His6-tag, a SNAP-tag, and a fluorescent protein. We characterized their binding in ELISA-type assays and their application in cell-surface binding. In the present study, we have constructed further fusion proteins of galectins with fluorescent protein color code. The fusion proteins of Gal-1, Gal-3, and Gal-8 were purified by affinity chromatography. For this, we have prepared glycoprotein affinity resins based on asialofetuin (ASF) and fetuin and combined this in a two-step purification with Immobilized Metal Affinity chromatography (IMAC) to get pure and active galectins. Purified galectin fractions were analyzed by size-exclusion chromatography. The binding characteristics to ASF of solely His6-tagged galectins and galectin fusion proteins were compared. As an example, we demonstrate a 1.6-3-fold increase in binding efficiency for HSYGal-3 (His6-SNAP-yellow fluorescent protein-Gal-3) compared to the HGal-3 (His6-Gal-3). Our results reveal an apparent higher binding efficiency for galectin SNAP-tag fusion proteins compared to His6-tagged galectins which are independent of the purification mode. This is also demonstrated by the binding of galectin fusion proteins to extracellular glycoconjugates laminin, fibronectin, and collagen IV. Our results indicate the probable involvement of the SNAP-tag in apparently higher binding signals, which we discuss in this study.
... Considering that glycopolymers have a tunable capacity to target galectins, [246][247][248] and based on the versatility of polymeric materials that permit the creation of multivalent systems by anchoring a repeating unit in a polymeric backbone, some methacrylamide-type copolymers presenting lactose units have been reported as strong binders of Gal-3. 249 The HPMA copolymer is an interesting scaffold due to its biocompatibility and water solubility. ...
Human lectins are involved in a wide variety of biological processes, both physiological and pathological, which have attracted the interest of the scientific community working in the glycoscience field. Multivalent glycosystems have been employed as useful tools to understand carbohydrate-lectin binding processes as well as for biomedical applications. The review shows the different scaffolds designed for a multivalent presentation of sugars and their corresponding binding studies to lectins and in some cases, their biological activities. We summarise this research by organizing based on lectin types to highlight the progression in this active field. The paper provides an overall picture of how these contributions have furnished relevant information on this topic to help in understanding and participate in these carbohydrate-lectin interactions.
... Galectin-carbohydrate interactions are reinforced by a dense carbohydrate presentation on glycoproteins that can induce high avidity through multivalent effect (i.e., glycocluster effect) [13]. Synthetic glycoconjugates are based on many types of scaffolds, such as polymers [14,15], dendrimers [16][17][18], calixarenes [19], serum albumins [20,21] or peptides [22,23]. Peptide structures are suitable for the development of therapeutic biomaterials due to their natural origin, biodegradability, and non-inflammatory behavior. ...
... In previous studies, Elling's group [10,20,27] used a thiourea linker for conjugation of carbohydrate units to serum albumins. In another study, Bojarová et al. [14] presented a structure-affinity relationship study with four different spacers and two different carbohydrate functionalities -O-ethyltriazole and N-triazole. It was found that the direct N-triazole linker more efficiently inhibited Gal-3 as the affinity to Gal-3 decreased in the order N-triazole > O-ethyltriazole > thiourea. ...
... Thus, we revealed the influence of the spacer length on the affinity of the glycocluster to Gal-1 and Gal-3. The present study, in combination with the existing literature [14,20], gives a detailed picture of the effect of different types of linkers and multivalent structural presentations of galectin ligands, and points a route to nanomolar glycocluster inhibitors of galectins with biomedical potential. ...
Galectins are proteins of the family of human lectins. By binding terminal galactose units of cell surface glycans, they moderate biological and pathological processes such as cell signaling, cell adhesion, apoptosis, fibrosis, carcinogenesis, and metabolic disorders. The binding of monovalent glycans to galectins is usually relatively weak. Therefore, the presentation of carbohydrate ligands on multivalent scaffolds can efficiently increase and/or discriminate the affinity of the glycoconjugate to different galectins. A library of glycoclusters and glycodendrimers with various structural presentations of the common functionalized N-acetyllactosamine ligand was prepared to evaluate how the mode of presentation affects the affinity and selectivity to the two most abundant galectins, galectin-1 (Gal-1) and galectin-3 (Gal-3). In addition, the effect of a one- to two-unit carbohydrate spacer on the affinity of the glycoconjugates was determined. A new design of the biolayer interferometry (BLI) method with specific AVI-tagged constructs was used to determine the affinity to galectins, and compared with the gold-standard method of isothermal titration calorimetry (ITC). This study reveals new routes to low nanomolar glycoconjugate inhibitors of galectins of interest for biomedical research.
... Statistical copolymers were prepared by reversible addition-fragmentation chain transfer (RAFT) polymerization of HPMA with respective monomers as follows: MA-AP-TT was employed for P0a, P0b, P1, and P5 copolymers; APMA-Boc for P2 [31]; MA-AHcholesterol for P3; MA-AH-cholesterol and APMA-Boc for P4. The chain transfer agent CTA-AIBN was used for all precursors except for P2, in which CTA-ACVA was employed instead. ...
Photo/radiosensitizers, such as octahedral molybdenum clusters (Mo6), have been intensively studied for photodynamic applications to treat various diseases. However, their delivery to the desired target can be hampered by its limited solubility, low stability in physiological conditions, and inappropriate biodistribution, thus limiting the therapeutic effect and increasing the side effects of the therapy. To overcome such obstacles and to prepare photofunctional nanomaterials, we employed biocompatible and water-soluble copolymers based on N-(2-hydroxypropyl)methacryla-mide (pHPMA) as carriers of Mo6 clusters. Several strategies based on electrostatic, hydrophobic, or covalent interactions were employed for the formation of polymer-cluster constructs. Importantly, the luminescent properties of the Mo6 clusters were preserved upon association with the polymers: all polymer-cluster constructs exhibited an effective quenching of their excited states, suggesting a production of singlet oxygen (O2(1 Δg)) species which is a major factor for a successful photodynamic treatment. Even though the colloidal stability of all polymer-cluster constructs was satisfactory in deionized water, the complexes prepared by electrostatic and hydrophobic interactions underwent severe aggregation in phosphate buffer saline (PBS) accompanied by the disruption of the cohesive forces between the cluster and polymer molecules. On the contrary, the conjugates prepared by covalent interactions notably displayed colloidal stability in PBS in addition to high luminescence quantum yields, suggesting that pHPMA is a suitable nanocarrier for molybdenum cluster-based photosensitizers intended for photodynamic applications.
... Due to the differential expression of some galectins in health and disease, the synthesis and development of new galectin-targeted ligands are gaining interest as potential diagnostic markers or therapeutics. The synthesis and structure affinity of the gal-3 specific epitope Gal-NAcβ1,4GlcNAc (LacdiNAc) decorating a nanostructured N-(2-hydroxypropyl)methacrylamide (HPMA) has recently been investigated [198]. LacdiNAc was successfully synthesized under the catalysis of mutant β-N-acetylhexosaminidases and used to decorate the HPMA nanocarriers. ...
... Gal-3 affinity in low μM range [198] Propargylglycine-N-carboxyanhydride (PG-NCA) to benzyl-l-glutamate-Ncarboxyanhydride (BLG-NCA) and poly (γ-benzyl-l-glutamate)-blockgalactan Amphiphilic nanostructures exhibiting lactose or galactan NA Gal-3 binding by multivalency effect ...
Among all the biological entities involved in the immune response, galectins, a family of glycan-binding proteins, have been described as key in immune cell homeostasis and modulation. More importantly, only some galectin family members are crucial in the resolution of inflammation, while others perpetuate the immune response in a pathological context. As they are expressed in most major diseases, their potential as targets for new therapies seems promising. Most of the galectin family members' ubiquitous expression points to the need for targeted treatments to ensure effectiveness. Engineered biomaterials are emerging as a promising method to improve galectin-targeted strategies' therapeutic performance. In this review, we provide an overview of the role of galectins in health and disease and their potential as therapeutic targets, as well as the state-of-the-art and future directions of galectin-targeted biomaterials.
... The used biomimetic strategies provide the possibility of influencing specific tissue or cellular activities. [22][23][24] For example, the controlled inhibition of a selected functional promotor [25] or receptor clustering can be used to make the diseased tissue more sensitive to a therapeutic agent. [26,27] Thus, macromolecules decorated with targeting ligands gained attention due to their capacity to bind specific receptors in the cell surface of target tissues, resulting in a higher cellular uptake. ...
... [70] Our group synthesized various HPMA-based glycopolymers by RAFT polymerization followed by the attachment of various functionalized saccharides to the polymer carrier by an azidealkyne click reaction or via aminolysis, and their potential as multivalent inhibitors towards Gal-1 or Gal-3 was evaluated. [24,25,36] N,N´-diacetyllactosamine (GalNAc 1,4GlcNAc, LacdiNAc), a selective carbohydrate epitope for Gal-3 but not for Gal-1, was bound via various spacers and with various loadings. It was found that their affinity to Gal-3 increased up to a certain glycan epitope content; a further addition of saccharide did not enhance the multivalent effect. ...
... Moreover, the binding affinity to Gal-3 was improved by the presence of an aromatic moiety directly attached to the glycan. [25] Another topic to solve is the similar binding specificity of Gal-1 and Gal-3, which leads to a challenging preparation of selective inhibitors. [128] Aiming to address this issue, Nacetyllactosamine (Gal 4GlcNAc; LacNAc) was bound to HPMAbased copolymers, as LacNAc is a common disaccharide ligand for both Gal-1 and Gal-3. ...
Inside Front Cover: The precise biorecognition of natural binding motifs by multiple vectors on the polymer construct is a crucial part of the designing of polymer-based drug-free macromolecular therapeutics, a novel class of therapeutics with tailored macromolecular design. It leads to the induction of cancer cell apoptosis or depletion of the efficacy of tumor-associated proteins. This is reported by Tomáš Etrych and co-workers in article 2100135.
