Thiodigalactoside–Bovine Serum Albumin Conjugates as High-Potency Inhibitors of Galectin-3: An Outstanding Example of Multivalent Presentation of Small Molecule Inhibitors

Abstract

Galectin inhibitors are urgently needed to understand the mode of action and druggability of different galectins, but potent and selective agents still evade researchers. Small-sized inhibitors based on thiodigalactoside (TDG) have shown their potential while modifications at their C3 position indicated a strategy to improve selectivity and potency. Considering the role of galectins as glycoprotein traffic police, involved in multivalent bridging interactions, we aimed to create multivalent versions of the potent TDG inhibitors. We herein present for the first time the multivalent attachment of a TDG derivative using bovine serum albumin (BSA) as the scaffold. An efficient synthetic method is presented to obtain a novel type of neoglycosylated proteins loaded with different numbers of TDG moieties. A polyethylene glycol (PEG)-spacer is introduced between the TDG and the protein scaffold maintaining appropriate accessibility for an adequate galectin interaction. The novel conjugates were evaluated in galectin binding and inhibition studies in vitro. The conjugate with a moderate density of 19 conjugated TDGs was identified as one of the most potent multivalent Gal-3 inhibitors so far, with a clear demonstration of the benefit of a multivalent ligand presentation. The described method may facilitate the development of specific galectin inhibitors and their application in biomedical research.

Biocompatible glyconanomaterials based on HPMA-copolymer for specific targeting of galectin-3

“…ASF is a multivalent glycoprotein carrying three LacNAc-capped triantennary N -glycans and it is commonly used as a standard ligand in binding assays with human Gal-3. For the aims of ELISA assay, we utilized a soluble His-tagged construct of human Gal-3, which was recombinantly expressed in E. coli and purified by immobilized metal-ion affinity chromatography as described before [ 28 , 36 ]. Gal-3 was incubated with increasing concentrations of the glyconanomaterials as competing ligands.…”

“…The production and purification of Gal-3 for ELISA assays was basically done as described before [ 28 , 36 ]. Briefly, Gal-3 was recombinantly expressed in E. coli Rosetta 2 (DE3) pLysS cells.…”

Galectins: Multitask signaling molecules linking fibroblast, endothelial and immune cell programs in the tumor microenvironment

“…Different galectin blocking compounds have been tested in vitro and/or in vivo, including specific anti-galectin neutralizing antibodies [80,81]. Moreover, glycan-based galectin inhibitors have also been investigated and proved to be promising lownanomolar galectin antagonists [148][149][150][151][152][153]. Furthermore, plant pectins and galactomannans have also shown antitumoral activities by virtue of their ability to bind galectins [154,155]; yet recent studies demonstrated that these naturally-occurring saccharides exhibit low inhibitory potency towards galectins CRDs [156].…”

Galectin-3 Is a Potential Mediator for Atherosclerosis

“…TD-139, a thiodigalactoside analogue, has been used as an inhaled powder for the treatment of idiopathic pulmonary fibrosis and is speculated to antagonize Gal-3 [92]. Multivalent attachment of a TDG derivative using the bovine serum albumin has been identified as one of the most potent Gal-3 inhibitors so far [93].…”