Structure-Guided Design of d-Galactal Derivatives with High Affinity and Selectivity for the Galectin-8 N-Terminal Domain

Abstract

Galectin-8 is a carbohydrate-binding protein that plays a crucial role in tumor progression and metastasis, antibacterial autophagy, modulation of the immune system, and bone remodeling. The design, synthesis, and protein affinity evaluation of a set of C-3 substituted benzimidazole and quinoline D-galactal derivatives identified a D-galactal-benzimidazole hybrid as a selective ligand for the galectin-8 N-terminal domain (galectin-8N), with a K d of 48 μM and 15-fold selectivity over galectin-3 and even better selectivity over the other mammalian galectins. X-ray structural analysis of galectin-8N in complex with one benzimidazole-and one quinoline-galactal derivative at 1.52 and 2.1 Å together with molecular dynamics simulations and quantum mechanical calculations of galectin-8N in complex with the benzimidazole derivative revealed orbital overlap between a NH LUMO of Arg45 with electron rich HOMOs of the olefin and O4 of the D-galactal. Such overlap is hypothesized to contribute to the high affinity of the D-galactal-derived ligands for galectin-8N. A (3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4sulfophenyl)-2H-tetrazolium) (MTS) assay evaluation of the D-galactal-benzimidazole hybrid and an analogous galactoside derivative on a panel of cell lines with MTS assay showed no effect on cell viability up to 100 μM concentration. A subsequent functional assay using the MDA-MB-231 cell line demonstrated that the D-galactal-benzimidazole hybrid and the analogous galactoside derivative reduced the secretion of the proinflammatory cytokines interleukin-6 (IL-6) and IL-8 in a dose-dependent manner. Therefore, these compounds represent potential probes for galectin-8N pharmacology investigations and possibly promising leads for the design and synthesis of potent and selective galectin-8 inhibitors as potential antitumor and anti-inflammatory agents.

Design and synthesis of novel 3-triazolyl-1-thiogalactosides as galectin-1, -3 and -8 inhibitors

“…13 C NMR (101 MHz, MeOD) d 158. 48, 143.12, 137.39, 131.81, 130.18, 129.24, 129.14, 123.45, 120.81, 114.36 (47). The compound was prepared according to general procedure A using 8 (45 mg, 0.14 mmol) and 21 (28 mg, 0.17 mmol).…”

“…gave product 44 as an off-white solid (36 mg, 47% yield). 1 (47). The compound was prepared according to general procedure A using 8 (45 mg, 0.14 mmol) and 21 (28 mg, 0.17 mmol).…”

Selective Galectin‐8N Ligands: The Design and Synthesis of Phthalazinone‐d‐Galactals

“…Using molecular dynamics (MD) simulations, we investigated the putative interactions that a dgalactal moiety carrying a phthalazinone heterocycle at position 3 might form in the galectin-8N binding site (Figure 3). The MD simulations of 4-phenylphthalazinone 5 in a d-galactal-based ligand-galectin-8N crystal structure (PDB accession code: 7AEN) [22] were run for 100 ns using Desmond in the Schrödinger suite. [28] During the simulations, we used light harmonic constraints (1 kcal mol À 1 Å À 2 ) on the oxygen at the C4 of the galactal and on all atoms in the backbone of the β-strands.…”

“…A visualisation of the structure-activity relationship (SAR) of the phthalazinone scaffold and its analogues is shown in Figure 4. Starting from the slightly galectin-8N-selective d- d-Galactal [22] 1300 � 72 1600 � 140 3300 � 650 4100 � 470 Lactose [16,34,35] 91 galactal, [22] introduction of the pyridazinone, the smallest fragment in the series, resulted in a two-fold increase in affinity for compound 13. Subsequent elaboration to the 1-phenylpyridazinone 14 and phthalazinone 16 gave an additional 2-and 4-fold increase in galectin-8N affinity, respectively.…”

“…However, it is a nearly equipotent ligand for galectin-3. [11] Recently, d-galactal analogue 2 was discovered, [22] in which the galactal moiety confers selectivity on galectin binding, but at the cost of affinity. The discovery of a selective tricyclic galactalbenzene hybrid 3 has been described, which has an affinity of 180 μM for galectin-8N.…”