Galectins, a family of endogenous
glycan-
binding proteins, play crucial roles in a broad range of physiological and
pathological processes.
Galectin-1 (Gal-1), a proto-type member of this family, is overexpressed in several
cancers and plays critical roles in tumor-immune escape, angiogenesis and
metastasis. Thus, generation of high-affinity
Gal-1 inhibitors emerges as an attractive therapeutic approach for a wide range of neoplastic conditions. Small-molecule
carbohydrate inhibitors based on
lactose (Lac) and
N-acetyllactosamine (LacNAc) structures have been tested showing different results. In this study, we evaluated Lac- and LacNAc-based compounds with specific chemical modifications at key positions as
Gal-1 ligands by competitive solid-phase assays (SPA) and isothermal titration calorimetry (ITC). Both assays showed excellent correlation, highlighting that
lactosides bearing bulky aromatic groups at the anomeric
carbon and
sulfate groups at the O3' position exhibited the highest binding affinities. To dissect the atomistic determinants for preferential affinity of the different tested
Gal-1 ligands, molecular docking simulations were conducted and
PRODIGY-LIG structure-based method was employed to predict binding affinity in
protein-
ligand complexes. Notably, calculated binding free energies derived from the molecular docking were in accordance with experimental values determined by SPA and ITC, showing excellent correlation between theoretical and experimental approaches. Moreover, this analysis showed that 3'-O-sulfate groups interact with residues of the
Gal-1 subsite B, mainly with Asn33, while the
ester groups of the aromatic anomeric group interact with Gly69 and Thr70 at
Gal-1 subsite E, extending deeper into the pocket, which could account for the enhanced binding affinity. This study contributes to the rational design of highly optimized
Gal-1 inhibitors to be further studied in
cancer models and other pathologic conditions.