Human Milk Oligosaccharides Multivalently Presented on Defined Synthetic Neo-Glycoproteins Are Nanomolar Ligands of Tandem-Repeat Galectins.

Galectins are small human proteins participating in inflammation processes, immune response, and cancerogenesis. Tandem-repeat galectins comprising Gal-4, Gal-8, and Gal-9 are a vital yet less studied part of the galectin fingerprint in cancer-related processes. The present work studies a library of prepared multivalent neo-glycoproteins decorated with poly--acetyllactosamine and human-milk-type oligosaccharides as ligands of this underexplored family of tandem-repeat galectins. A thorough binding evaluation by ELISA and biolayer interferometry was complemented with a detailed epitope mapping both from the galectin and the glycoconjugate viewpoints by nuclear magnetic resonance. The found interactions in the galectin binding site were correlated to data from molecular modeling. The present work reveals pioneer information on the binding of tandem-repeat galectins to multivalent glycoconjugates carrying complex carbohydrate ligands and represents an invaluable starting point for the development of new high-affinity tailored ligands of tandem-repeat galectins, needed both for diagnosis and therapy.