Structure-based optimization of angiostatic agent 6DBF7, an allosteric antagonist of galectin-1

Galectin-1 (gal-1), which binds β-galactoside groups on various cell surface receptors, is crucial to cell adhesion and migration, and is found to be elevated in several cancers. Previously, we reported on 6DBF7, a dibenzofuran (DBF)-based peptidomimetic of the gal-1 antagonist anginex. In the present study, we used a structure-based approach to optimize 6DBF7. Initial NMR studies showed that 6DBF7 binds to gal-1 on one side of the β-sandwich away from the lectin’s carbohydrate binding site. Although an alanine scan of 6DBF7 showed that the two cationic groups (lysines) in the partial peptide are crucial to its angiostatic activity, it is the hydrophobic face of the amphipath that appears to interact directly with the surface of gal-1. Based on this structural information, we designed and tested additional DBF analogs. In particular, substitution of the C-terminal Asp for alanine and branched alkyl side chains (Val, Leu, Ile) for linear ones (Nle, Nva) rendered the greatest improvements in activity. Flow cytometry with gal-1−/− splenocytes showed that 6DBF7 and two of its more potent analogs (DB16 and DB21) can fully inhibit fluorescein isothiocyanate–gal-1 binding. Moreover, heteronuclear single-quantum coherence NMR titrations showed that the presence of DB16 decreases gal-1 affinity for lactose, indicating that the peptidomimetic targets gal-1 as a noncompetitive, allosteric inhibitor of glycan binding. Using tumor mouse models (B16F10 melanoma, LS174 lung, and MA148 ovarian), we found that DB21 inhibits tumor angiogenesis and tumor growth significantly better than 6DBF7, DB16, or anginex. DB21 is currently being developed further and holds promise for the management of human cancer in the clinic.