Direct and Allosteric Inhibition of the FGF2/HSPGs/FGFR1 Ternary Complex Formation by an Antiangiogenic, Thrombospondin-1-Mimic Small Molecule.

Fibroblast growth factors (FGFs) are recognized targets for the development of therapies against angiogenesis-driven
diseases, including cancer. The formation of a ternary complex with the transmembrane tyrosine kinase receptors (FGFRs),
and heparan sulphate proteoglycans (HSPGs) is required for FGF2 pro-angiogenic activity. Here by using a combination of
techniques including Nuclear Magnetic Resonance, Molecular Dynamics, Surface Plasmon Resonance and cell-based binding
assays we clarify the molecular mechanism of inhibition of an angiostatic small molecule, sm27, mimicking the endogenous
inhibitor of angiogenesis, thrombospondin-1. NMR and MD data demonstrate that sm27 engages the heparin-binding site
of FGF2 and induces long-range dynamics perturbations along FGF2/FGFR1 interface regions. The functional consequence
of the inhibitor binding is an impaired FGF2 interaction with both its receptors, as demonstrated by SPR and cell-based
binding assays. We propose that sm27 antiangiogenic activity is based on a twofold–direct and allosteric–mechanism,
inhibiting FGF2 binding to both its receptors.