Ligand selection from the analysis of protein conformational substates: new leads targeting the N-terminal domain of Hsp90

The dynamic properties of proteins underlie every aspect of their functions in the cell. The atomistic description of protein motions and their inclusion in ligand selection processes may provide new opportunities for hit identification and drug discovery. Herein, we present a novel rational strategy that allowed us to computationally select new N-terminal targeted inhibitors of the molecular chaperone Hsp90 starting from the atomistic analysis of the conformational dynamics of the complex between the protein and its natural ligand ATP. First of all, we have identified the relevant representatives of distinct conformational substates of the Hsp90-ATP complex through the application of a novel structural clustering strategy and, for each of them, we have afterwards characterized the nucleotide-protein interactions to build a pharmacophore model recapitulating the binding hotspots conserved in different ensembles of protein conformations. The resulting pharmacophore has been finally used to screen a database of small molecules and allowed us to identify novel drug-like molecules with interesting activities against Hsp90 functions in experimental models of cancer cells. The results and the experimental validation of the selected molecules provide support for the feasibility of including protein flexibility in drug selection strategies through the characterization of relevant substates.