Flexible docking and scoring using the Internal Coordinate Mechanics software (ICM) was benchmarked for ligand binding mode prediction against the 85 co-crystal structures in the modified Astex data set. The ICM virtual ligand screening was tested against the 40 DUD target benchmarks and 11-target WOMBAT sets. The self-docking accuracy was evaluated for the top 1 and top 3 scoring poses at each ligand binding site with near native conformations below 2 Å RMSD found in 91% and 95% of the predictions, respectively. The virtual ligand screening using single rigid pocket conformations provided the median area under the ROC curves equal to 69.4 with 22.0% true positives recovered at 2% false positive rate. Significant improvements up to ROC AUC= 82.2 and ROC(2%)= 45.2 were achieved following our best practices for flexible pocket refinement and out-of-pocket binding rescore. The virtual screening can be further improved by considering multiple conformations of the target.

The study of allosteric functional modulation in dynamic proteins is attracting increasing attention. In particular, the discovery of new allosteric sites may generate novel opportunities and strategies for drug development, overcoming the limits of classical active-site oriented drug design. In this paper, we report on the results of a novel, ab initio, fully computational approach for the discovery of allosteric inhibitors based on the physical characterization of signal propagation mechanisms in proteins and apply it to the important molecular chaperone Hsp90. We first characterize the allosteric "hot spots" involved in interdomain communication pathways from the nucleotide-binding site in the N-domain to the distal C-domain. On this basis, we develop dynamic pharmacophore models to screen drug libraries in the search for small molecules with the functional and conformational properties necessary to bind these "hot spot" allosteric sites. Experimental tests show that the selected moelcules bind the Hsp90 C-domain, exhibit antiproliferative activity in different tumor cell lines, while not affecting proliferation of normal human cells, destabilize Hsp90 client proteins, and disrupt association with several cochaperones known to bind the N- and M-domains of Hsp90. These results prove that the hits alter Hsp90 function by affecting its conformational dynamics and recognition properties through an allosteric mechanism. These findings provide us with new insights on the discovery and development of new allosteric inhibitors that are active on important cellular pathways through computational biology. Though based on the specific case of Hsp90, our approach is general and can readily be extended to other target proteins and pathways.