Abstract
The initial discovery phase of protein modulators, which consists of filtering molecular libraries and in vitro direct binding validation, is central in drug discovery. Thus, virtual screening of large molecular libraries, together with the evaluation of binding affinity by isothermal calorimetry, generates an efficient experimental setup. Herein, we applied virtual screening for discovering small molecule inhibitors of MDM2, a major negative regulator of the tumor suppressor p53, and thus a promising therapeutic target. A library of 20 million small molecules was screened against an averaged model derived from multiple structural conformations of MDM2 based on published structures. Selected molecules originating from the computational filtering were tested in vitro for their direct binding to MDM2 via isothermal titration calorimetry. Three new molecules, representing distinct chemical scaffolds, showed binding to MDM2. These were further evaluated by exploring structure-similar chemical analogues. Two scaffolds were further evaluated by de novo synthesis of molecules derived from the initial molecules that bound MDM2, one with a central oxoazetidine acetamide and one with benzene sulfonamide. Several molecules derived from these scaffolds increased wild-type p53 activity in MCF7 cancer cells. These set a basis for further chemical optimization and the development of new chemical entities as anticancer drugs.