Proposing novel MDM2 inhibitors: Combined physics-driven high-throughput virtual screening and in vitro studies.

The mouse double minute 2 (MDM2) protein acts as a negative regulator of the p53 tumor suppressor. It directly binds to the N terminus of p53 and promotes p53 ubiquitination and degradation. Since the most common p53-suppressing mechanisms involve the MDM2, proposing novel inhibitors has been the focus of many in silico and also experimental studies. Thus, here we screened around 500,000 small organic molecules from Enamine database at the binding pocket of this oncogenic target. The screening was achieved systematically with starting from the high-throughput virtual screening method followed by more sophisticated docking approaches. The initial high number of screened molecules was reduced to 100 hits which then were studied extensively for their therapeutic activity and pharmacokinetic properties using binary QSAR models. The structural and dynamical profiles of the selected molecules at the binding pocket of the target were studied thoroughly by all-atom molecular dynamics simulations. The free energy of the binding of the hit molecules was estimated by the MM/GBSA method. Based on docking simulations, binary QSAR model results, and free energy calculations, 11 compounds (E1-E11) were selected for in vitro studies. HUVEC vascular endothelium, colon cancer, and breast cancer cell lines were used for testing the binding affinities of the identified hits and for further cellular effects on human cancer cell. Based on in vitro studies, six compounds (E1, E2, E5, E6, E9, and E11) in breast cancer cell lines and six compounds (E1, E2, E5, E6, E8, and E10) in colon cancer cell lines were found as active. Our results showed that these compounds inhibit proliferation and lead to apoptosis.