Computational Analysis of the Homologous Mouse Double RINGs Interface for Targeted Anticancer Drug Design

The new yearly 1.1 million cancer surges and considerably increasing mortality, post-decades of decline, require urgent and effective therapy. Targeted anticancer therapy is an attractive strategy for treating cancer by targeting critical receptors. Homologous Mouse Double-Minute RINGs (MDM2-MDMX RING) negatively regulate the p53 tumour-suppressing role by promoting proteasomal degradation. Interventively, Hinok., MMRi62, MMRi64, and MMRi71 exert a therapeutic downregulation against MDM2-MDMX RING heterodimer by activating p53’s apoptotic arm, but their binding mechanisms remain poorly understood due to the RING domain’s high structural flexibility and lack of co-crystal structures. This study reveals the underlying dynamic conformational stability and flexibility of the MDM_2G443T-MDMX interface, and the potential mechanistic inhibitor binding using the all-atom MD simulations. The MMRi64-bound and Hinok-bound systems exhibited the lowest RMSD values (2.28 Å and 2.34 Å, respectively), indicating significant protein stabilization. MMRi71-bound (1.03 Å) and MMRi64-bound (1.30 Å) complexes displayed the lowest mean RMSF values, demonstrating that these ligands lock the protein into a rigid conformation. Conversely, the MMRi62-bound system showed the highest average RMSD and the highest RMSF (1.74 Å), identifying it as the most flexible complex. The ligand-bound MDM2G443T-MDMX complex underwent notable secondary structure transitions. MMGB/PBSA calculations revealed favorable binding free energies (-22.69, -26.42, and -20.30 kcal/mol) for the key interactions. Arg444, Pro445, Lys473, and Pro476 provided the highest binding free energy contributions to the RING domains. Over the 200 ns simulation, Hinok demonstrated dual interactions with both MDM2G443T-MDMX RING domains, highlighting a potential dual-inhibition mechanism, whereas MMRis interacted selectively with MDM2.