Selective mechanism of inhibitors to two bromodomains of BRD4 revealed by multiple replica molecular dynamics simulations and free energy analyses

Bromodomain-containing protein 4 (BRD4) is critical in cell cycle regulation and has emerged as a potential target for treatment of various cancers. BRD4 contains two bromodomains, namely BDl and BD2. Research suggests that selectively inhibiting BDl or BD2 may provide more effective treatment options. Therefore, understanding the selective mechanism of inhibitor binding to BDl and BD2 is essential for development of high selective inhibitors to BDl and BD2. Multiple replica molecular dynamics (MRMD) simulations are utilized to investigate the binding selectivity of inhibitors SG3-179, GSK778, and GSK620 for BDl and BD2. The results show that BDl has stronger structural flexibility than BD2, moreover BDl and BD2 exhibit different internal dynamics. The analyses of free energy landscapes reveal significant differences in the conformational distribution of BDl and BD2. Binding free energy predictions suggest that entropy changes, electrostatic interactions, and van der Waals interactions are key factors in the selective binding of BDl and BD2 by SG3-179, GSK778, and GSK620. The calculations of the energy contributions of individual residues demonstrate that residues (W81, W374), (P82, P375), (Q85, K378), (V87, V380), (192, 1385), (N93, G386), (194, 1387), (C136, C429), (N140, N433), (K141, P434), (D144, H437) and (1146, V439) corresponding to (BDl, BD2) generate significant energy difference in binding of SG3-179, GSK778, and GSK620 to BDl and BD2, and they can serve as effective targets for development of high selective inhibitors against BDl or BD2. The related information may provide significant theoretical guidance for improving the selectivity of inhibitors for BDl and BD2.