Co-Inhibition of P-gp and Hsp90 by an Isatin-Derived Compound Contributes to the Increase of the Chemosensitivity of MCF7/ADR-Resistant Cells to Doxorubicin

Breast cancer is a complex and multi-drug resistant (MDR) disease, which could result in the failure of many chemotherapeutic clinical agents. Discovering effective molecules from natural products or by derivatization from known compounds is the interest of many research studies. The first objective of the present study is to investigate the cytotoxic combinatorial, chemosensitizing, and apoptotic effects of an isatin derived compound (5,5-diphenylimidazolidine-2,4-dione conjugated with 5-substituted isatin, named HAA(2021) in the present study) against breast cancer cells (MCF7) and breast cancer cells resistant to doxorubicin (MCF7/ADR) when combined with doxorubicin. The second objective is to investigate the binding mode of HAA(2021) withP-glycoprotein (P-gp) and heat shock protein 90 (Hsp90), and to determine whether their co-inhibition by HAA(2021) contribute to the increase of the chemosensitization of MCF7/ADR cells to doxorubicin. The combination of HAA(2021), at non-toxic doses, with doxorubicin synergistically inhibited the proliferation while inducing significant apoptosis in MCF7 cells. Moreover, HAA(2021) increased the chemosensitization of MCF7/ADR cells to doxorubicin, resulting in increased cytotoxicity/selectivity and apoptosis-inducing efficiency compared with the effect of doxorubicin or HAA(2021) alone against MCF7/ADR cells. Molecular modeling showed that two molecules of HAA(2021) bind to P-gp at the same time, causing P-gp inhibitory effect of the MDR efflux pump, and accumulation of Rhodamine-123 (Rho123) in MCF7/ADR cells. Furthermore, HAA(2021) stably interacted with Hsp90 alpha more efficiently compared with 17-N-allylamino-17-demethoxygeldanamycin (17-AAG), which was confirmed with the surface plasmon resonance (SPR) and molecular modeling studies. Additionally, HAA(2021) showed multi-target effects via the inhibition of Hsp90 and nuclear factor kappa B (NF-B) proteins in MCF7 and MCF7/ADR cells. Results of real time-PCR also confirmed the synergistic co-inhibition of P-gp/Hsp90 alpha genes in MCF7/ADR cells. Further pharmacokinetic and in vivo studies are warranted for HAA(2021) to confirm its anticancer capabilities.