Design, synthesis, and biological evaluation of novel phenylindole derivatives as P-gp inhibitors to overcome multidrug resistance in MCF-7/ADR cells.

Overcoming multidrug resistance (MDR) remains a formidable obstacle in cancer chemotherapy, largely attributable to drug efflux mediated by the P-glycoprotein (P-gp) pump. To address this challenge, we designed and synthesized 27 novel phenylindole derivatives and systematically assessed their MDR-reversal activity in MCF-7/ADR cells. Among these compounds, Ina4 demonstrated potent reversal activity (RF = 229.4), exceeding that of the reference P-gp inhibitors verapamil (RF = 51.0) and cyclosporine A (RF = 103.3), while exhibiting low intrinsic cytotoxicity. Mechanistic investigations, including western blot and Rhodamine 123 (Rh123) accumulation assays, revealed that Ina4 effectively inhibits the efflux function of P-gp without altering its protein expression levels. Furthermore, molecular docking analysis indicated that Ina4 may bind to the active pocket of P-gp, primarily via π-π stacking interactions. Notably, in a 3D tumor spheroid model, co-administration of Ina4 with doxorubicin (DOX) resulted in significant suppression of spheroid growth. Collectively, these findings indicate that Ina4 is a promising P-gp inhibitor.