NPB304 inhibited the MAPK pathway in MCF-7/paclitaxel cells.

<p>Cell lysates were subjected to Western blotting. Relative protein levels were quantified by densitometry. (A) The MEK-ERK pathway was activated in MCF-7/paclitaxel cells compared with MCF-7 cells. (B) After treatment with 200 nM TPA (PMA) for 6 to 24 h, the MEK-ERK pathway was activated in MCF-7 cells, and p-ERK1/2 was overexpressed maximally at 24 h. MCF-7 cells were pretreated with paclitaxel and 200 nM TPA for 24 h and subsequently exposed to paclitaxel for 48 h. (C) The expression of p-ERK1/2 was decreased in a concentration-dependent manner when MCF-7/paclitaxel cells were treated with 10 to 50 µM U0126 for 24 h. Then, MCF-7/paclitaxel cells were treated with paclitaxel for 48 h after being exposed to paclitaxel and U0126 for 24 h. (D) NPB304 inhibited p-ERK in a dose-dependent manner in MCF-7/paclitaxel cells after treatment for 72 h. (E) MCF-7/paclitaxel cells were treated with 7.5 µM NPB304 for different time periods. (F) TPA partially rescued the sensitization effect of NPB304 in the MCF-7/paclitaxel cells. The resistant cells were treated with paclitaxel and NPB304 in the presence or absence of TPA (200 nM) for 72 h. (G) p-p38 was also overexpressed in MCF-7/paclitaxel cells compared with MCF-7 cells. The IC₅₀ of paclitaxel was decreased when MCF-7/paclitaxel cells were treated with paclitaxel for 48 h after being exposed to paclitaxel and the p38 inhibitor SB203580 for 24 h. NPB304 suppressed the expression of p-p38 in a dose-dependent manner in MCF-7/paclitaxel cells after treatment for 72 h. The expression is shown relative to actin. The experiments were performed three times. The data represent the mean ± SD. *p<0.05, **p<0.01, one-way ANOVA (n = 3).