Treatment with β-elemene combined with paclitaxel inhibits growth, migration, and invasion and induces apoptosis of ovarian cancer cells by activation of STAT-NF-κB pathway

Abstract

In this study, we aimed to analyze the anti-cancer effects of β-elemene combined with paclitaxel for ovarian cancer. RT-qPCR, MTT assay, western blot, flow cytometry, and immunohistochemistry were used to analyze in vitro and in vivo anti-cancer effects of combined treatment of β-elemene and paclitaxel. The in vitro results showed that β-elemene+paclitaxel treatment markedly inhibited ovarian cancer cell growth, migration, and invasion compared to either paclitaxel or β-elemene treatment alone. Results demonstrated that β-elemene+paclitaxel induced apoptosis of SKOV3 cells, down-regulated anti-apoptotic Bcl-2 and Bcl-xl gene expression and up-regulated pro-apoptotic P53 and Apaf1 gene expression in SKOV3 cells. Administration of β-elemene+paclitaxel arrested SKOV3 cell cycle at S phase and down-regulated CDK1, cyclin-B1, and P27 gene expression and apoptotic-related resistant gene expression of MDR1, LRP, and TS in SKOV3 cells. In vivo experiments showed that treatment with β-elemene+paclitaxel significantly inhibited ovarian tumor growth and prolonged the overall survival of SKOV3-bearing mice. In addition, the treatment inhibited phosphorylated STAT3 and NF-κB expression in vitro and in vivo. Furthermore, it inhibited migration and invasion through down-regulation of the STAT-NF-κB signaling pathway in SKOV3 cells. In conclusion, the data suggested that β-elemene+paclitaxel can inhibit ovarian cancer growth via down-regulation of the STAT3-NF-κB signaling pathway, which may be a potential therapeutic strategy for ovarian cancer therapy.

Figure 1. Dose-dependent assay of β-elemene (A) and paclitaxel (B) for SKOV3 cells growth. Data are reported as means±SD. *P<0.05, **P<0.01 (ANOVA followed by Tukey's multiple comparison post hoc test). ns: not significant.

Figure 2. Effect of β-elemene (100 mg/mL), paclitaxel (20 mg/mL), or their combination on cell growth of SKOV3 cells (A and B), and on cell cycle (C to E). F, Effect of β-elemene and/or paclitaxel on genes CDK1, cyclin-B1, and P27 expression in SKOV3 cells. Data are reported as means±SD. *P<0.05, **P<0.01 (ANOVA followed by Tukey's multiple comparison post hoc test). ns: not significant.

Figure 3. Effect of β-elemene (100 mg/mL), paclitaxel (20 mg/mL), or their combination on apoptosis of SKOV3 cells (A), and the relative mRNA expression of Bcl-2 and Bcl-xl (B), P53 and Apaf1 (C), and MDR1, LRP, and TS (D). Data are reported as means±SD. **P<0.01 (ANOVA followed by Tukey's multiple comparison post hoc test). ns: not significant.

Figure 4. Effect of β-elemene+paclitaxel on migration (A) and invasion (B) of SKOV3 cells (magnification bar, 100 μm). C and D, Relative E-cadherin and MTA3 mRNA (C) and protein (D) expression in SKOV3 cells after treatment with β-elemene and/or paclitaxel. Data are reported as means±SD. **P<0.01 (ANOVA followed by Tukey's multiple comparison post hoc test). ns: not significant.

Figure 5. Effect of β-elemene+paclitaxel on relative STAT3 and NF-κB mRNA (A) and protein (B) expression levels in SKOV3 cells. C, Effects of STAT3 overexpression (STAT3OP) and NF-κB expression and phosphorylation in SKOV3 cells. D, Effects of STAT3 overexpression (STAT3OP) on β-elemene+paclitaxel-regulated growth. Data are reported as means±SD. *P<0.05, **P<0.01 (ANOVA followed by Tukey's multiple comparison post hoc test). ns: not significant.

Figure 6. Effects of STAT3 overexpression (STAT3OP) on β-elemene+paclitaxel-regulated migration (A) and invasion (B) of SKOV3 cells. Data are reported as means±SD. **P<0.01 (ANOVA followed by Tukey's multiple comparison post hoc test). ns: not significant.

Figure 7. Effects of STAT3 overexpression (STAT3OP) on β-elemene+paclitaxel-regulated apoptosis of SKOV3 cells. Data are reported as means±SD. **P<0.01 (ANOVA followed by Tukey's multiple comparison post hoc test). ns: not significant.

Figure 8. Anti-cancer effect of β-elemene+paclitaxel in SKOV3-bearing mice. A, Tumor growth in SKOV3-bearing mice after receiving β-elemene and/or paclitaxel. B, Apoptotic tumor cells in tumor tissue after treatment with β-elemene and/or paclitaxel determined by TUNEL assay (magnification bar: 50x). C, Protein expression of STAT3 and NF-κB in tumor tissue after treatment with β-elemene and/or paclitaxel (magnification bar: 50x). D, Survival time of tumor-bearing mice after treatment with β-elemene and/or paclitaxel. Data are reported as means±SD. **P<0.01 (ANOVA followed by Tukey's multiple comparison post hoc test).