Camptothecin inhibits the progression of NPC by regulating TGF-β-induced activation of the PI3K/AKT signaling pathway

Abstract

Nasopharyngeal carcinoma (NPC) is a type of cancer that is characterized by increased invasiveness, metastatic potential and tumor recurrence. Camptothecin has been demonstrated to exhibit anticancer activity. However, the potential underlying molecular mechanisms mediated by camptothecin in NPC cells remain elusive. In the present study, the efficacy of camptothecin for NPC was investigated in vitro and in vivo. Additionally, the potential signaling pathway mediated by camptothecin in NPC cells was also examined. The results indicated that the viability and aggressiveness of NPC cells were suppressed by camptothecin treatment in a dose-dependent manner. Camptothecin administration downregulated the expression levels of cell-cycle-associated proteins including cyclin 1, cyclin-dependent kinase (CDK)1 and CDK2 in NPC cells. Expression levels of migration-associated proteins including vimentin, fibronectin and epithelial cadherin were regulated by camptothecin treatment in NPC cells. Additionally, camptothecin inhibited the expression of transforming growth factor-β (TGF-β), phosphoinositide 3-kinase (PI3K) and protein kinase B (AKT), whereas TGF-β overexpression abrogated camptothecin-mediated inhibition of PI3K and AKT expression and camptothecin-mediated inhibition of the viability and aggressiveness of NPC cells. Camptothecin significantly inhibited tumor growth and increased survival times in a mouse model of cancer. In conclusion, these results indicate that camptothecin treatment may inhibit the viability of NPC cells and aggressiveness by regulating the TGF-β-induced PI3K/AKT signaling pathways, which in turn may be a potential molecular target for the treatment of NPC.

Keywords: camptothecin; nasopharyngeal carcinoma; phosphoinositide 3-kinase/protein kinase B; transforming growth factor-β.

Figure 1.

Camptothecin suppresses cellular viability and induces cell cycle arrest in NPC cells. (A) An MTT assay revealed that camptothecin inhibits the viability of HK1 and C666-1 cells in a dose-dependent (2, 6 and 10 µM) manner. (B) Camptothecin inhibits the viability of HK1 and C666-1 cells in a time-dependent (24, 48 and 72 h) manner as assessed using an MTT assay. (C) Flow cytometric analysis of the cell cycle demonstrated that camptothecin (6 µM) induces cell cycle arrest at S-phase in HK1 and C666-1 cells. (D) Western blot analysis indicated that camptothecin (6 µM) downregulates the expression of cell-cycle-associated proteins, cyclin 1, CDK1 and CDK2 in NPC cells. β-actin was used as a loading control. **P<0.01. NPC, nasopharyngeal carcinoma; NS, not significant; CDK, cyclin-dependent kinase.

Figure 2.

Camptothecin treatment inhibits migration and invasion of NPC cells. (A) At 48 h post-treatment, camptothecin (6 µM) significantly inhibits migration (B) and invasion of NPC cells. Scale bar, 50 µm. **P<0.01. (C) Western blot analysis revealed that camptothecin (6 µM) treatment downregulates the expression of vimentin and fibronectin, and upregulates the expression of E-cadherin in NPC cells. β-actin was used as a loading control. NPC, nasopharyngeal carcinoma.

Figure 3.

Camptothecin regulates the viability of NPC cells by regulating TGF-β-induced activation of the PI3K/AKT signaling pathway. (A) Western blot analysis revealed that camptothecin (6 µM) treatment inhibits the expression of TGF-β, PI3K and AKT in NPC cells. (B) Western blot analysis demonstrated that TGF-β overexpression abrogates camptothecin-mediated inhibition of PI3K, pPI3K pAKT and AKT expression in NPC cells. (C) An MTT assay revealed that TGF-β overexpression abolishes camptothecin-mediated inhibition of viability of NPC cells. (D) TGF-β overexpression abrogates camptothecin-mediated inhibition of migration (E) and invasion of NPC cells. β-actin was used as a loading control. **P<0.01. NPC, nasopharyngeal carcinoma; TGF-β, transforming growth factor-β; PI3K, phosphoinositide 3-kinase; AKT, protein kinase B; pPI3K, phosphorylated PI3K; pAKT, phosphorylated AKT; NS, not significant.

Figure 4.

Camptothecin treatment inhibits tumor growth and increases survival times in vivo. (A) Camptothecin treatment (10 mg/kg) inhibited tumor growth in tumor-bearing mice compared with the control group. (B) Camptothecin treatment (10 mg/kg) decreased TGF-β, PI3K and AKT expression in tumor sections as assessed by immunohistochemistry. (C) Camptothecin treatment increased the survival times of mice during the 120-day treatment. Scale bars, 20 µm. **P<0.01. TGF-β, transforming growth factor-β; PI3K, phosphoinositide 3-kinase; AKT, protein kinase B.