Overexpression of FOXM1 is associated with metastases of nasopharyngeal carcinoma

Abstract

Background: The forkhead box M1 (FOXM1) transcription factor plays an important role in the metastases of many cancers. Down-regulation of FOXM1 by its inhibitor, thiostrepton, can inhibit the metastatic potential of some cancers; however, there are few studies regarding the functional significance of FOXM1 and thiostrepton in the metastases of nasopharyngeal carcinoma (NPC) and the underlying mechanism.

Methods: Expression of FOXM1 in NPC, normal nasopharyngeal tissues, a NPC cell line (C666-1), and a nasopharyngeal epithelial cell line (NP69) was investigated by immunohistochemical staining, qRT-PCR, and Western blot. The correlation between FOXM1 expression and the clinical characteristics of patients was analyzed. Moreover, the effects of thiostrepton on expression of FOXM1 in C666-1 and NP69 cells, and the invasion and migration ability of C666-1 cells were examined. The expressions of MMP-2, MMP-9, fascin-1, ezrin, and paxillin were determined after treatment with thiostrepton.

Results: FOXM1 was overexpressed in NPC and C666-1 cells compared with normal nasopharyngeal tissues and NP69 cells. Overexpression of FOXM1 was associated with lymph node metastasis and advanced tumor stage. Moreover, thiostrepton inhibited expression of FOXM1 in C666-1 cells in a dose-dependent manner, but had a minimal effect on NP69 cells. Thiostrepton inhibited the migration and invasion ability of C666-1 cells by down-regulating the expression of MMP-2, MMP-9, fascin-1, and paxillin.

Conclusions: Overexpression of FOXM1 is associated with metastases of NPC patients. Thiostrepton inhibits the metastatic ability of NPC cells by down-regulating the expression of FOXM1, MMP-2, MMP-9, fascin-1, and paxillin.

Keywords: Forkhead box M1 transcription factor (FOXM1); metastases; nasopharyngeal carcinoma; oncogene; thiostrepton.

Figure 1.

Expression of FOXM1 in NPC and normal nasopharyngeal tissues (NT). A: Overexpression of FOXM1 in NPC and low expression in NT as evidenced by immunohistochemical staining. B: Relative FOXM1 mRNA levels in NPC and NT were determined by qRT-PCR. C: Overexpression of FOXM1 in NPC and low expression in NT were determined by Western blot. Experiments were independently repeated three times. *p < 0.05.

Figure 2.

Thiostrepton significantly reduces the expression of FOXM1 mRNA and protein in a NPC cell line other than a nasopharyngeal epithelial cell line. A: The expression of FOXM1 mRNA in C666-1 cells was inhibited after thiostrepton treatment at 2, 4, or 6 µM for 48 h. B: Thiostrepton down-regulated the expression of FOXM1 protein in C666-1 cells. C: The expression of FOXM1 mRNA in NP69 cells was lower than in C666-1 cells and was less affected by thiostrepton. Experiments were independently repeated three times. *p < 0.05.

Figure 3.

Thiostrepton reduces the invasion and migration activities of NPC cells. A: Invasion and migration assays were used to detect the invasion and migration ability of C666-1 cells following treatment with 2, 4, or 6 µM thiostrepton for 48 h. B: Manual counts of invasion and migration experiments. Experiments were independently repeated three times. *p < 0.05.

Figure 4.

Thiostrepton reduces mRNA and protein expressions of metastatic associated genes in NPC cells. A: Relative MMP-2, MMP-9, fascin-1, ezrin, and paxillin mRNA levels were determined by qRT-PCR. B: An ELISA assay was used to detect protein levels of MMP-2 and MMP-9. C: Fascin-1, ezrin, and paxillin protein levels were determined by Western blot. Experiments were independently repeated three times. *p < 0.05.