Icotinib inhibits the invasion of Tca8113 cells via downregulation of nuclear factor κB-mediated matrix metalloproteinase expression

Abstract

Icotinib is an epidermal growth factor receptor tyrosine kinase inhibitor, which has been revealed to inhibit proliferation in tumor cells. However, the effect of icotinib on cancer cell metastasis remains to be explained. This study examines the effect of icotinib on the migration and invasion of squamous cells of tongue carcinoma (Tca8113 cells) in vitro. The results of the Boyden chamber invasion assay demonstrated that icotinib reduced cell invasion, suppressed the protein levels of matrix metalloproteinases (MMPs), MMP-2 and MMP-9, and increased the expression of tissue inhibitor of metalloproteinase-1. In addition, icotinib was found to significantly decrease the protein levels of nuclear factor κB (NF-κB) p65, which suggested that icotinib inhibits NF-κB activity. Furthermore, treatment with the NF-κB inhibitor, pyrrolidine dithiocarbamate, suppressed cell invasion and MMP-2 expression. These results suggested that icotinib inhibits the invasion of Tca8113 cells by downregulating MMP via the inactivation of the NF-κB signaling pathways.

Keywords: Tca8113 cells; icotinib; matrix metallopeptidase; nuclear factor κB; tumor invasion.

Figure 1

Effects of icotinib on Tca8113 cell viability. MTT assay indicated that treatment with icotinib for 24 h at various concentrations (0 and 2 μM) exhibited no cytotoxicity on Tca8113 cells. Data are presented as the mean ± standard error of the mean (n=5) for each group. ^*P<0.05, vs. the control group.

Figure 2

Effect of icotinib on the invasion of Tca8113 cells. The invaded Tca8113 cells were counted in five random fields following treatment with various concentrations of icotinib (0–2 μM) for 24 h. Data are presented as the mean ± standard error of the mean (n=5) for each group. ^*P<0.05 and ^**P<0.01, vs. the control group.

Figure 3

Effects of icotinib on MMP-2, MMP-9, TIMP-1 and TIMP-2 expression. The expression of these proteins was analyzed in cells following treatment with various concentrations of icotinib (0–2 μM) for 24 h. MMP, matrix metalloproteinase; TIMP, tissue inhibitor of metalloproteinases.

Figure 4

Effects of icotinib on the expression of NF-κB p65. The nuclear extracts were prepared and the protein levels of NF-κB p65 were determined using western blot analysis following treatment with various concentrations of icotinib (0–2 μM) for 24 h. His H1 was used as a loading control and the densitometry was quantified. Data are presented as the mean ± standard error of the mean (n=5) for each group. ^*P<0.05 and ^***P<0.001, vs. the control group. NF-κB, nuclear factor κB.

Figure 5

Effects of the nuclear factor κB inhibitor, PDTC, and icotinib on MMP-2 expression in Tca8113 cells. Cells were pretreated with PDTC (5 μM) for 1 h and then incubated in the presence or absence of icotinib (1 μM) for 24 h. The protein levels of MMP-2 were measured by western blot analysis. β-actin was used as a loading control and the densitometry was quantified. Data are presented as the mean ± standard error of the mean (n=5) for each group. ^*P<0.05 and ^###P<0.001, vs. the control group. PDTC, pyrrolidine dithiocarbamate; MMP, matrix metalloproteinase.