Anti-oncogenic activity of Chibby in the development of human nasopharyngeal carcinoma

Abstract

The Wnt/β-catenin pathway serves important roles in cancer development. The expression and function of Chibby (Cby), as a direct antagonist of β-catenin, in nasopharyngeal carcinoma (NPC) has not been fully investigated. The present study revealed that the mRNA and protein expression of Cby was significantly lower in NPC tissue than in the adjacent normal tissue. Low expression of Cby was significantly associated with the tumor and the clinical staging. Furthermore, Cby overexpression inhibited the proliferation of human NPC SUNE1 cells and induced cell cycle arrest. In addition, Cby overexpression also significantly enhanced the susceptibility of SUNE1 cells to apoptosis. These results indicated that Cby might serve as an anti-oncogenic gene in the development of NPC and could represent a potential therapeutic target for the human NPC therapy.

Keywords: Chibby; Wnt/β-catenin; apoptosis; cell cycle; nasopharyngeal carcinoma.

Figure 1.

Expression of Cby in the clinical NPC tissues. (A) Scatter graph depicting the relative level of Cby mRNA of NPC tumor tissue (0.0725±0.0061) and adjacent normal tissue (0.0951±0.0509; Student's t-test, *P=0.0219, n=45). (B) The expression of Cby protein in NPC tissue and adjacent normal tissue. Cby expression is clearly observed in the nuclei of adjacent normal tissue cells, but expression is significant lower in NPC cells (original magnification, ×200). Cby, Chibby; NPC, nasopharyngeal cancer; T, NPC tissue; P, adjacent normal tissue.

Figure 2.

Establish of Cby overexpression SUNE1 cells model. (A and B) Expression of Cby in NP69 and SUNE1 cells detected by western blot analysisfollowing transfection withplv-cs2.0 (empty vector) and plv-cs2.0-Cby. (C) Activation of Wnt signaling assessed by β-catenin/T-cell factor-responsive luciferase reporter activity, normalized to β-Gal. Wnt3a, 30 nM. *P<0.05; **P<0.01; ***P<0.001. Cby, Chibby.

Figure 3.

Effect of Cby overexpression on the proliferation of SUNE1 cells. (A) The growth of NP69 and SUNE1 cells transfected with plv-cs2.0 (empty vector) or plv-cs2.0-Cby were detected by MTT assay every 24 h from day 0 to day 5. (B) The anti-oncogenic potential of Cby, assessed by colony formation assays. The number of foci containing >50 cells was counted. **P<0.01; ***P<0.001. Cby, Chibby.

Figure 4.

Effect of Cby overexpression on the cell cycle distributions and apoptosis of SUNE1 cells. (A) Cell cycle analysis was performed by flow cytometry following plasmid transfection. The proportion of G₁ phase cells for plv-cs2.0 (empty vector) and plv-cs2.0-Cby groups were 52.5±2.5 and 73.6±3.2%, respectively (P=0.0051). (B) Apoptosis analysis of the control group plv-cs2.0 and plv-cs2.0-Cby SUNE1 cells. Hoechst 33342 staining showed apoptotic cells with condensed chromatin or fragmented nuclei. Magnification, ×200. (C) Assessment of apoptosis of SUNE1 cells using flow cytometry with Annexin V-fluorescein isothiocyanate/phosphoinositide staining following treatment with 20 µM 5-fluorouracil (left) or 30 nM Wnt 3a (right). *P<0.05, **P<0.01. Cby, Chibby.