Short hairpin RNA- mediated gene knockdown of FOXM1 inhibits the proliferation and metastasis of human colon cancer cells through reversal of epithelial-to-mesenchymal transformation

Journal of Experimental & Clinical Cancer Research

XinGuo Zhu²  YiZhou Yao²  HenFeng Chen²  Jing Yu²  You Hu¹  LinHua Jiang²  KanKan Yang² 

[1] Department of Laparoscopic Surgery, The First Affiliated Hospital of Soochow University, Suzhou, 215006, Jiangsu Province, China;Department of General Surgery, The First Affiliated Hospital of Soochow University, Suzhou, 215006, Jiangsu Province, China

关键词: Colorectal cancer;    Metastasis;    EMT;    FOXM1;   

Others  :  1219934
DOI  :  10.1186/s13046-015-0158-1

 received in 2015-01-27, accepted in 2015-04-14,  发布年份 2015

【 摘 要 】

Background

The Forkhead box M1 (FOXM1) is an oncogenic transcription factor and plays a significant role in cell EMT, proliferation, metastasis in a multitude of human solid tumors including colorectal cancer (CRC). However, the underlying molecular mechanisms by which FoxM1 contributes to epithelial-to-mesenchymal (EMT) and metastasis have not been fully elucidated in CRC.

Methods

In our study, we investigated FOXM1 protein expression in 87 CRC tissue specimens, invasive lymph nodes and adjacent paired normal colorectal tissues by immunohistochemical analysis. Then we transfected FOXM1 specific shRNA into SW620 cells to examine effect of FOXM1 on proliferation, colony formation, migration and invasion in vitro. Western blotting and real-time PCR were used to detect the protein and mRNA expression of FOXM1 and EMT-related markers.

Results

FOXM1 was overexpressed in CRC tissues, invasive lymph nodes and CRC cell lines. FoxM1 overexpression was significantly associated with lymph node metastasis (P < 0.001), and tumor recurrence (P < 0.001). Moreover, downregulation of FOXM1 in SW620 cells by shRNA approach inhibited cell growth, clonogenicity, migration and invasion in vitro. In addition, decreased FOXM1 expression in SW620 cells reversed the acquisition of EMT phenotype by up-regulating E-cadherin, as well as reduction Vimentin and Snail expressions at protein and mRNA levels.

Conclusions

FOXM1 may regulate CRC cells metastasis through EMT program and FOXM1 may be a potential target for treatment of CRC.

【 授权许可】

   
2015 Yang et al.; licensee BioMed Central.

【 图 表 】

Figure 1.

Figure 2.

Figure 3.

Figure 4.

Figure 5.

Figure 6.

【 参考文献 】

• [1]Siegel R, DeSantis C, Jemal A: Colorectal cancer statistics, 2014. CA Cancer J Clin 2014, 64:104-17.
• [2]DeSantis CE, Lin CC, Mariotto AB, Siegel RL, Stein KD, Kramer JL, et al.: Cancer treatment and survivorship statistics, 2014. CA Cancer J Clin. 2014, 64:252-71.
• [3]Ferlay J, Shin HR, Bray F, Forman D, Mathers C, Parkin DM: Estimates of worldwide burden of cancer in 2008: GLOBOCAN 2008. Int J Cancer 2010, 127:2893-917.
• [4]Halasi M, Gartel AL: FOX (M1) news—It is cancer. Mol Cancer Ther 2013, 12:245-54.
• [5]Kalin TV, Ustiyan V, Kalinichenko VV: Multiple faces of FoxM1 transcription factor. Cell Cycle 2011, 10:396-405.
• [6]Pilarsky C, Wenzig M, Specht T, Saeger HD, Grützmann R: Identification and validation of commonly overexpressed genes in solid tumors by comparison of microarray data. Neoplasia 2004, 6:744-50.
• [7]Kalinichenko VV, Major ML, Wang X, Petrovic V, Kuechle J, Yoder HM, et al.: Foxm1b transcription factor is essential for development of hepatocellular carcinomas and is negatively regulated by the p19ARF tumor suppressor. Genes Dev 2004, 18:830-50.
• [8]Wang Z, Ahmad A, Banerjee S, Azmi A, Kong D, Li Y, et al.: FoxM1 is a novel target of a natural agent in pancreatic cancer. Pharm Res 2010, 27:1159-68.
• [9]Ahmad A, Wang Z, Kong D, Ali S, Li Y, Banerjee S, et al.: FoxM1 down-regulation leads to inhibition of proliferation, migration and invasion of breast cancer cells through the modulation of extra-cellular matrix degrading factors. Breast Cancer Res Treat 2010, 122:337-46.
• [10]Wen N, Wang Y, Wen L, Zhao S-H, Ai Z-H, Wang Y, et al.: Overexpression of FOXM1 predicts poor prognosis and promotes cancer cell proliferation, migration and invasion in epithelial ovarian cancer. J Transl Med 2014, 12:134. BioMed Central Full Text
• [11]Uddin S, Ahmed M, Hussain A, Abubaker J, Al-Sanea N, AbdulJabbar A, et al.: Genome-wide expression analysis of Middle Eastern colorectal cancer reveals FOXM1 as a novel target for cancer therapy. Am J Pathol 2011, 178:537-47.
• [12]Kim I-M, Ackerson T, Ramakrishna S, Tretiakova M, Wang I-C, Kalin TV, et al.: The Forkhead Box m1 transcription factor stimulates the proliferation of tumor cells during development of lung cancer. Cancer Res 2006, 66:2153-61.
• [13]Wierstra I, Alves J: FOXM1, a typical proliferation-associated transcription factor. Biol Chem 2007, 388:1257-74.
• [14]Leung TW, Lin SS, Tsang AC, Tong CS, Ching JC, Leung WY, et al.: Over-expression of FoxM1 stimulates cyclin B1 expression. FEBS Lett 2001, 507:59-66.
• [15]Kong F-F, Qu Z-Q, Yuan H-H, Wang J-Y, Zhao M, Guo Y-H, et al.: Overexpression of FOXM1 is associated with EMT and is a predictor of poor prognosis in non-small cell lung cancer. Oncol Rep 2014, 31:2660-8.
• [16]Yang C, Chen H, Tan G, Gao W, Cheng L, Jiang X, et al.: FOXM1 promotes the epithelial to mesenchymal transition by stimulating the transcription of Slug in human breast cancer. Cancer Lett 2013, 340:104-12.
• [17]Li Q, Zhang N, Jia Z, Le X, Dai B, Wei D, et al.: Critical role and regulation of transcription factor FoxM1 in human gastric cancer angiogenesis and progression. Cancer Res 2009, 69:3501-9.
• [18]Zhang Y, Zhang N, Dai B, Liu M, Sawaya R, Xie K, et al.: FoxM1B transcriptionally regulates vascular endothelial growth factor expression and promotes the angiogenesis and growth of glioma cells. Cancer Res 2008, 68:8733-42.
• [19]Wang Z, Banerjee S, Kong D, Li Y, Sarkar FH: Down-regulation of Forkhead Box M1 transcription factor leads to the inhibition of invasion and angiogenesis of pancreatic cancer cells. Cancer Res 2007, 67:8293-300.
• [20]Chen H, Zou Y, Yang H, Wang J, Pan H: Downregulation of FoxM1 inhibits proliferation, invasion and angiogenesis of HeLa cells in vitro and in vivo. Int J Oncol 2014, 45:2355-64.
• [21]Bates RC, Mercurio AM: Review the epithelial-mesenchymal transition (EMT) and colorectal cancer progression. Cancer Biol Ther 2005, 4:365-70.
• [22]Li D, Wei P, Peng Z, Huang C, Tang H, Jia Z, et al.: The critical role of dysregulated FOXM1–PLAUR signaling in human colon cancer progression and metastasis. Clin Cancer Res 2013, 19:62-72.
• [23]Huang C, Qiu Z, Wang L, Peng Z, Jia Z, Logsdon CD, et al.: A novel FoxM1-caveolin signaling pathway promotes pancreatic cancer invasion and metastasis. Cancer Res 2012, 72:655-65.
• [24]Bao B, Wang Z, Ali S, Kong D, Banerjee S, Ahmad A, et al.: Over‐expression of FoxM1 leads to epithelial–mesenchymal transition and cancer stem cell phenotype in pancreatic cancer cells. J Cell Biochem 2011, 112:2296-306.
• [25]Miao L, Xiong X, Lin Y, Cheng Y, Lu J, Zhang J, et al.: Down-regulation of FoxM1 leads to the inhibition of the epithelial-mesenchymal transition in gastric cancer cells. Cancer Gene 2014, 207:75-82.
• [26]He S-B, Yuan Y, Wang L, Yu M-J, Zhu Y-B, Zhu X-G: Effects of cyclin-dependent kinase 8 specific siRNA on the proliferation and apoptosis of colon cancer cells. J Exp Clin Cancer Res 2011, 30:109. BioMed Central Full Text
• [27]Livak KJ, Schmittgen TD: Analysis of relative gene expression data using real-time quantitative PCR and the 2(-Delta Delta C(T)) Method. Methods 2001, 25:402-8.
• [28]Chu X-Y, Zhu Z-M, Chen L-B, Wang J-H, Su Q-S, Yang J-R, et al.: FOXM1 expression correlates with tumor invasion and a poor prognosis of colorectal cancer. Acta Histochem 2012, 114:755-62.
• [29]Wang I-C, Chen Y-J, Hughes DE, Ackerson T, Major ML, Kalinichenko VV, et al.: FoxM1 regulates transcription of JNK1 to promote the G1/S transition and tumor cell invasiveness. J Biol Chem 2008, 283:20770-8.
• [30]Yoshida Y, Wang I, Yoder HM, Davidson NO, Costa RH: The forkhead box M1 transcription factor contributes to the development and growth of mouse colorectal cancer. Gastroenterology 2007, 132:1420-31.
• [31]Valastyan S, Weinberg RA: Tumor metastasis: molecular insights and evolving paradigms. Cell 2011, 147:275-92.
• [32]Thiery JP, Sleeman JP: Complex networks orchestrate epithelial–mesenchymal transitions. Nat Rev Mol Cell Biol 2006, 7:131-42.
• [33]Yue D, Li H, Che J, Zhang Y, Tseng H-HK, Jin JQ, et al.: Hedgehog/Gli promotes epithelial-mesenchymal transition in lung squamous cell carcinomas. J Exp Clin Cancer Res 2014, 33:1-7. BioMed Central Full Text
• [34]Loboda A, Nebozhyn MV, Watters JW, Buser CA, Shaw PM, Huang PS, et al.: EMT is the dominant program in human colon cancer. BMC Med Genomics 2011, 4:9. BioMed Central Full Text
• [35]Huang C, Xie D, Cui J, Li Q, Gao Y, Xie K: FOXM1c promotes pancreatic cancer epithelial-to-mesenchymal transition and metastasis via upregulation of expression of the urokinase plasminogen activator system. Clin Cancer Res 2014, 20:1477-88.
• [36]Yang J, Weinberg RA: Epithelial-mesenchymal transition: at the crossroads of development and tumor metastasis. Dev Cell 2008, 14:818-29.
• [37]Batlle E, Sancho E, Francí C, Domínguez D, Monfar M, Baulida J, et al.: The transcription factor snail is a repressor of E-cadherin gene expression in epithelial tumour cells. Nat Cell Biol 2000, 2:84-9.
• [38]Dong C, Wu Y, Yao J, Wang Y, Yu Y, Rychahou PG, et al.: G9a interacts with snail and is critical for snail-mediated E-cadherin repression in human breast cancer. J Clin Invest 2012, 122:1469-86.
• [39]Wu Y, Zhou BP: Snail: more than EMT. Cell Adh Migr 2010, 4:199-203.
• [40]Kaufhold S, Bonavida B: Central role of snail1 in the regulation of EMT and resistance in cancer: a target for therapeutic intervention. J Exp Clin Cancer Res 2014, 33:62. BioMed Central Full Text
• [41]Wang H, Wang H-S, Zhou B-H, Li C-L, Zhang F, Wang X-F, et al.: Epithelial–mesenchymal transition (EMT) induced by TNF-α requires AKT/GSK-3β-mediated stabilization of snail in colorectal cancer. PLoS One 2013, 8:e56664.
• [42]Barrallo-Gimeno A, Nieto MA: The snail genes as inducers of cell movement and survival: implications in development and cancer. Development 2005, 132:3151-61.
• [43]Balli D, Ustiyan V, Zhang Y, Wang I, Masino AJ, Ren X, et al.: Foxm1 transcription factor is required for lung fibrosis and epithelial‐to‐mesenchymal transition. EMBO J 2013, 32:231-44.
• [44]Fan X-J, Wan X-B, Yang Z-L, Fu X-H, Huang Y, Chen D-K, et al.: Snail promotes lymph node metastasis and twist enhances tumor deposit formation through epithelial-mesenchymal transition in colorectal cancer. Hum Pathol 2013, 44:173-80.
• [45]Park HJ, Gusarova G, Wang Z, Carr JR, Li J, Kim KH, et al.: Deregulation of FoxM1b leads to tumour metastasis. EMBO Mol Med 2011, 3:21-34.
• [46]Wei P, Zhang N, Wang Y, Li D, Wang L, Sun X, et al.: FOXM1 promotes lung adenocarcinoma invasion and metastasis by upregulating SNAIL. Int J Biol Sci 2015, 11:186-98.