Putative Tumor-Suppressor Function Of Krüppel-Like Factor 4 In Primary Lung Carcinoma

Krüppel-like factor 4 (KLF4) is a zinc-finger protein highly expressed in epithelial tissues such as the gut and skin, and plays important roles in stem cells and in development of gastric cancers. However, the role of KLF4 in primary lung cancer is still unknown.Purpose: To explore the function of KLF4 gene in lung cancer development.Methods: First, collect lung cancer tissues and matched normal lung tissues, and test KLF4 expression in lung tissues by Western blotting, RT-PCR and Real-time PCR methods; Second, test the level of KLF4 promoter methylation in lung cancer tissues and matched normal tissues by methylation-specific PCR and bisulphate cloning-squening methods; To observe the effect of Ad-KLF4 on cell viability and cell cycle in various of lung cancer cell lines by SRB and PI staing methods; To establish stable KLF4 expression lung cancer cell line, KLF4 knockdown HBE and HBE/K-ras cell lines, then observe the changement of cell growth, and clonegenic formation; By subcutenous animaltumor model, observe the effect of KLF4 gene on tumor growt; In addition, after Ad-KLF4 treatment, to investigate the changement of KLF4-asssociated gene p21 and cyclin D1, and the changement of telomerase activity for learning the mechanism of KLF4-induced effect in lung cancer development.Results: The levels of KLF4 protein and mRNA were dramatically decreased in most of primary lung tumors when compared to that of matched normal lung tissues. Moreover, level of hypermethylation in KLF4 promoter was increased in primary lung cancer tissues when compared with that of matched normal lung tissues. Enforced expression of KLF4 in resulted in marked inhibition of cell growth and clonogenic formation in cultured lung cancer cell lines with lower KLF4-expression. Furthermore, knockdown of KLF4 expression promoted cell growth, and clonogenic formation in immortalized humean bronchial epithelial cells. Delivery of KLF4 gene to lung cancer lcells by ex vivo transfecation or by adenovector-mediated gene transfer suppressed tumor growth in vivo. The tumor-suppressive mechanism of KLF4 was associated with its role in the cell cycle arrest at the G₁-S checkpoint and suppression of telomerase activity.Conclusion: From lung tissues, lung cancer cell lines and animal model, our results suggest that KLF4 plays an important suppressive role in the development and progression of primary lung carcinoma.