Experimental Study Of Its Biological Role And Mechanism Of KLF9 In Prostate Cancer

[Objective]:This study was performed to explore the potential biological role and mechanisms of transcription factor KLF9 in the formation and progress of prostate cancer.[Methods]:The quantitative real-time polymerase chain reaction (qRT-PCR) and Western blot were used to detect the levels of KLF9 in LNCaP cells treated by flutamide and the expression of KLF9 in prostate cell lines. PC3 and DU145 cells were transfected with pTRIPZ-KLF9 vector to measure the effect of KLF9 on cell proliferation and apoptosis; cell proliferation was tested by MTT assay, the changes of cell cycle and apoptosis were determined by flow cytometry using propidium iodide (PI) staining and Annexin V and 7-AAD staining respectively. Western blot was also used to detect the changes of apoptosis-related proteins and proteins involved in the pathways. The pLVX-AKT vector was transfected into the PC3/KLF9 and DU145/KLF9 cells to evaluate the effect of AKT overexpression. Tumor xenograft model was established with PC3/KLF9 and LNCaP/KLF9 cells and used to determine the effect of KLF9 overexpression in vivo.[Results]:We found that KLF9 was induced in a time-dependent manner in response to apoptosis caused by flutamide and knockdown of KLF9 could decrease flutamide-induced growth inhibition in LNCaP cells. Moreover, the levels of KLF9 are lower in prostate cancer cell lines, particularly in androgen-independent cell lines (DU145、PC3 and C4-2B) compared with 2 non-tumor cell lines (RWPE-1 and BPH1).Overexpression of KLF9 dramatically suppressed cell growth, colony formation, causing cell cycle arrest in G2/M phase and induced cell apoptosis in PC3 and DU145 cells (p< 0.01); Consistent with the flow cytometry analysis results, overexpression of KLF9 significantly increased the expression of apoptosis-related proteins, cleaved PARP and cleaved caspase3 and decreased the expression of anti-apoptotic proteins, Bcl-2 and Bcl-xl. In xenografts model, KLF9 overexpression also efficiently suppressed the tumorigenicity, tumor growth and induced tumor cell apoptosis in prostate cancer in vivo. Mechanistically, the activation of AKT and the downstream targets were suppressed by the overexpressing KLF9; Furthermore, AKT reactivation could partially rescue the KLF9-overexpressing-mediated growth inhibition and apoptosis in prostate cancer cells.[Conclusions]:1) The expression of KLF9 is lower in prostate cancer cell lines compared with non-tumor cell lines; 2) KLF9 is induced in flutamide-caused apoptosis in LNCaP cells; 3) KLF9 overexpression significantly suppresses cell growth and enhances apoptosis in vitro and in vivo, may act as a tumor suppressor gene in prostate cancer; 4) The decreased strength of AKT activation is involved in KLF9-mediated inhibition of prostate cancer cells.