Synergistic Effect of Sulindac and Simvastatin on Apoptosis in Lung Cancer A549 Cells through AKT-Dependent Downregulation of Survivin / Journal of the Korean Cancer Association, 대한암학회지

PURPOSE: Non-steroidal anti-inflammatory drugs (NSAIDs) and statins are potential chemopreventive or chemotherapeutic agents. The mechanism underlying the deregulation of survivin by NSAIDs and statins in human non-small cell lung cancer cells has not been elucidated. In this study, we investigated the synergistic interaction of sulindac and simvastatin in lung cancer A549 cells. MATERIALS AND METHODS: Cell viability was measured by an MTT assay, while the expression of apoptotic markers, AKT, and survivin in response to sulindac and simvastatin was examined by Western blotting. DNA fragmentation by apoptosis was analyzed by flow cytometry in A549 cells. Reactive oxygen species (ROS) generation was measured by flow cytometry using H2DCFDA and MitoSOX Red, and the effects of pretreatment with N-acetylcysteine were tested. The effects of AKT on survivin expression in sulindac- and simvastatin-treated cells were assessed. Survivin was knocked down or overexpressed to determine its role in apoptosis induced by sulindac and simvastatin. RESULTS: Sulindac and simvastatin synergistically augmented apoptotic activity and intracellular ROS production in A549 cells. Inhibition of AKT by siRNA or LY294002 inhibited survivin, while AKT overexpression markedly increased survivin expression, even in the presence of sulindac and simvastatin. Moreover, survivin siRNA enhanced sulindac- and simvastatininduced apoptosis. In contrast, survivin upregulation protected against sulindac- and simvastatin-induced apoptosis. CONCLUSION: Combined treatment with sulindac and simvastatin augmented their apoptotic potential in lung cancer cells through AKT signaling-dependent downregulation of survivin. These results indicate that sulindac and simvastatin may be clinically promising therapies for the prevention of lung cancer.

Medulative effect of dihydroartemisinin on UCHL1 gene expression in human prostate cancer PC-3 cells and its mechanism / 肿瘤

Objective: To investigate the effect of dihydroartemisinin (DHA) on expression of tumor suppressor gene ubiquitin carboxyl-terminal hydrolase L1 (UCHL1) in human prostate cancer cell line PC-3, and explore its regulative mechanism. Methods: PC-3 cells were treated with different concentrations (25, 50, and 100 μmol/L) of DHA for 48 h, while PC-3 cells without DHA treatment were used as the control. Then the apoptosis and cell cycle distribution were detected by flow cytometry. The expressions and cellular locations of DNA methyltransferase 1 (DNMT1) and UCHL1 proteins were detected by immunofluorescence staining. The expression levels of UCHL1, DNMT1, phospho-Akt (p-Akt) and Akt proteins were detected by Western blotting. The 1 μmol/L phosphoinositide-3-kinase (PI3K) inhibitor Wortmanin was used as a positive control to analyze the expression of p-Akt protein in DHA-treated group. Results: DHA significantly induced the apoptosis of PC-3 cells and arrested the cell cycle distribution at phase G2/M as compared with those of the control group (both P 0.05) in the DHA-treated group and the positive control group. The downregulation of p-Akt expression was more obvious in high-concentration of DHA-treated group, much closer to that in the positive control group. Conclusion: DHA can inhibit the expression of DNMT1, restore the function of UCHL1 gene, induce the apoptosis of PC-3 cells, and block the cell cycle progression. These mechanisms may be related to the suppressive activity of PI3K/Akt pathway.

Effect of PTEN/PI3K mutations on gene expression profiling of lung cancer cells / 肿瘤

Objective: To investigate the effect of PTEN (phosphatase and tensin homolog deleted on chromosome ten)/ PI 3K (phosphoinositide 3-kinase) mutation on gene expression profiling of lung cancer cell lines. Methods: PTEN protein expression levels were analyzed by Western blotting, and the PTEN gene mutation was confirmed by sequencing in 19 NSCLC (non-small cell lung cancer) cell lines. The effect of PTEN gene mutation on the downstream AKT and mTOR pathways was also examined by Western blotting. GSEA (Gene set enrichment analysis) was employed to investigate the difference of gene expression profiling induced by PTEN/PI 3K mutation in 19 NSCLC cell lines. Results: In this study, 19 NSCLC cell lines had been screened, and 5 cell lines had no PTEN protein expression and harbored PTEN mutation. Other 2 cell lines had PIK 3CA mutation, which was found by database retrieval. The mutation of PTEN gene had a significant impact on the continuous activation of its downstream molecules AKT and mTOR pathways. GSEA was used to compare the gene expression profiling characters between the CON (without PTEN or PIK 3CA mutation) and the MUT (with PTEN or PIK 3CA mutation) groups of NSCLC cell lines. It was surprising that the most enriched gene set affected by PTEN/PI3K dysfunction was the mitochondrial-metabolism gene set. Conclusion: Mutation of PTEN/PI 3K may affect its downstream molecules AKT and mTOR pathways as well as the lung cancer cells' gene expression profiling, especially for the mitochondrial-metabolism gene set. This finding implies the importance of mitochondrial-metabolism regulation in lung cancer development. © 2012 by Tumor.