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Objective:That there is a connection between inflammation and cancer has been suspected for a long time. The aim of the present study was to provide new mechanistic insight into the effect of pitavastatin at low dose on NF-κB activated by TNF-α in human breast cancer cell line (MCF 7). Methods:(1)MCF 7 cells (1x104 cells/well) were seeded into 96-well plates and then treated with various concentrations of pitavastatin for 48 h. Cell viability was determinedby WST-8 assay. (2) MCF 7 cells were treated with pitavastatin (1 μM) alone, or pitavastatin (1μM) and TNF-α (1 ng/ml) for 24 h. After the treatment, the nuclear protein was extracted and NF-κB activation was examined by immunoblotting with an antibody specific for NF-κB p65. (3) To further investigate the effect of pitavastatin on NF-κB activation in MCF 7 cells, nuclear extracts were prepared and analyzed by EMSA. (4) MCF 7 cells were cotransfected NF-κB-dependent luciferase reporter (3x κB-luc) using LipofectamineTM with Plus Reagents. After transfection for 24 h, the cells were treated with pitavastatin alone or with pitavastatin, mevalonic acid, Y27632 and TNF-α for 24 h, and then harvested for the luciferase assay. (5) IL-6 concentrations in the supernatants were measured using commercial enzyme linked immunosorbent assay (ELISA) kits. Results:We found that treatment of MCF 7 with 1μM pitavastatin inhibited the proliferation and suppressed the nuclear expression of NF-κB p65 induced by TNF-α with western blot. Furthermore, EMSA showed that pitavastatin significantly reduced the DNA binding activity of NF-κB induced by TNF-α. Subsequently, luciferase assay revealed that pitavastatin (1μM) inhibited the transcriptional activity of the NF-κB promoter, which was clearly related to the HMG-CoA reductase activity because addition of Mevalonic acid (MEV) could elevate the NF-κB activity. Moreover, the Rho kinase inhibitor Y27632 abolished the effect of pitavastatin on NF-κB activity. Finally, the addition of TNF-α significantly increased IL-6 protein production, which was suppressed by the addition of pitavastatin. Conclusion:These results suggest that pitavastatin at low dose (1 μM) inhibits NF-κB activation and decreases IL-6 production induced by TNF-α. It is dependent on Rho kinase pathway in human breast cancer cells.

98th AACR Annual Meeting-- Apr 14-18, 2007; Los Angeles, CA