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Anticancer activity of a cyclooxygenase inhibitor, CX9051, in human prostate cancer cells: the roles of NF-κB and crosstalk between the extrinsic and intrinsic apoptotic pathways

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Abstract

Comprehensive studies support the notion that selective inhibitors of cyclooxygenase-2 (COX-2) display anticancer activities in numerous types of cancer cells, including prostate cancers. Our previous study showed that the benzodithiazolium-based compound CX9051 selectively inhibited COX-2 activity. We now show that CX9051 inhibits cell proliferation and induces apoptosis in numerous human cancer cell types. Biochemical analyses, including flow cytometry, showed that CX9051 induced apoptosis in the absence of cell cycle checkpoint arrest and down-regulated the expression of Bcl-2, Bcl-xL, and Mcl-1, but up-regulated tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) expression, leading to proteolytic activation of caspase-8, -9, -7, and -3. These data suggest that CX9051 functions in both mitochondria-mediated intrinsic and death receptor-induced extrinsic apoptosis pathways. Moreover, confocal microscopy demonstrated that CX9051 induced nuclear translocation of nuclear factor-kappa B (NF-κB) at initial stage and then caused a marked decrease of total cellular NF-κB at later stage in both PC-3 and DU145 cells. Taken together, our data suggest that CX9051 induces TRAIL up-regulation and activation of extrinsic apoptotic signaling, which in turn activates mitochondria-mediated intrinsic apoptotic signaling, leading to cancer cell apoptosis.

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Acknowledgments

This study was supported by research grants NSC94-2320-B002-097, NSC95-2320-B002-035, and NSC96-2320-B002-017 from the National Science Council, Taiwan.

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Authors and Affiliations

  1. School of Pharmacy, National Taiwan University, No. 1, Section 1, Jen-Ai Road, Taipei, 100, Taiwan

    Chiung-Hua Huang, Jih-Hwa Guh, Grace Shiahuy Chen, Pin-Hsuan Lu & Ji-Wang Chern

  2. Department of Applied Chemistry, Providence University, 200 Chung-Chi Road, Shalu, Taichung, Taiwan

    Grace Shiahuy Chen

  3. Department of Life Science, College of Life Science, National Taiwan University, Taipei, Taiwan

    Ji-Wang Chern

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  1. Chiung-Hua Huang
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  2. Jih-Hwa Guh
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Correspondence to Ji-Wang Chern.

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Fig. S1

Effect of CX9051 on lipopolysaccharide-induced COX-2 expression and prostaglandin E2 production in THP-1 macrophages. a COX-2 protein expression. The level of COX-2 protein was monitored at the indicated times after treatment of cells with lipopolysaccharide (100 ng/mL), with or without CX9051. Cell lysates were then prepared and subjected to Western blotting using an antibody specific for human COX-2. Equal loading of proteins was verified by GAPDH. b Prostaglandin E2 production was measured in THP-1 macrophages cultured with lipopolysaccharide (100 ng/mL) with or without various concentrates of CX9051 for 24 h. Values represent the mean ± SD of duplicate determinations from two separate experiments (DOC 51 kb)

Fig. S2

The cell cycle distribution of the respective control. Flow cytometric analysis of PC-3 and DU145 prostate cancer cells in the absence of CX9051 for the indicated time interval was performed (DOC 100 kb)

Fig. S3

Effect of CX9051 on Bad protein expression in PC-3 cells mitochondria fractions. Western blot analysis of mitochondria fraction from PC-3 cells treated with 10 μM CX9051 for the indicated times was performed using specific antibody against Bad and complex IV was used as a mitochondrial loading control (DOC 63 kb)

Fig. S4

Effect of CX9051 on the nuclear translocation of NF-κB p65 in DU145 cells. Immunocytometry analysis of NF-κB p65 localization in DU145 cells was performed after treatment with 10 μM CX9051 for the indicated times. FITC staining of NF-κB p65 is indicated by green fluorescence, and PI staining of DNA is indicated by red fluorescence (DOC 15947 kb)

Table S1

Effect of CX9051 and other known COX-2 inhibitors on recombinant COX-2 enzyme activity (DOC 37 kb)

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Huang, CH., Guh, JH., Chen, G.S. et al. Anticancer activity of a cyclooxygenase inhibitor, CX9051, in human prostate cancer cells: the roles of NF-κB and crosstalk between the extrinsic and intrinsic apoptotic pathways. Naunyn-Schmied Arch Pharmacol 382, 159–169 (2010). https://doi.org/10.1007/s00210-010-0528-3

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  • DOI: https://doi.org/10.1007/s00210-010-0528-3

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