Suppression of constitutive and tumor necrosis factor alpha-induced nuclear factor (NF)-kappaB activation and induction of apoptosis by apigenin in human prostate carcinoma PC-3 cells: correlation with down-regulation of NF-kappaB-responsive genes.

Development of androgen independence and resistance to apoptosis in prostate cancer are often correlated with high levels of serum tumor necrosis factor (TNF)-alpha in these patients. The loss of sensitivity to TNF-alpha-induced apoptosis in androgen-insensitive prostate carcinoma cells is due in part to constitutive activation of Rel/nuclear factor (NF)-kappaB transcription factors that regulate several cell survival and antiapoptotic genes. Our previous studies have demonstrated growth inhibitory and apoptotic effects of apigenin, a common plant flavonoid, in a variety of human prostate carcinoma cells. Here we examined whether apigenin is effective in inhibiting NF-kappaB expression in androgen-insensitive human prostate carcinoma cells exhibiting high constitutive levels of NF-kappaB.
Using androgen-insensitive human prostate carcinoma PC-3 cells, the effect of apigenin was assessed on NF-kappaB activation by electrophoretic mobility shift assay and reporter gene assay. Expression of NF-kappaB subunits p65 and p50, IkappaBalpha, p-IkappaBalpha, in-beads kinase assay and NF-kappaB-regulated genes were determined by Western blot analysis. Apoptosis was determined by annexin V/propidium iodide staining after fluorescence-activated cell-sorting analysis.
Treatment of cells with 10-40- micro M doses of apigenin inhibited DNA binding and reduced nuclear levels of the p65 and p50 subunits of NF-kappaB. Apigenin inhibited IkappaBalpha degradation and IkappaBalpha phosphorylation and significantly decreased IKKalpha kinase activity. Apigenin also inhibited TNF-alpha-induced activation of NF-kappaB via the IkappaBalpha pathway, thereby sensitizing the cells to TNF-alpha-induced apoptosis. The inhibition of NF-kappaB activation correlated with a decreased expression of NF-kappaB-dependent reporter gene and suppressed expression of NF-kappaB-regulated genes [specifically, Bcl2, cyclin D1, cyclooxygenase-2, matrix metalloproteinase 9, nitric oxide synthase-2 (NOS-2), and vascular endothelial growth factor].
Our results indicate that inhibition of NF-kappaB by apigenin may lead to prostate cancer suppression by transcriptional repression of NF-kappaB-responsive genes as well as selective sensitization of prostate carcinoma cells to TNF-alpha-induced apoptosis.
AuthorsSanjeev Shukla, Sanjay Gupta
JournalClinical cancer research : an official journal of the American Association for Cancer Research (Clin Cancer Res) Vol. 10 Issue 9 Pg. 3169-78 (May 1 2004) ISSN: 1078-0432 [Print] United States
PMID15131058 (Publication Type: Journal Article, Research Support, Non-U.S. Gov't, Research Support, U.S. Gov't, P.H.S.)
Chemical References
  • Flavonoids
  • I-kappa B Proteins
  • Isoenzymes
  • Membrane Proteins
  • NF-kappa B
  • NF-kappa B p50 Subunit
  • Proto-Oncogene Proteins c-bcl-2
  • Recombinant Fusion Proteins
  • Transcription Factor RelA
  • Tumor Necrosis Factor-alpha
  • Vascular Endothelial Growth Factor A
  • Cyclin D1
  • NF-kappaB inhibitor alpha
  • Apigenin
  • Luciferases
  • Nitric Oxide Synthase
  • Nitric Oxide Synthase Type II
  • Cyclooxygenase 2
  • PTGS2 protein, human
  • Prostaglandin-Endoperoxide Synthases
  • IKBKE protein, human
  • Protein-Serine-Threonine Kinases
  • CHUK protein, human
  • I-kappa B Kinase
  • IKBKB protein, human
  • Matrix Metalloproteinase 9
  • Apigenin
  • Apoptosis (drug effects)
  • Biological Transport (drug effects)
  • Cell Line, Tumor
  • Cell Nucleus (drug effects, metabolism)
  • Cyclin D1 (genetics)
  • Cyclooxygenase 2
  • Dose-Response Relationship, Drug
  • Down-Regulation (drug effects)
  • Flavonoids (pharmacology)
  • Gene Expression Regulation, Neoplastic (drug effects)
  • Humans
  • I-kappa B Kinase
  • I-kappa B Proteins (metabolism)
  • Isoenzymes (genetics)
  • Luciferases (genetics, metabolism)
  • Male
  • Matrix Metalloproteinase 9 (genetics)
  • Membrane Proteins
  • NF-kappa B (genetics, metabolism)
  • NF-kappa B p50 Subunit
  • Nitric Oxide Synthase (genetics)
  • Nitric Oxide Synthase Type II
  • Phosphorylation (drug effects)
  • Prostaglandin-Endoperoxide Synthases (genetics)
  • Prostatic Neoplasms (genetics, metabolism, pathology)
  • Protein-Serine-Threonine Kinases (metabolism)
  • Proto-Oncogene Proteins c-bcl-2 (genetics)
  • Recombinant Fusion Proteins (genetics, metabolism)
  • Transcription Factor RelA
  • Transfection
  • Tumor Necrosis Factor-alpha (pharmacology)
  • Vascular Endothelial Growth Factor A (genetics)

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