Silibinin inhibits constitutive and TNFalpha-induced activation of NF-kappaB and sensitizes human prostate carcinoma DU145 cells to TNFalpha-induced apoptosis.
Prostate cancer (PCA) is one of the most common invasive malignancies of men in the US, however, there have been limited successes so far in its therapy. Even most potent agents (e.g. TNFalpha) are ineffective in killing human PCA cells possibly due to constitutive activation of NF-kappaB that subsequently activates a large number of anti-apoptotic genes. In such a scenario, strong apoptotic agent TNFalpha, further induces NF-kappaB activation rather than inducing apoptosis. In several recent studies, we have demonstrated both cancer preventive and anti-cancer efficacy of silymarin and its constituent silibinin in a variety of experimental tumor models and cell culture systems. Here we examined whether silibinin is effective in inhibiting constitutive NF-kappaB activation in human PCA cells, which would help in overcoming TNFalpha-insensitivity. Our studies reveal that silibinin effectively inhibits constitutive activation of NF-kappaB in advanced human prostate carcinoma DU145 cells. Consistent with this, nuclear levels of p65 and p50 sub-units of NF-kappaB were also reduced. In the studies assessing molecular mechanism of this effect, silibinin treatment resulted in a significant increase in the level of IkappaBalpha with a concomitant decrease in phospho-IkappaBalpha. Kinase assays revealed that silibinin dose-dependently decreases IKKalpha kinase activity. The effect of silibinin on IKKalpha seemed to be direct as evidenced by the in vitro kinase assay, where immunoprecipitated IKKalpha was incubated with silibinin. This shows that silibinin does not necessarily need an upstream event to bring about its inhibitory effect on IKKalpha and downstream effectors. Additional studies showed that silibinin also inhibits TNFalpha-induced activation of NF-kappaB via IkappaBalpha pathway and subsequently sensitizes DU145 cells to TNFalpha-induced apoptosis. These results indicate that silibinin could be used to enhance the effectiveness of TNFalpha-based chemotherapy in advanced PCA.
Dhanalakshmi
S
S
Department of Pharmaceutical Sciences, School of Pharmacy, University of Colorado Health Sciences Center, Denver, Colorado, CO 80262, USA.
Singh
R P
RP
Agarwal
C
C
Agarwal
R
R
D021581:Active Transport, Cell Nucleus; D017209:Apoptosis / Q000187:drug effects*; D004268:DNA-Binding Proteins / Q000502:physiology; D006801:Humans; D051550:I-kappa B Kinase; D020672:I-kappa B Proteins; D008297:Male; D000072000:NF-KappaB Inhibitor alpha; D016328:NF-kappa B / Q000378:metabolism*; D010766:Phosphorylation; D011471:Prostatic Neoplasms / Q000188:drug therapy* / Q000473:pathology; D017346:Protein Serine-Threonine Kinases / Q000037:antagonists & inhibitors; D012838:Silymarin / Q000494:pharmacology*; D014407:Tumor Cells, Cultured; D014409:Tumor Necrosis Factor-alpha / Q000494:pharmacology*
D004268:DNA-Binding Proteins; D020672:I-kappa B Proteins; D016328:NF-kappa B; C000606204:NFKBIA protein, human; D012838:Silymarin; D014409:Tumor Necrosis Factor-alpha; D000072000:NF-KappaB Inhibitor alpha; D017346:Protein Serine-Threonine Kinases; C496559:CHUK protein, human; D051550:I-kappa B Kinase; C496560:IKBKB protein, human; C496565:IKBKE protein, human