Plumbagin (5-Hydroxy-2-methyl-1,4-naphthoquinone) Suppresses NF-κB Activation and NF-κB-regulated Gene Products Through Modulation of p65 and IκBα Kinase Activation, Leading to Potentiation of Apoptosis Induced by Cytokine and Chemotherapeutic Agents

Gautam Sethi,Santosh K. Sandur,Haruyo Ichikawa,Bharat B. Aggarwal,Kwang Seok Ahn

doi:10.1074/jbc.m601595200

Gautam Sethi, Santosh K. Sandur + Show 3 more

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https://doi.org/10.1074/jbc.m601595200

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Abstract

Plumbagin, derived from the medicinal plant Plumbago zeylanica, modulates cellular proliferation, carcinogenesis, and radioresistance, all known to be regulated by the activation of the transcription factor NF-kappaB, suggesting plumbagin might affect the NF-kappaB activation pathway. We found that plumbagin inhibited NF-kappaB activation induced by TNF, and other carcinogens and inflammatory stimuli (e.g. phorbol 12-myristate 13-acetate, H2O2, cigarette smoke condensate, interleukin-1beta, lipopolysaccharide, and okadaic acid). Plumbagin also suppressed the constitutive NF-kappaB activation in certain tumor cells. The suppression of NF-kappaB activation correlated with sequential inhibition of the tumor necrosis factor (TNF)-induced activation of IkappaBalpha kinase, IkappaBalpha phosphorylation, IkappaBalpha degradation, p65 phosphorylation, p65 nuclear translocation, and the NF-kappaB-dependent reporter gene expression activated by TNF, TNFR1, TRAF2, NIK, IKK-beta, and the p65 subunit of NF-kappaB. Plumbagin also suppressed the direct binding of nuclear p65 and recombinant p65 to the DNA, and this binding was reversed by dithiothreitol both in vitro and in vivo. However, plumbagin did not inhibit p65 binding to DNA when cells were transfected with the p65 plasmid containing cysteine 38 mutated to serine. Plumbagin down-regulated the expression of NF-kappaB-regulated anti-apoptotic (IAP1, IAP2, Bcl-2, Bcl-xL, cFLIP, Bfl-1/A1, and survivin), proliferative (cyclin D1 and COX-2), and angiogenic (matrix metalloproteinase-9 and vascular endothelial growth factor) gene products. This led to potentiation of apoptosis induced by TNF and paclitaxel and inhibited cell invasion. Overall, our results indicate that plumbagin is a potent inhibitor of the NF-kappaB activation pathway that leads to suppression of NF-kappaB-regulated gene products. This may explain its cell growth modulatory, anticarcinogenic, and radiosensitizing effects previously described.

Highlights

Plumbagin has been shown to exert anticancer and antiproliferative activities in animal models as well as in cells in culture [7,8,9,10,11]
We used tumor necrosis factor (TNF) to examine the effect of plumbagin on the NF-␬B activation pathway because the pathway activated by this agent is well understood
Plumbagin Down-modulated TNF-induced NF-␬B-dependent Antiapoptotic Gene Expression—Because NF-␬B regulates the expression of the anti-apoptotic proteins such as inhibitor of apoptosis protein 1 (IAP1)/2 [45, 46], Bcl-2 [47], Bcl-xL [48], cFLIP [49], Bfl-1/A1 [50], and survivin [51], we examined whether plumbagin can modulate the expression of these anti-apoptotic gene products induced by TNF in KBM-5 cells

Summary

Introduction

Plumbagin has been shown to exert anticancer and antiproliferative activities in animal models as well as in cells in culture [7,8,9,10,11]. Because plumbagin has been reported to exhibit chemopreventive, growth inhibitory, and radiosensitizing effects, we postulated that this quinone may be mediating its effects through modulation of the NF-␬B activation pathway. To test this hypothesis, we investigated the effect of plumbagin on NF-␬B activation induced by several inflammatory agents in various cell types, including those in which NF-␬B is constitutively active. We found that plumbagin inhibited NF-␬B activation and NF-␬B-regulated gene products, leading to an increase in apoptosis and suppression of cellular invasion

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