Evodiamine Abolishes Constitutive and Inducible NF-κB Activation by Inhibiting IκBα Kinase Activation, Thereby Suppressing NF-κB-regulated Antiapoptotic and Metastatic Gene Expression, Up-regulating Apoptosis, and Inhibiting Invasion

Yasunari Takada,Yoshinori Kobayashi,Bharat B Aggarwal

doi:10.1074/jbc.m500077200

Abstract

Evodiamine, an alkaloidal component extracted from the fruit of Evodiae fructus (Evodia rutaecarpa Benth., Rutaceae), exhibits antiproliferative, antimetastatic, and apoptotic activities through a poorly defined mechanism. Because several genes that regulate cellular proliferation, carcinogenesis, metastasis, and survival are regulated by nuclear factor-kappaB (NF-kappaB), we postulated that evodiamine mediates its activity by modulating NF-kappaB activation. In the present study, we investigated the effect of evodiamine on NF-kappaB and NF-kappaB-regulated gene expression activated by various carcinogens. We demonstrate that evodiamine was a highly potent inhibitor of NF-kappaB activation, and it abrogated both inducible and constitutive NF-kappaB activation. The inhibition corresponded with the sequential suppression of IkappaBalpha kinase activity, IkappaBalpha phosphorylation, IkappaBalpha degradation, p65 phosphorylation, p65 nuclear translocation, and p65 acetylation. Evodiamine also inhibited tumor necrosis factor (TNF)-induced Akt activation and its association with IKK. Suppression of Akt activation was specific, because it had no effect on JNK or p38 MAPK activation. Evodiamine also inhibited the NF-kappaB-dependent reporter gene expression activated by TNF, TNFR1, TRADD, TRAF2, NIK, and IKK but not that activated by the p65 subunit of NF-kappaB. NF-kappaB-regulated gene products such as Cyclin D1, c-Myc, COX-2, MMP-9, ICAM-1, MDR1, Survivin, XIAP, IAP1, IAP2, FLIP, Bcl-2, Bcl-xL, and Bfl-1/A1 were all down-regulated by evodiamine. This down-regulation potentiated the apoptosis induced by cytokines and chemotherapeutic agents and suppressed TNF-induced invasive activity. Overall, our results indicated that evodiamine inhibits both constitutive and induced NF-kappaB activation and NF-kappaB-regulated gene expression and that this inhibition may provide a molecular basis for the ability of evodiamine to suppress proliferation, induce apoptosis, and inhibit metastasis.

Highlights

Evodiamine, an alkaloidal component extracted from the fruit of Evodiae fructus (Evodia rutaecarpa Benth., Rutaceae), exhibits antiproliferative, antimetastatic, and apoptotic activities through a poorly defined mechanism
Evodiamine Inhibits nuclear factor-␬B (NF-␬B) Activation Induced by Various Carcinogens and Inflammatory Stimuli—tumor necrosis factor (TNF), IL-1␤, phorbol 12-myristate 13-acetate (PMA), okadaic acid, H2O2, and cigarette smoke condensate are all potent activators of NF-␬B, but they differ in how they activate NF-␬B [30, 33, 37, 38]
The concentrations of evodiamine and NF-␬B activators used and the duration of exposure had minimal effect on cell viability. These results suggested that evodiamine acts at a step in the NF-␬B activation pathway that is common to all six agents

Summary

Introduction

Evodiamine, an alkaloidal component extracted from the fruit of Evodiae fructus (Evodia rutaecarpa Benth., Rutaceae), exhibits antiproliferative, antimetastatic, and apoptotic activities through a poorly defined mechanism. NF-␬B-regulated gene products such as Cyclin D1, c-Myc, COX-2, MMP-9, ICAM-1, MDR1, Survivin, XIAP, IAP1, IAP2, FLIP, Bcl-2, Bcl-xL, and Bfl-1/A1 were all down-regulated by evodiamine This down-regulation potentiated the apoptosis induced by cytokines and chemotherapeutic agents and suppressed TNF-induced invasive activity. How evodiamine regulates cell proliferation, apoptosis, invasion, and migration is incompletely understood We postulated that this compound mediates its effects by modulating nuclear factor ␬B (NF-␬B), a transcription factor that plays a major role in tumorigenesis [13]. NF-␬B regulates several genes that mediate proliferation (e.g. Cyclin D1 and c-Myc), antiapoptosis (e.g. Survivin, tumor necrosis factor (TNF) receptor-associated factor 1 (TRAF1), cellular Fas-associated death domain protein-like interleukin-1␤ (IL-1␤)-converting enzyme-inhibitory protein (FLIP), inhibitor-of-apoptosis protein (IAP), X chromosome-linked IAP (XIAP), Bcl-2, and Bcl-xL), drug resistance (e.g. multidrug resistance protein 1 (MDR1)), immunomodulation (e.g. chemokines and interleukins), and metastasis (e.g. cyclooxygenase-2 (COX-2), matrix metalloproteinase-9 (MMP9), and intracellular adhesion molecule-1 (ICAM-1)) (14 –28). Upon activation of NF-␬B, I␬B␣ kinase (IKK) is activated, leading to I␬B␣ phosphorylation, ubiquitination, and degradation; release of p50/p65 the heterodimer, translocates to the nucleus, binds to its consensus sequence, and induces gene transcription

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