Kurarinone promotes TRAIL-induced apoptosis by inhibiting NF-κB-dependent cFLIP expression in HeLa cells

Ok-Won Seo,Kyu-Sun Lee,Moo-Ho Won,Kwang-Soon Lee,Young-Myeong Kim,Kwon-Soo Ha,Ji-Hee Kim,Young-Guen Kwon,Jung Hwan Kim

doi:10.3858/emm.2012.44.11.074

Abstract

This study was designed to investigate the effects of the prenylated flavonoid kurarinone on TNF-related apoptosis inducing ligand (TRAIL)-induced apoptosis and its underlying mechanism. A low dose of kurarinone had no significant effect on apoptosis, but this compound markedly promoted tumor cell death through elevation of Bid cleavage, cytochrome c release and caspase activation in HeLa cells treated with TRAIL. Caspase inhibitors inhibited kurarinone-mediated cell death, which indicates that the cytotoxic effect of this compound is mediated by caspase-dependent apoptosis. The cytotoxic effect of kurarinone was not associated with expression levels of Bcl-2 and IAP family proteins, such as Bcl-2, Bcl-xL, Bid, Bad, Bax, XIAP, cIAP-1 and cIAP-2. In addition, this compound did not regulate the death-inducing receptors DR4 and DR5. On the other hand, kurarinone significantly inhibited TRAIL-induced IKK activation, IκB degradation and nuclear translocation of NF-κB, as well as effectively suppressed cellular FLICE-inhibitory protein long form (cFLIPL) expression. The synergistic effects of kurarinone on TRAIL-induced apoptosis were mimicked when kurarinone was replaced by the NF-κB inhibitor withaferin A or following siRNA-mediated knockdown of cFLIPL. Moreover, cFLIP overexpression effectively antagonized kurarinone-mediated TRAIL sensitization. These data suggest that kurarinone sensitizes TRAIL-induced tumor cell apoptosis via suppression of NF-κB-dependent cFLIP expression, indicating that this compound can be used as an anti-tumor agent in combination with TRAIL.

Highlights

Soluble recombinant tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) is thought to be therapeutic alternative against tumors (Plantivaux et al, 2009)
Microscopic analysis showed that TNF-related apoptosis inducing ligand (TRAIL) (75 ng/ml) alone induced cell death of about 15%, and TRAIL co-treated with 5 μM kurarinone further elevated cell death to 70%, while kurarinone alone did not demonstrate a significant cytotoxic effect (Figures 1A and 1B)
To differentiate between apoptosis and necrosis, we examined whether caspase inhibitors would regulate kurarinone-mediated synergistic cell death

Summary

Introduction

Soluble recombinant tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) is thought to be therapeutic alternative against tumors (Plantivaux et al, 2009). TRAIL can induce tumor cell apoptosis with little apoptotic effect to normal cells (Cho et al, 2005). Despite this attractive selective tumoricidal activity of TRAIL, many tumor cells show resistance to TRAIL-induced apoptosis and the mechanism of TRAIL resistance is not fully elucidated (Plantivaux et al, 2009). Vol 44(11), 653-664, 2012 suppressing survival signals and gene expression as well as elevating apoptosis-associated gene expression (Neuzil et al, 2004; Seol et al, 2005; Griffith et al, 2011; Gupta et al, 2011; Jung et al, 2012)

Methods

Results

Conclusion