A Smac-mimetic sensitizes prostate cancer cells to TRAIL-induced apoptosis via modulating both IAPs and NF-kappaB

Yao Dai,Liang Xu,Wenhua Tang,Jiqin Lian,Meilan Liu,Theodore S Lawrence,Yongming Li

doi:10.1186/1471-2407-9-392

Abstract

BackgroundAlthough tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) is a promising agent for human cancer therapy, prostate cancer still remains resistant to TRAIL. Both X-linked inhibitor of apoptosis (XIAP) and nuclear factor-kappaB function as key negative regulators of TRAIL signaling. In this study, we evaluated the effect of SH122, a small molecule mimetic of the second mitochondria-derived activator of caspases (Smac), on TRAIL-induced apoptosis in prostate cancer cells.MethodsThe potential of Smac-mimetics to bind XIAP or cIAP-1 was examined by pull-down assay. Cytotoxicity of TRAIL and/or Smac-mimetics was determined by a standard cell growth assay. Silencing of XIAP or cIAP-1 was achieved by transient transfection of short hairpin RNA. Apoptosis was detected by Annexin V-PI staining followed by flow cytometry and by Western Blot analysis of caspases, PARP and Bid. NF-kappaB activation was determined by subcellular fractionation, real time RT-PCR and reporter assay.ResultsSH122, but not its inactive analog, binds to XIAP and cIAP-1. SH122 significantly sensitized prostate cancer cells to TRAIL-mediated cell death. Moreover, SH122 enhanced TRAIL-induced apoptosis via both the death receptor and the mitochondrial pathway. Knockdown of both XIAP and cIAP-1 sensitized cellular response to TRAIL. XIAP-knockdown attenuated sensitivity of SH122 to TRAIL-induced cytotoxicity, confirming that XIAP is an important target for IAP-inhibitor-mediated TRAIL sensitization. SH122 also suppressed TRAIL-induced NF-kappaB activation by preventing cytosolic IkappaB-alpha degradation and RelA nuclear translocation, as well as by suppressing NF-kappaB target gene expression.ConclusionThese results demonstrate that SH122 sensitizes human prostate cancer cells to TRAIL-induced apoptosis by mimicking Smac and blocking both IAPs and NF-kappaB. Modulating IAPs may represent a promising approach to overcoming TRAIL-resistance in human prostate cancer with constitutively active NF-kappaB signaling.

Highlights

Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) is a promising agent for human cancer therapy, prostate cancer still remains resistant to tumor necrosis factor-related apoptosis-inducing ligand (TRAIL)
ShFmuigmaucar-nmepi1mroesttiactceocmanpcoeurncdeilnlsteracted with X-linked inhibitor of apoptosis (XIAP) and cIAP-1 in second mitochondria-derived activator of caspases (Smac)-mimetic compound interacted with XIAP and cIAP-1 in human prostate cancer cells
TRAIL alone (50 ng/ml) moderately induced apoptosis, and SH122 alone showed a minor effect on apoptosis, with a slightly increased apoptotic cell population compared with the control even at the highest concentration (Figure 3). Both death receptor and mitochondrial pathways are involved in SH122-sensitized, TRAIL-induced apoptosis Based on TRAIL-induced apoptotic signaling, we examined the expression of caspases treated with SH122 and TRAIL

Summary

Introduction

Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) is a promising agent for human cancer therapy, prostate cancer still remains resistant to TRAIL. Both X-linked inhibitor of apoptosis (XIAP) and nuclear factor-kappaB function as key negative regulators of TRAIL signaling. The mitochondrial pathway is engaged by the release of multiple pro-apoptotic factors from mitochondria into the cytosol, such as cytochrome c, Smac and apoptosis inducing factor (AIF). These factors execute cells through apoptosis in either a caspasedependent or independent manner [9]

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