Abstract
TRAIL is a promising anticancer agent, capable of inducing apoptosis in a wide range of treatment-resistant tumor cells. In ‘type II' cells, the death signal triggered by TRAIL requires amplification via the mitochondrial apoptosis pathway. Consequently, deregulation of the intrinsic apoptosis-signaling pathway, for example, by loss of Bax and Bak, confers TRAIL-resistance and limits its application. Here, we show that despite resistance of Bax/Bak double-deficient cells, TRAIL-treatment resulted in caspase-8 activation and complete processing of the caspase-3 proenzymes. However, active caspase-3 was degraded by the proteasome and not detectable unless the XIAP/proteasome pathway was inhibited. Direct or indirect inhibition of XIAP by RNAi, Mithramycin A or by the SMAC mimetic LBW-242 as well as inhibition of the proteasome by Bortezomib overcomes TRAIL-resistance of Bax/Bak double-deficient tumor cells. Moreover, activation and stabilization of caspase-3 becomes independent of mitochondrial death signaling, demonstrating that inhibition of the XIAP/proteasome pathway overcomes resistance by converting ‘type II' to ‘type I' cells. Our results further demonstrate that the E3 ubiquitin ligase XIAP is a gatekeeper critical for the ‘type II' phenotype. Pharmacological manipulation of XIAP therefore is a promising strategy to sensitize cells for TRAIL and to overcome TRAIL-resistance in case of central defects in the intrinsic apoptosis-signaling pathway.
Highlights
Death ligands initiate receptor oligomerization and formation of the death inducing signaling complex (DISC), resulting in activation of the initiator caspase-8
To evaluate strategies to overcome resistance to tumor necrosis factor-related apoptosis-inducing ligand (TRAIL), we addressed the role of X-linked inhibitor of apoptosis (XIAP) in a HCT116 colon cancer cell line model
We compared the impact of myeloid cell leukemia 1 (Mcl-1) and XIAP downregulation on TRAIL-treated isogenic HCT116 wt cells, HCT116 B-cell lymphoma 2 (Bcl-2)-associated x protein (Bax)-deficient cells and cells devoid of Bax and Bcl-2 homologous antagonist/killer (Bak).[15]
Summary
Death ligands initiate receptor oligomerization and formation of the death inducing signaling complex (DISC), resulting in activation of the initiator caspase-8. Activated, truncated (t)Bid triggers Bax activation to induce mitochondrial membrane permeabilization (MMP) accompanied by the release of apoptogenic factors from the mitochondrial inter-membrane space into the cytosol. One of these factors, cytochrome c, associates with APAF-1 and procaspase-9 to form the ‘apoptosome’, a platform that facilitates autocatalytic activation of caspase-9, which in turn triggers the effector caspases. Cytochrome c, associates with APAF-1 and procaspase-9 to form the ‘apoptosome’, a platform that facilitates autocatalytic activation of caspase-9, which in turn triggers the effector caspases Another pro-apoptotic factor released upon MMP is SMAC/DIABLO.[8,9] SMAC potentiates apoptosis by neutralizing cytosolic inhibitor of apoptosis proteins (IAPs). Inhibition of the endogenous Bak antagonist Mcl-1 enables TRAIL to kill cells via Bak.[14]
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