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2008 Annual Meeting Abstracts
Zinc Chelation Induces Rapid Depletion of the X-linked Inhibitor of Apoptosis Protein (XIAP) and Sensitizes Prostate Cancer Cells to TRIAL-mediated Apoptosis
Benjamin J Scoll, Peter Makhov*, Konstantin Golovine*, Vladimir M Kolenko, Jason Rothman, Paul L Crispen, Tavis Shaw, Robert G Uzzo
Fox Chase Cancer Center, Philadelphia, PA
Introduction: X-linked inhibitor of apoptosis (XIAP), the most potent member of the inhibitor of apoptosis protein (IAP) family of endogenous caspase inhibitors, blocks the initiation and execution phases of the apoptotic cascade. As such, XIAP represents an attractive target in treating apoptosis-resistant forms of cancer. We evaluate the effect of treating a prostate cancer cell line with the membrane-permeable zinc chelator, N,N,N’,N’,-tetrakis(2-pyridylmethyl) ethylenediamine (TPEN).
Methods: Various cancer cell lines, including PC-3 prostate cancer cells, were incubated with 8 µM TPEN. Cell lysates were evaluated with Western blotting. XIAP mRNA levels were detected by RT-PCR. Inhibition was tested with inhibitors of proteasomes (MG-132), caspases (Z-VAD-FMK), and cathepsin (ALLM and CA-074), lysosomal proteases (chloroquine), cysteine proteases (E64d), aspartic proteases (pepstatin A), and serine proteases (Pefabloc). Caspase-3 activity was measured using the flurometric tetrapeptide substrate DEVD-AMC.
Results: Treatment of PC-3 cells with TPEN induced rapid depletion of XIAP. The depletion of XIAP was selective, as TPEN had no effect on the expression of other zinc-binding members of the IAP family including cIAP1, cIAP2 and survivin. The down-regulation of XIAP in TPEN-treated cells occured via proteasome- and caspase-independent mechanisms and was completely prevented by the serine protease inhibitor, Pefabloc. In addition, administration of TPEN sensitized PC-3 cells to caspase-dependent apoptosis in response to treatment with death ligands CD95/Fas, TNF-α or tumor necrosis factor-related apoptosis inducing ligand (TRAIL).
Conclusions: Zinc-chelating agents may be used to sensitize malignant cells to established cytotoxic agents via down-regulation of XIAP.
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