The therapeutic potential of XIAP suppression in cancer treatment has encouraged research to identify and characterize mechanisms that regulate XIAP expression as well as antagonists that block XIAP function. The inhibition or down-regulation of XIAP in cancer cells lowers the apoptotic threshold, thereby inducing cell death and/or enhancing the cytotoxic action of chemotherapeutic agents. Indeed, XIAP antagonist 1396-34 sensitized PC-3 and DU-145 prostate cancer cells to chemotherapeutic agents and TRAIL 22
. Moreover, XIAP antisense enhanced pro-apoptotic TRAIL potency by 12- to 13-fold in DU-145 prostate cancer cells 23
demonstrating that knock-down of XIAP itself was sufficient to reverse TRAIL resistance.
Zinc, in addition to its catalytic role in more than 300 zinc metalloenzymes, is a known inhibitor of enzymes in general. Removal of zinc from an inhibitory zinc-specific enzymatic site results in a marked increase of enzyme activity 24
. For example, human tissue kallikreins, a subgroup of extracellular serine proteases including prostate specific antigen (PSA), are inactivated by reversible binding of zinc (reviewed in 25
). Such findings provide a rationale for our observation that the addition of zinc chelators triggers serine protease dependent depletion of XIAP. Nevertheless, the exact mechanism underlying the depletion of XIAP under zinc-deficient conditions remains to be elucidated. A possibility also exists that the TPEN-mediated pro-apoptotic effect is not exclusively due to XIAP depletion. Further studies are needed to fully explore the potential mechanisms of action of zinc chelating agents.
Zinc is not the only metal ion influencing XIAP function. Elevated copper levels result in a conformational change in XIAP, which accelerates its degradation and significantly decreases its ability to inhibit caspase-3 in HEK 293 cells 26
. In contrast to our observation that zinc chelation induces specific degradation of XIAP (), studies by Mufti et al. show that intracellular copper accumulation also affects other IAPs such as Op-IAP and cIAP2. Mufti et al. also report that copper induced no significant changes in XIAP mRNA expression 26
. This finding is in agreement with our data, in that XIAP mRNA levels were not decreased in TPEN-treated cells () indicating that both copper and TPEN can modulate XIAP expression at the posttranscriptional level. Furthermore, it has been suggested that direct binding of copper to cysteine residues within XIAP induces a conformational change that is associated with decreased stability 26
. An intriguing possibility exists that in addition to direct binding of copper to XIAP, it can also substitute for zinc and thus, promote instability of XIAP. Indeed, the ability of various metals, including copper and cadmium, to substitute zinc in zinc finger domains is well established 27,28
Recent studies suggest that the presence or development of resistance may ultimately limit the use of small-molecule XIAP inhibitors 3
. It is unknown whether malignant cells acquire mutations in the BIR domains of XIAP and whether these mutations affect XIAP’s function or its response to inhibitors. It is also unknown whether mutations in XIAP can develop after treatment with small-molecule XIAP inhibitors similar to the development of mutations in ABL kinase after treatment with the inhibitor Gleevec 3,29
. Therefore, agents able to completely deplete XIAP in malignant cells might be superior to small-molecule XIAP inhibitors. Taken together, our findings indicate that zinc-chelating molecules, capable of reducing XIAP expression, in combination with other treatment modalities may be beneficial in treating apoptosis resistant tumors.