Cooperative Cytotoxicity of Proteasome Inhibitors and Tumor Necrosis Factor–Related Apoptosis-Inducing Ligand in Chemoresistant Bcl-2-Overexpressing Cells

Abstract

Purpose: Bcl-2 overexpression is frequently detected in lymphoid malignancies, being associated with poor prognosis and reduced response to therapy. Here, we evaluated whether Bcl-2 overexpression affects the cytotoxic activity of proteasome inhibitors taken alone or in association with conventional anticancer drugs or tumor necrosis factor–related apoptosis-inducing ligand (TRAIL). Experimental Design: Jurkat cells engineered to overexpress Bcl-2 were treated with proteasome inhibitors (MG132, epoxomicin, and bortezomib), anticancer drugs (etoposide and doxorubicin), TRAIL, or combinations of these compounds. Cell death and loss of mitochondrial transmembrane potential were detected by flow cytometry. Cytosolic relocalization of cytochrome c and SMAC/Diablo, caspase cleavage, and Bcl-2 and Mcl-1 levels were determined by immunoblotting. Nuclear factor-κB inhibition was done by retroviral transduction with a dominant-negative mutant of IκBα. Results: Bcl-2 overexpression results in significant inhibition of apoptosis in response to proteasome inhibitors, antiblastics, and TRAIL. Addition of TRAIL to proteasome inhibitors results in a synergistic cytotoxic effect in Bcl-2-overexpressing cells, whereas this result is not reproduced by the combination of proteasome inhibitors with antiblastic drugs. Importantly, proteasome inhibitors plus TRAIL induce mitochondrial dysfunction irrespective of up-regulated Bcl-2. Bcl-2 cleavage to a fragment with putative proapoptotic activity and elimination of antiapoptotic Mcl-1 may both play a role in proteasome inhibitors-TRAIL cooperation. Conversely, nuclear factor-κB inhibition by proteasome inhibitors is per se insufficient to explain the observed synergy. Conclusions: Combined proteasome inhibitors and TRAIL overcome the apoptotic threshold raised by Bcl-2 and may prove useful in the treatment of chemoresistant malignancies with up-regulated Bcl-2.

Green tea polyphenol EGCG sensitizes human prostate carcinoma LNCaP cells to TRAIL-mediated apoptosis and synergistically inhibits biomarkers associated with angiogenesis and metastasis

“…There exists a synergism in the apoptotic process presumably by downregulation of prosurvival signaling molecule AKT in addition to modulation of other important anti/proapoptotic molecules. It has been shown that proteasome inhibitors cooperate with TRAIL in inducing apoptosis in cancers with over expression of bcl2 (Nencioni et al, 2005). Our results demonstrate that EGCG that has also been shown to act as a proteasome inhibitor (Lam et al, 2004;Landis-Piwowar et al, 2005) might actually synergize with TRAIL in inducing apoptosis in LNCaP cells.…”

NF-κB-independent sensitization of glioblastoma cells for TRAIL-induced apoptosis by proteasome inhibition

“…Although some potential mediators of this sensitization effect have been indicated, for example p21 (Lashinger et al, 2005), c-FLIP (Sayers et al, 2003), Bik and Bim (Nikrad et al, 2005), whereas others have been excluded, for instance Bcl-2 (Nencioni et al, 2005), Bax (He et al, 2004) and Bcl-xL (Johnson et al, 2003), the specific contribution of NF-kB to this phenomenon remained unclear. In fact, there is only one other report pointing to a differential role of proteasome versus NF-kB inhibition in the regulation of TRAIL-induced apoptosis in glioblastoma cells, possibly by blocking degradation of active caspases (Kim et al, 2004).…”

Salinomycin in cancer: A new mission for an old agent

“…It was shown that salinomycin is capable of inducing apoptosis in both wild-type p53 cells and p53-lacking cells, indicating that the induction of apoptosis by salinomycin is independent of the p53 status of the cell (34). Next, p53-lacking lymphoblastic leukemia cells were transfected with a plasmid encoding the human anti-apoptotic protein Bcl-2, leading to stable overexpression of Bcl-2 and resistance of the cells to apoptosis induced by chemotherapeutic drugs (37). In these cells, salinomycin was able to markedly induce apoptosis even at the low concentrations used for the induction of apoptosis in non-resistant cells (34).…”