Quercetin sensitizes human hepatoma cells to TRAIL‐induced apoptosis via Sp1‐mediated DR5 up‐regulation and proteasome‐mediated c‐FLIP_S down‐regulation

Abstract

This study demonstrates that combined treatment with subtoxic doses of quercetin (3 0 ,3 0 ,4 0 ,5,7-pentahydroxyflavone), a flavonoid found in many fruits and vegetables, plus tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) induces rapid apoptosis in TRAIL-resistant hepatocellular carcinoma (HCC) cells. Effective induction of apoptosis by the combined treatment with quercetin and TRAIL was not blocked by overexpression of Bcl-xL, which is known to confer resistance to various chemotherapeutic agents. These results suggest that this combined treatment may provide an attractive strategy for treating resistant HCCs. While the proteolytic processing of procaspase-3 by TRAIL was partially blocked in various HCC cells treated with TRAIL alone, co-treatment with quercetin efficiently recovered TRAIL-induced caspase activation. We found that quercetin treatment of HCC cells significantly up-regulated the mRNA and protein levels of DR5, a death receptor of TRAIL, in a transcription factor Sp1-dependent manner. Furthermore, treatment with quercetin significantly decreased the protein levels of c-FLIP, an inhibitor of caspase-8, through proteasome-mediated degradation. Finally, administration of small interfering RNA against DR5 or overexpression of c-FLIP S , but not c-FLIP L , significantly attenuated quercetin-stimulated TRAIL-induced apoptosis. Collectively, these findings show that quercetin recovers TRAIL sensitivity in various HCC cells via up-regulation of DR5 and down-regulation of c-FLIP S . J. Cell. Biochem. 105: 1386-1398. ß 2008 KEY WORDS: QUERCETIN; TRAIL; HEPATOMA; DR5; c-FLIP S H epatocellular carcinoma (HCC) is the fifth most frequent neoplasm worldwide (>500,000 death/year) [Bruix et al., 2004]. Most hepatocellular carcinoma patients are diagnosed at advanced stages that are unsuitable for the current curative therapies of resection and transplantation [Avila et al., 2006]. The current chemotherapeutic options yield poor responses and low patient survival, making them ineffective for treatment of advanced HCC [Avila et al., 2006]. Therefore, new therapeutic alternatives are needed for more effective treatment of this malignancy.Tumor necrosis factor (TNF)-related apoptosis-inducing ligand (TRAIL), a member of the TNF family, is considered a promising anticancer agent due to its ability to induce apoptosis in a variety of tumor cell types while having only negligible effects on normal cells [Ashkenazi et al., 1999]. Binding of TRAIL to either DR4 or DR5, two death receptors of TRAIL, leads to oligomerization and clustering of their intracellular death domains. The subsequent interaction of DR4 or DR5 with the adaptor molecule, FADD (Fas-associated death domain), via their respective death domains leads to recruitment and activation of caspase-8 [ Thomas et al., 2004]. Finally, caspase-8 activates the executioner caspases (e.g., caspase-3 and caspase-7), leading to apoptotic cell death [Srivastava, 2001]. Although TRAIL has garnered considerable attention as a novel anticancer agent...

Quercetin-mediated Mcl-1 and survivin downregulation restores TRAIL-induced apoptosis in non-Hodgkin's lymphoma B cells

“…Quercetin has been reported to synergize with TRAIL, [24][25][26][27][28][29] but the molecular mechanisms underlying this sensitization remain largely unknown. At the proximal level, quercetin-mediated sensitization to TRAIL has been correlated with TRAIL-R2 stabilization, 24 increased TRAIL-R2 expression at the cell surface, 25,29 enhanced TRAIL DISC formation 27 and even c-FLIP downregulation. 25 In our cellular models, regulation of proximal events is unlikely to explain the synergy since quercetin induced no change in TRAIL receptor or c-FLIP expression and only modest differences in TRAIL-R1, FADD, caspase-8 and caspase-10 recruitment within the DISC.…”

“…At the proximal level, quercetin-mediated sensitization to TRAIL has been correlated with TRAIL-R2 stabilization, 24 increased TRAIL-R2 expression at the cell surface, 25,29 enhanced TRAIL DISC formation 27 and even c-FLIP downregulation. 25 In our cellular models, regulation of proximal events is unlikely to explain the synergy since quercetin induced no change in TRAIL receptor or c-FLIP expression and only modest differences in TRAIL-R1, FADD, caspase-8 and caspase-10 recruitment within the DISC. As compared to conventional chemotherapeutic drugs, such as cisplatin or 5FU, which induce a robust increase in caspase-8 recruitment and activation within the TRAIL DISC in VAL cells, 17 quercetin only weakly enhanced initiator caspase-8/10 or TRAIL-R1 recruitment.…”

“…At this point, it should be emphasized that the concentrations of quercetin used in our study, 20 mM, are lower than those used in most studies (50-200 mM). [24][25][26] Besides, cell specificities may also give rise to discrepant results. Likewise, the mitochondrial pathway is strongly inhibited in resistant B-lymphoma cell lines, yet quercetin achieves restoration of the TRAIL apoptotic machinery.…”

Sensitization of rhTRAIL-resistant Triple-negative Breast Carcinoma Through Silibinin Co-Treatment

“…rhTRAIL is a more favorable therapeutic agent because many cancers have a non-functional TP53 gene that leads to necrosis over ANTICANCER RESEARCH 37: 6593-6599 (2017) 6596 apoptosis, so if chemotherapy and radiation are applied, then negative side effects could be observed in patients. Despite this, there are factors that are implicated in resistance to rhTRAIL that lead to clinical trials being terminated due to the absence of clinical efficacy (25)(26)(27)(28). Our study sought to combat rhTRAIL resistance in TNBC BT-20 and HCC1937 cells through the application of the "mother-nature"-derived silibinin as a sensitizing agent.…”

“…Some rhTRAIL-resistant cancer cell lines have displayed low expression levels of DR4 and DR5, and some studies have postulated that this lack of expression of DRs hinders rhTRAIL from inducing apoptosis in those cancer cell lines (25,28). Accordingly, this study inspected the protein and cell surface expressions of DR4 and DR5 in silibinin-treated TNBC cells via western blot and FACS analyses.…”