Deficient tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) death receptor transport to the cell surface in human colon cancer cells selected for resistance to TRAIL-induced apoptosis

Many tumor cell types are sensitive to tumor necrosis factor-related apoptosis-inducing ligand (TRAIL)-induced apoptosis. Incubation of TRAIL-sensitive cells with TRAIL invariably leads to resistant survivors even when high doses of TRAIL are used. Because the emergence of resistance to apoptosis is a major concern in successful treatment of cancer, and TRAIL survivors may contribute to therapeutic failure, we investigated potential resistance mechanisms. We selected TRAIL-resistant SW480 human colon adenocarcinoma cells by repeatedly treating them with high and/or low doses of TRAIL. The resulting TRAIL-resistant clones were not cross-resistant to Fas or paclitaxel. Expression of modulators of apoptosis was not changed in the resistant cells, including TRAIL receptors, cFLIP, Bax, Bid, or IAP proteins. Surprisingly, we found that DISC formation was deficient in multiple selected TRAIL-resistant clones. DR4 was not recruited to the DISC upon TRAIL treatment, and caspase-8 was not activated at the DISC. Although total cellular DR4 mRNA and protein were virtually identical in TRAIL-sensitive parental and TRAIL-resistant clones, DR4 protein expression on the cell surface was essentially undetectable in the TRAIL-resistant clones. Moreover, exogenous DR4 and KILLER/DR5 were not properly transported to the cell surface in the TRAIL-resistant cells. Interestingly, TRAIL-resistant cells were resensitized to TRAIL by tunicamycin pretreatment, which increased cell surface expression of DR4 and KILLER/DR5. Our data suggest that tumor cells may become resistant to TRAIL through regulation of the death receptor cell surface transport and that resistance to TRAIL may be overcome by the glycosylation inhibitor/endoplasmic reticulum stress-inducing agent tunicamycin.     

E1A sensitizes cancer cells to TRAIL-induced apoptosis through enhancement of caspase activation

Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) has been shown to induce apoptosis of cancer cells. Sensitization of cancer cells to TRAIL, particularly TRAIL-resistant cancer cells, could improve the effectiveness of TRAIL as an anticancer agent. The adenovirus type 5 E1A that associates with anticancer activities including sensitization to apoptosis induced by tumor necrosis factor is currently being tested in clinical trials. In this study, we investigated the sensitivity to TRAIL in the E1A transfectants ip1-E1A2 and 231-E1A cells and the parental TRAIL-resistant human ovarian cancer SKOV3.ip1 and TRAIL-sensitive human breast cancer MDA-MB-231 cells. The results indicated that the percentage of TRAIL-induced apoptotic cells was significantly higher in the E1A transfectants of both cell lines than it was in the parental cell lines. To further investigate the cellular mechanism of this effect, we found that E1A enhances TRAIL-induced activation of caspase-8, caspase-9, and caspase-3. Inhibition of caspase-3 activity by a specific inhibitor, Z-DEVD-fmk, abolished TRAIL-induced apoptosis. In addition, E1A enhanced TRAIL expression in ip1-E1A2 cells, but not in 231-E1A cells, and the anti-TRAIL neutralizing antibody N2B2 blocked the E1A-mediated bystander effect in vitro. Taken together, these results suggest that E1A sensitizes both TRAIL-sensitive and TRAIL-resistant cancer cells to TRAIL-induced apoptosis, which occurs through the enhancement of caspase activation; activation of caspase-3 is required for TRAIL-induced apoptosis; and E1A-induced TRAIL expression is involved in the E1A-mediated bystander effect. Combination of E1A and TRAIL could be an effective treatment for cancer.     

The flavonolignan silibinin potentiates TRAIL-induced apoptosis in human colon adenocarcinoma and in derived TRAIL-resistant metastatic cells

Silibinin, a flavonolignan, is the major active component of the milk thistle plant (Silybum marianum) and has been shown to possess anti-neoplastic properties. TNF-related apoptosis-inducing ligand (TRAIL) is a promising anti-cancer agent which selectively induces apoptosis in cancer cells. However, resistance to TRAIL-induced apoptosis is an important and frequent problem in cancer treatment. In this study, we investigated the effect of silibinin and TRAIL in an in vitro model of human colon cancer progression, consisting of primary colon tumor cells (SW480) and their derived TRAIL-resistant metastatic cells (SW620). We showed by flow cytometry that silibinin and TRAIL synergistically induced cell death in the two cell lines. Up-regulation of death receptor 4 (DR4) and DR5 by silibinin was shown by RT-PCR and by flow cytometry. Human recombinant DR5/Fc chimera protein that has a dominant-negative effect by competing with the endogenous receptors abrogated cell death induced by silibinin and TRAIL, demonstrating the activation of the death receptor pathway. Synergistic activation of caspase-3, -8, and -9 by silibinin and TRAIL was shown by colorimetric assays. When caspase inhibitors were used, cell death was blocked. Furthermore, silibinin and TRAIL potentiated activation of the mitochondrial apoptotic pathway and down-regulated the anti-apoptotic proteins Mcl-1 and XIAP. The involvement of XIAP in sensitization of the two cell lines to TRAIL was demonstrated using the XIAP inhibitor embelin. These findings demonstrate the synergistic action of silibinin and TRAIL, suggesting chemopreventive and therapeutic potential which should be further explored.     

Ionizing radiation can overcome resistance to TRAIL in TRAIL-resistant cancer cells

Although the majority of cancer cells are killed by TRAIL (tumor necrosis factor-related apoptosis-inducing ligand treatment), certain types show resistance to it. Ionizing radiation also induces cell death in cancer cells and may share common intracellular pathways with TRAIL leading to apoptosis. In the present study, we examined whether ionizing radiation could overcome TRAIL resistance in the variant Jurkat clones. We first selected TRAIL-resistant or -sensitive Jurkat clones and examined cross-responsiveness of the clones between TRAIL and radiation. Treatment with gamma-radiation induced significant apoptosis in all the clones, indicating that there seemed to be no cross-resistance between TRAIL and radiation. Combined treatment of radiation with TRAIL synergistically enhanced killing of TRAIL-resistant cells, compared to TRAIL or radiation alone. Apoptosis induced by combined treatment of TRAIL and radiation in TRAIL-resistant cells was associated with cleavage of caspase-8 and the proapoptotic Bid protein, resulting in the activation of caspase-9 and caspase-3. No changes in the expressions of TRAIL receptors (DR4 and DR5) and Bcl-2 or Bax were found after treatment. The caspase inhibitor z-VAD-fmk completely counteracted the synergistic cell killing induced by combined treatment of TRAIL and gamma-radiation. These results demonstrated that ionizing radiation in combination with TRAIL could overcome resistance to TRAIL in TRAIL-resistant cells through TRAIL receptor-independent synergistic activation of the cascades of the caspase-8 pathway, suggesting a potential clinical application of combination treatment of TRAIL and ionizing radiation to TRAIL-resistant cancer cells.     

Cellular inhibitor of apoptosis 1 (cIAP-1) degradation by caspase 8 during TNF-related apoptosis-inducing ligand (TRAIL)-induced apoptosis

TNF-related apoptosis-inducing ligand (TRAIL) is a potential chemotherapeutic agent with high selectivity for malignant cells. Many tumors, however, are resistant to TRAIL cytotoxicity. Although cellular inhibitors of apoptosis 1 and 2 (cIAP-1 and -2) are often over-expressed in cancers, their role in mediating TRAIL resistance remains unclear. Here, we demonstrate that TRAIL-induced apoptosis of liver cancer cells is associated with degradation of cIAP-1 and X-linked IAP (XIAP), whereas cIAP-2 remains unchanged. Lower concentrations of TRAIL causing minimal or no apoptosis do not alter cIAP-1 or XIAP protein levels. Silencing of cIAP-1 expression, but not XIAP or cIAP-2, as well as co-treatment with a second mitochondrial activator of caspases (SMAC) mimetic (which results in rapid depletion of cIAP-1), sensitizes the cells to TRAIL. TRAIL-induced loss of cIAP-1 and XIAP requires caspase activity. In particular, caspase 8 knockdown stabilizes both cIAP-1 and XIAP, while caspase 9 knockdown prevents XIAP, but not cIAP-1 degradation. Cell-free experiments confirmed cIAP-1 is a substrate for caspase 8, with likely multiple cleavage sites. These results suggest that TRAIL-mediated apoptosis proceeds through caspase 8-dependent degradation of cIAP-1. Targeted depletion of cIAP-1 by SMAC mimetics in conjunction with TRAIL may be beneficial for the treatment of human hepatobiliary malignancies.     

Thiourea compound AW00178 sensitizes human H1299 lung carcinoma cells to TRAIL-mediated apoptosis

The tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) triggers apoptosis in a wide variety of cancer cells. Recently, cancer cell resistance to TRAIL-mediated apoptosis has become a challenging issue in the development of TRAIL-based anti-cancer therapies. In this study, we found that 1-(5-chloro-2-methyl-phenyl)-3-[4-(5-trifluoromethyl-pyrazol-1-yl)-phenyl]-thiourea (AW00178) was able to sensitize TRAIL-resistant human lung cancer H1299 cells to TRAIL-mediated apoptosis. Treatment with AW00178, either alone or in combination with TRAIL, induced the expression of CHOP, a protein related to TRAIL sensitivity, and reduced the expression of survivin, an anti-apoptotic protein involved in TRAIL resistance. Additionally, AW00178, alone or in combination with TRAIL, induced the activation of c-Jun and inactivation of Akt. A pharmacologic inhibition study revealed that c-Jun activation and Akt inactivation were strongly related to CHOP induction and survivin down-regulation, respectively. In summary, these results suggested that AW00178 mediated sensitization to TRAIL-mediated apoptosis in H1299 cells by increasing sensitivity and decreasing resistance to TRAIL via the induction of c-Jun-dependent CHOP expression and the reduction of Akt-dependent survivin expression, respectively.     

Enhanced caspase-8 recruitment to and activation at the DISC is critical for sensitisation of human hepatocellular carcinoma cells to TRAIL-induced apoptosis by chemotherapeutic drugs

Tumour necrosis factor (TNF)-related apoptosis-inducing ligand (TRAIL) exhibits potent antitumour activity upon systemic administration in mice without showing the deleterious side effects observed with other apoptosis-inducing members of the TNF family such as TNF and CD95L. TRAIL may, thus, have great potential in the treatment of human cancer. However, about 60% of tumour cell lines are not sensitive to TRAIL. To evaluate the mechanisms of tumour resistance to TRAIL, we investigated hepatocellular carcinoma (HCC) cell lines that exhibit differential sensitivity to TRAIL. Pretreatment with chemotherapeutic drugs, for example, 5-fluorouracil (5-FU), rendered the TRAIL-resistant HCC cell lines sensitive to TRAIL-induced apoptosis. Analysis of the TRAIL death-inducing signalling complex (DISC) revealed upregulation of TRAIL-R2. Caspase-8 recruitment to and its activation at the DISC were substantially increased after 5-FU sensitisation, while FADD recruitment remained essentially unchanged. 5-FU pretreatment downregulated cellular FLICE-inhibitory protein (cFLIP) and specific cFLIP downregulation by small interfering RNA was sufficient to sensitise TRAIL-resistant HCC cell lines for TRAIL-induced apoptosis. Thus, a potential mechanism for TRAIL sensitisation by 5-FU is the increased effectiveness of caspase-8 recruitment to and activation at the DISC facilitated by the downregulation of cFLIP and the consequent shift in the ratio of caspase-8 to cFLIP at the DISC.     

Curcumin enhances the apoptosis-inducing potential of TRAIL in prostate cancer cells: molecular mechanisms of apoptosis, migration and angiogenesis

Background

We have recently shown that curcumin (a diferuloylmethane) inhibits growth and induces apoptosis, and also demonstrated that TRAIL induces apoptosis by binding to specific cell surface death receptors in prostate cancer cells. The objectives of this paper were to investigate the molecular mechanisms by which curcumin enhanced the apoptosis-inducing potential of TRAIL in prostate cancer cells.

Results

Curcumin enhanced the apoptosis-inducing potential of TRAIL in androgen-unresponsive PC-3 cells and sensitized androgen-responsive TRAIL-resistant LNCaP cells. Curcumin inhibited the expressions of Bcl-2, Bcl-X_L, survivin and XIAP, and induced the expressions Bax, Bak, PUMA, Bim, and Noxa and death receptors (TRAIL-R1/DR4 and TRAIL-R2/DR5) in both cell lines. Overexpression of dominant negative FADD inhibited the interactive effects of curcumin and TRAIL on apoptosis. Treatment of these cells with curcumin resulted in activation of caspase-3, and caspase-9, and drop in mitochondrial membrane potential, and these events were further enhanced when combined with TRAIL. Curcumin inhibited capillary tube formation and migration of HUVEC cells and these effects were further enhanced in the presence of MEK1/2 inhibitor PD98059.

Conclusion

The ability of curcumin to inhibit capillary tube formation and cell migration, and enhance the therapeutic potential of TRAIL suggests that curcumin alone or in combination with TRAIL can be used for prostate cancer prevention and/or therapy.     

Suppression of NF-kappa B survival signaling by nitrosylcobalamin sensitizes neoplasms to the anti-tumor effects of Apo2L/TRAIL

We have previously demonstrated the anti-tumor activity of nitrosylcobalamin (NO-Cbl), an analog of vitamin B12 that delivers nitric oxide (NO) and increases the expression of tumor necrosis factor-related apoptosis-inducing ligand (Apo2L/TRAIL) and its receptors in human tumors. The specific aim of this study was to examine whether NO-Cbl could sensitize drug-resistant melanomas to Apo2L/TRAIL. Antiproliferative effects of NO-Cbl and Apo2L/TRAIL were assessed in malignant melanomas and non-tumorigenic melanocyte and fibroblast cell lines. Athymic nude mice bearing human melanoma A375 xenografts were treated with NO-Cbl and Apo2L/TRAIL. Apoptosis was measured by TUNEL and confirmed by examining levels and activity of key mediators of apoptosis. The activation status of NF-kappa B was established by assaying DNA binding, luciferase reporter activity, the phosphorylation status of I kappa B alpha, and in vitro IKK activity. NO-Cbl sensitized Apo2L/TRAIL-resistant melanoma cell lines to growth inhibition by Apo2L/TRAIL but had minimal effect on normal cell lines. NO-Cbl and Apo2L/TRAIL exerted synergistic anti-tumor activity against A375 xenografts. Treatment with NO-Cbl followed by Apo2L/TRAIL induced apoptosis in Apo2L/TRAIL-resistant tumor cells, characterized by cleavage of caspase-3, caspase-8, and PARP. NO-Cbl inhibited IKK activation, characterized by decreased phosphorylation of I kappa B alpha and inhibition of NF-kappa B DNA binding activity. NO-Cbl suppressed Apo2L/TRAIL- and TNF-alpha-mediated activation of a transfected NF-kappa B-driven luciferase reporter. XIAP, an inhibitor of apoptosis, was inactivated by NO-Cbl. NO-Cbl treatment rendered Apo2L/TRAIL-resistant malignancies sensitive to the anti-tumor effects of Apo2L/TRAIL in vitro and in vivo. The use of NO-Cbl and Apo2L/TRAIL capitalizes on the tumor-specific properties of both agents and represents a promising anti-cancer combination.     

IAP antagonists target cIAP1 to induce TNFalpha-dependent apoptosis

XIAP prevents apoptosis by binding to and inhibiting caspases, and this inhibition can be relieved by IAP antagonists, such as Smac/DIABLO. IAP antagonist compounds (IACs) have therefore been designed to inhibit XIAP to kill tumor cells. Because XIAP inhibits postmitochondrial caspases, caspase 8 inhibitors should not block killing by IACs. Instead, we show that apoptosis caused by an IAC is blocked by the caspase 8 inhibitor crmA and that IAP antagonists activate NF-kappaB signaling via inhibtion of cIAP1. In sensitive tumor lines, IAP antagonist induced NF-kappaB-stimulated production of TNFalpha that killed cells in an autocrine fashion. Inhibition of NF-kappaB reduced TNFalpha production, and blocking NF-kappaB activation or TNFalpha allowed tumor cells to survive IAC-induced apoptosis. Cells treated with an IAC, or those in which cIAP1 was deleted, became sensitive to apoptosis induced by exogenous TNFalpha, suggesting novel uses of these compounds in treating cancer.     

Hypoxia inducing factor-1α regulates tumor necrosis factor-related apoptosis-inducing ligand sensitivity in tumor cells exposed to hypoxia

Hypoxia is a common environmental stress. Particularly, the center of rapidly-growing solid tumors is easily exposed to hypoxic conditions. Hypoxia is well known to attenuate the therapeutic response to radio and chemotherapies including tumor necrosis factor (TNF)-related apoptosis-inducing ligand (TRAIL) protein. HIF-1α is a critical mediator of the hypoxic response. However, little is known about the function of hypoxia-inducible factor-1α (HIF-1α) on hypoxic inhibition of TRAIL-mediated apoptosis. In this study, we investigated whether hypoxic inhibition of TRAIL-mediated apoptosis can be regulated by modulating HIF-1α protein. Hypoxia- and DEF-induced HIF-1α activation inhibited the TRAIL-mediated apoptosis in SK-N-SH, HeLa, A549 and SNU-638 cells. And also, HIF-1α inactivating reagents including DOX increased the sensitivity to TRAIL protein in tumor cells exposed to hypoxia. Furthermore, knock-down of HIF-1α using lentiviral RNA interference sensitized tumor cells to TRAIL-mediated cell death under hypoxic condition. Taken together, these results indicate that HIF-1α inactivation increased TRAIL sensitivity in hypoxia-induced TRAIL-resistant tumor cells and also suggest that HIF-1α inhibitors may have benefits in combination therapy with TRAIL against hypoxic tumor cells.     

Proteasome-mediated degradation of Smac during apoptosis: XIAP promotes Smac ubiquitination in vitro

During apoptosis, Smac (second mitochondria-derived activator of caspases)/DIABLO, an IAP (inhibitor of apoptosis protein)-binding protein, is released from mitochondria and potentiates apoptosis by relieving IAP inhibition of caspases. We demonstrate that exposure of MCF-7 cells to the death-inducing ligand, TRAIL (tumor necrosis factor-related apoptosis-inducing ligand), results in rapid Smac release from mitochondria, which occurs before or in parallel with loss of cytochrome c. Smac release is inhibited by Bcl-2/Bcl-xL or by a pan-caspase inhibitor demonstrating that this event is caspase-dependent and modulated by Bcl-2 family members. Following release, Smac is rapidly degraded by the proteasome, an effect suppressed by co-treatment with a proteasome inhibitor. As the RING finger domain of XIAP possesses ubiquitin-protein ligase activity and XIAP binds tightly to mature Smac, an in vitro ubiquitination assay was performed which revealed that XIAP functions as a ubiquitin-protein ligase (E3) in the ubiquitination of Smac. Both the association of XIAP with Smac and the RING finger domain of XIAP are essential for ubiquitination, suggesting that the ubiquitin-protein ligase activity of XIAP may promote the rapid degradation of mitochondrial-released Smac. Thus, in addition to its well characterized role in inhibiting caspase activity, XIAP may also protect cells from inadvertent mitochondrial damage by targeting pro-apoptotic molecules for proteasomal degradation.     

Codium fragile F2 sensitize colorectal cancer cells to TRAIL-induced apoptosis via c-FLIP ubiquitination

This study demonstrates that combined treatment with subtoxic doses of Codium extracts (CE), a flavonoid found in many fruits and vegetables, and tumor necrosis factor-related apoptosis-inducing ligand (TRAIL), induces apoptosis in TRAIL-resistant colorectal cancer (CRC) cells. Effective induction of apoptosis by combined treatment with CE and TRAIL was not blocked by Bcl-xL overexpression, which is known to confer resistance to various chemotherapeutic agents. While TRAIL-mediated proteolytic processing of procaspase-3 was partially blocked in various CRC cells treated with TRAIL alone, co-treatment with CE efficiently recovered TRAIL-induced caspase activation. We observed that CE treatment of CRC cells did not change the expression of anti-apoptotic proteins and pro-apoptotic proteins, including death receptors (DR4 and DR5). However, CE treatment markedly reduced the protein level of the short form of the cellular FLICE-inhibitory protein (c-FLIPS), an inhibitor of caspase-8, via proteasome-mediated degradation. Collectively, these observations show that CE recovers TRAIL sensitivity in various CRC cells via down-regulation of c-FLIPS.     

15-Deoxy-Δ<Superscript>12,14</Superscript>-prostaglandin J<Subscript>2</Subscript> (15d-PGJ<Subscript>2</Subscript>) sensitizes human leukemic HL-60 cells to tumor necrosis factor-related apoptosis-inducing ligand (TRAIL)-induced apoptosis through Akt downregulation

While tumor necrosis factor (TNF)-related apoptosis-inducing ligand (TRAIL) is a promising new agent for the treatment of cancer, resistance to TRAIL remains a therapeutic challenge. Identifying agents to use in combination with TRAIL to enhance apoptosis in leukemia cells would increase the potential utility of this agent as a therapy for leukemia. Here, we show that 15-deoxy-Δ^12,14-prostaglandin J₂ (15d-PGJ₂), a natural ligand for peroxisome proliferator-activated receptor γ (PPARγ), can sensitize TRAIL-resistant leukemic HL-60 cells to TRAIL-induced apoptosis. The sensitization to TRAIL-induced apoptosis by 15d-PGJ₂ was not blocked by a PPARγ inhibitor (GW9662), suggesting a PPARγ-independent mechanism. This process was accompanied by activation of caspase-8, caspase-9, and caspase-3 and was concomitant with Bid and PARP cleavage. We observed significant decreases in XIAP, Bcl-2, and c-FLIP after cotreatment with 15d-PGJ₂ and TRAIL. We also observed the inhibition of Akt expression and phosphorylation by cotreatment with 15d-PGJ₂ and TRAIL. Furthermore, inactivation of Akt by Akt inhibitor IV sensitized human leukemic HL-60 cells to TRAIL, indicating a key role for Akt inhibition in these events. Taken together, these findings indicate that 15d-PGJ₂ may augment TRAIL-induced apoptosis in human leukemia cells by down-regulating the expression and phosphorylation of Akt.     

Epidermal growth factor receptor-mediated tissue transglutaminase overexpression couples acquired tumor necrosis factor-related apoptosis-inducing ligand resistance and migration through c-FLIP and MMP-9 proteins in lung cancer cells

Acquired chemoresistance not only blunts anticancer therapy but may also promote cancer cell migration and metastasis. Our previous studies have revealed that acquired tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) resistance in lung cancer cells is associated with Akt-mediated stabilization of cellular caspase 8 and Fas-associated death domain (FADD)-like apoptosis regulator-like inhibitory protein (c-FLIP) and myeloid cell leukemia 1 (Mcl-1). In this report, we show that cells with acquired TRAIL resistance have significantly increased capacities in migration and invasion. By gene expression screening, tissue transglutaminase (TGM2) was identified as one of the genes with the highest expression increase in TRAIL-resistant cells. Suppressing TGM2 dramatically alleviated TRAIL resistance and cell migration, suggesting that TGM2 contributes to these two phenotypes in TRAIL-resistant cells. TGM2-mediated TRAIL resistance is likely through c-FLIP because TGM2 suppression significantly reduced c-FLIP but not Mcl-1 expression. The expression of matrix metalloproteinase 9 (MMP-9) was suppressed when TGM2 was inhibited, suggesting that TGM2 potentiates cell migration through up-regulating MMP-9 expression. We found that EGF receptor (EGFR) was highly active in the TRAIL-resistant cells, and suppression of EGFR dramatically reduced TGM2 expression. We further determined JNK and ERK, but not Akt and NF-κB, are responsible for EGFR-mediated TGM2 expression. These results identify a novel pathway that involves EGFR, MAPK (JNK and ERK), and TGM2 for acquired TRAIL resistance and cell migration in lung cancer cells. Because TGM2 couples TRAIL resistance and cell migration, it could be a molecular target for circumventing acquired chemoresistance and metastasis in lung cancer.     

Delivery of sTRAIL variants by MSCs in combination with cytotoxic drug treatment leads to p53-independent enhanced antitumor effects

Mesenchymal stem cells (MSCs) are able to infiltrate tumor tissues and thereby effectively deliver gene therapeutic payloads. Here, we engineered murine MSCs (mMSCs) to express a secreted form of the TNF-related apoptosis-inducing ligand (TRAIL), which is a potent inducer of apoptosis in tumor cells, and tested these MSCs, termed MSC.sTRAIL, in combination with conventional chemotherapeutic drug treatment in colon cancer models. When we pretreated human colorectal cancer HCT116 cells with low doses of 5-fluorouracil (5-FU) and added MSC.sTRAIL, we found significantly increased apoptosis as compared with single-agent treatment. Moreover, HCT116 xenografts, which were cotreated with 5-FU and systemically delivered MSC.sTRAIL, went into remission. Noteworthy, this effect was protein 53 (p53) independent and was mediated by TRAIL-receptor 2 (TRAIL-R2) upregulation, demonstrating the applicability of this approach in p53-defective tumors. Consequently, when we generated MSCs that secreted TRAIL-R2-specific variants of soluble TRAIL (sTRAIL), we found that such engineered MSCs, labeled MSC.sTRAIL^DR5, had enhanced antitumor activity in combination with 5-FU when compared with MSC.sTRAIL. In contrast, TRAIL-resistant pancreatic carcinoma PancTu1 cells responded better to MSC.sTRAIL^DR4 when the antiapoptotic protein XIAP (X-linked inhibitor of apoptosis protein) was silenced concomitantly. Taken together, our results demonstrate that TRAIL-receptor selective variants can potentially enhance the therapeutic efficacy of MSC-delivered TRAIL as part of individualized and tumor-specific combination treatments.