Combination Treatment Of Tumor Necrosis Factor-related Apoptosis-inducing Ligand Research Articles

The HDAC1 Inhibitor CBUD-1001 Enhances TRAIL-induced Apoptosis in Colorectal Cancer Cells.

Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) is a potential anti-tumor agent. However, resistance to TRAIL has been reported in a number of clinical trials. In this study, we investigated the molecular mechanisms by which a novel histone deacetylase (HDAC) inhibitor, CBUD-1001, sensitizes colorectal cancer (CRC) cells to TRAIL-induced apoptosis. Apoptotic cell death induced by CBUD-1001 and/or TRAIL was assessed on human CRC cells using the MTT assay, FACS analysis and nuclei staining. The involved molecular mechanisms were explored through western blotting analysis. We demonstrated that combined with CBUD-1001, TRAIL significantly enhanced TRAIL-induced apoptosis in CRC cells via mitochondria-mediated pathways. We also found that hyper-acetylation of histone by CBUD-1001 treatment leads to up-regulation of death receptor (DR) 5 in a dose- and time-dependent manner. Furthermore, we identified that enhanced sensitivity to TRAIL by combination with CBUD-1001 depends on the MAPK/CHOP axis, being a key mediator of DR5. A novel HDAC inhibitor CBUD-1001 sensitizes TRAIL-induced apoptosis via up-regulation of DR5, and that CBUD-1001 and TRAIL combination treatment offers an effective strategy to overcome TRAIL resistance in CRC cells.

Downregulation of c‑FLIP and upregulation of DR‑5 by cantharidin sensitizes TRAIL‑mediated apoptosis in prostate cancer cells via autophagy flux.

Tumor necrosis factor (TNF)-related apop-tosis-inducing ligand (TRAIL), a type II transmembrane protein, is a part of the TNF superfamily of cytokines. Cantharidin, a type of terpenoid, is extracted from the blister beetles (Mylabris genus) used in Traditional Chinese Medicine. Cantharidin elicits antibiotic, antiviral and antitumor effects, and can affect the immune response. The present study demonstrated that a cantharidin and TRAIL combination treatment regimen elicited a synergistic outcome in TRAIL-resistant DU145 cells. Notably, it was also identified that cantharidin treatment initiated the downregulation of cellular FLICE-like inhibitory protein (c-FLIP) and upregulation of death receptor 5 (DR-5), and sensitized cells to TRAIL-mediated apoptosis by initiating autophagy flux. In addition, cantharidin treatment increased lipid-modified microtubule-associated proteins 1A/1B light chain 3B expression and significantly attenuated sequestosome 1 expression. Attenuation of autophagy flux by a specific inhibitor such as chloroquine and genetic modification using ATG5 small interfering RNA abrogated the cantharidin-mediated TRAIL-induced apoptosis. Overall, the results of the present study revealed that cantharidin effectively sensitized cells to TRAIL-mediated apoptosis and its effects are likely to be mediated by autophagy, the downregulation of c-FLIP and the upregulation of DR-5. They also suggested that the combination of cantharidin and TRAIL may be a successful therapeutic strategy for TRAIL-resistant prostate cancer.

Trichothecenes from an Endophytic Fungus Alternaria sp. sb23.

Three new (alterchothecenes A - C, 1: -3: ) and 3 known (4: -6: ) trichothecenes, along with 9 known compounds (7: -15: ), were isolated from the culture of Alternaria sp. sb23, an endophytic fungus separated from the root of Schisandra sphenanthera Rehd. et Wils. Their structures were elucidated by spectroscopic analyses, and the absolute configurations of 1: -3: were determined through comparison of the experimental electronic circular dichroism (ECD) spectra and optical rotations with similar analogues. In vitro cytotoxicity tests of compounds 1: -6: against human HT-29 colon carcinoma and human MCF-7 breast cancer cell lines indicated that 4: -6: exhibited significant cytotoxic effects, with IC50 values ranging from 0.89 to 9.38 µM. And the potential of compounds 1: -6: as tumor necrosis factor (TNF)-related apoptosis-inducing ligand (TRAIL) sensitizers in HT-29 cells was evaluated. The results revealed that combination treatment of TRAIL with compounds 1: -6: synergistically decreased cell viability compared with the sole treatment with those compounds.

Herbal Medicines Showing Synergistic Effects with Tumor Necrosis Factor-Related Apoptosis-Inducing Ligand (TRAIL) against A549 TRAIL-resistant Lung Cancer Cells: A Screening Study.

Background:Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) is a cytokine that activates apoptosis through death receptors on the cell surface and is regarded as a potential anticancer agent. However, many cancer cells are resistant to TRAIL-induced apoptosis.Objective:The aim is to identify the herbal medicines that could help overcome resistance in TRAIL-resistant lung cancer cells.Materials and Methods;TRAIL-resistant A549 cells and 13 herbal medicines with known apoptosis-related anticancer effects were used in this study: Clematidis Radix, Corydalis Tuber Rhizoma, Paeoniae Radix Rubra, Corni Fructus, Curcumae longae Rhizoma (CLR), Moutan Cortex, Salviae miltiorrhizae Radix, Phellodendri Cortex, Farfarae Flos, Paeoniae Radix Alba, Angelicae gigantis Radix, Coptidis Rhizoma (CR), and Taraxaci Herba. Cytotoxic effects were investigated after a 48-h incubation, using an 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide assay, to identify the herbal medicines with the most potent synergistic effects with TRAIL.Results:The majority of the 13 medicines exhibited concentration-dependent cytotoxicity against A549 cells. Among them, CR and CLR showed the most potent cytotoxic effects, based on the IC50. We then investigated the use of these two medicines in combination with TRAIL and identified synergistic cytotoxic effects against TRAIL-resistant A549 cells.Conclusion:Synergistic cytotoxic effects of the combination of TRAIL and herbal medicines, in particular, CR and CLR, were confirmed in A549 cells. Therefore, CR and CLR showed potential to be used as candidates to overcome TRAIL resistance. Future studies to identify their underlying mechanism of action are required.SUMMARY Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) is an attractive anticancer agent which can induce apoptosis in tumor cells without causing cytotoxicity to normal cellsHowever, resistance to TRAIL is often observed in some tumor cells, including nonsmall cell lung cancers, which may limit its cytotoxic efficacy in cancer treatmentThe combination treatment of TRAIL and herbal medicines, particularly Coptidis Rhizoma (CR) and Curcumae longae Rhizoma (CLR), can induce the synergistic cytotoxic effects against TRAIL-resistant A549 cells, indicating that TRAIL resistance was reduced by combination therapy. Abbreviations used: TRAIL: Tumor necrosis factor-related apoptosis-inducing ligand; CLR: Curcumae longae Rhizoma; CR: Coptidis Rhizoma; NSCLC: non-small cell lung cancer.

Morusin Induces TRAIL Sensitization by Regulating EGFR and DR5 in Human Glioblastoma Cells.

Glioblastoma is one of the most malignant primary tumors, and the prognosis for glioblastoma patients remains poor. Tumor-necrosis-factor-related apoptosis-inducing ligand (TRAIL) is considered a promising anticancer agent due to its remarkable ability to selectively kill tumor cells. However, since many cancers are resistant to TRAIL, strategies to overcome resistance are required for the successful use of TRAIL in the clinic. In the present study, the potential of morusin as a TRAIL sensitizer in human glioblastoma cells was evaluated. Treatment with TRAIL or morusin alone showed weak cytotoxicity in human glioblastoma cells. However, combination treatment of TRAIL with morusin synergistically decreased cell viability and increased apoptosis compared with single treatment. Morusin induced expression of death receptor 5 (DR5), but not DR4 or decoy receptors (DcR1 and DcR2). Furthermore, morusin significantly decreased anti-apoptotic molecules survivin and XIAP. In addition, morusin reduced expression of EGFR and PDFGR as well as phosphorylation of STAT3, possibly mediating down-regulation of survivin and XIAP. Together these results suggest that morusin enhances TRAIL sensitivity in human glioblastoma cells through regulating expression of DR5 and EGFR. Therefore, the combination treatment of TRAIL and morusin may be a new therapeutic strategy for malignant glioma patients.

Downregulation of Nrf2 by the combination of TRAIL and Valproic acid induces apoptotic cell death of TRAIL-resistant papillary thyroid cancer cells via suppression of Bcl-xL

Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) represents an effective agent for the treatment of many cancers, though the majority of thyroid cancers are found to be resistant. Therefore it would be necessary to identify agents capable of increasing the sensitivity of these cancers to TRAIL-mediated cell death. Here, we examined the therapeutic effect and its underlying mechanism of combination treatment of TRAIL and histone deacetylase inhibitor, Valproic acid (VPA) in vitro using human papillary thyroid cancer (PTC) cells and in vivo using an orthotopic mouse model of PTC. TRAIL-VPA combination therapy synergistically induced apoptotic cell death in TRAIL-resistant PTC through caspase activation. In addition, downregulation of antioxidant transcription factor, Nrf2 by co-treatment of TRAIL-VPA induces cell death via suppression of Bcl-xL in vitro and in vivo; these effects were further enhanced following siRNA inhibition of these proteins in combination with TRAIL or TRAIL-VPA. Taken together, VPA sensitized TRAIL-resistant PTC cells to apoptotic cell death through involvement of Nrf2 and Bcl-xL. Thus, the combination of VPA and TRAIL may be a promising therapy for TRAIL-resistant PTC.

Abstract B34: Sensitization of TRAIL-resistant malignant melanomas by ellagic acid

Abstract The incidence of malignant melanoma has been on the rise for the past 30 years with a lifetime risk of 1 in 75. In its early stage, melanoma is easily cured, but the prognosis associated with metastatic malignant melanoma remains very poor and is one of the most treatment-refractory malignancies. FDA-approved melanoma therapeutics, Dacarbazine (DTIC) and Interleukin-2 (IL-2) are associated with response rates of only 10-20% and do not yield long-lasting remissions. A direct and selective p53-independent cancer therapy is the application of Tumor necrosis factor-Related Apoptosis-Inducing Ligand (TRAIL). TRAIL can induce apoptosis in a broad range of transformed human cells while showing minimal toxicity towards non-malignant cells. However, some cancers are resistant to rhTRAIL-induced apoptosis as a direct result of overexpressed antiapoptotic proteins. One key driver mediating the expression of such antiapoptotic proteins is Casein Kinase 2 (CK2). Upregulated in the nucleus of all cancer cells, CK2 serves as an oncoprotein, promoting the expression of inhibitors of apoptosis resulting in resistance to cell death. By inhibiting CK2 and, therefore, downregulating its antiapoptotic products, TRAIL-resistant cells may become more sensitive to TRAIL-induced apoptosis. To date, Ellagic Acid (EA) is the most potent, naturally-occurring CK2 inhibitor, possessing a Ki of 20 nM. To test whether the addition of EA could sensitize cancer cells to the effects of TRAIL, we utilized cotreatment with TRAIL and EA against the V600E BRAF-mutated melanoma cell lines A375 and WM164, which have previously been reported as TRAIL-resistant. Our in vitro analyses showed that the cotreatment of TRAIL plus EA does not augment the ability of TRAIL to induce apoptosis in A375 as determined by Annexin-V Apoptosis assays and Western Blot analysis. TRAIL-alone was able to induce apoptosis in A375 characterized by the formation of Annexin-V positive cells, PARP fragmentation and the activation of caspase 3. TRAIL initiated both the extrinsic and intrinsic pathways of apoptosis marked by caspase 8-activation and the mitochondrial release of cytochrome c, respectively. However, EA treatment alone did not induce apoptosis marked by the absence of Annexin-V positive cells and PARP fragmentation. Additionally, both TRAIL and EA, as single-agents, inhibited the growth of A375 in a dose-dependent manner, and the combination treatment of TRAIL plus EA enhanced the cytostatic effects of the single-agent treatments as determined by Sulforhodamine-B (SRB) colorimetric assays. Cell cycle analysis through Propidium Iodide (PI) staining and FACS analysis revealed that EA impeded cell growth through an EA-mediated S-phase arrest in A375. Microarray analysis showed that EA downregulates antiapoptotic proteins survivin, XIAP, cIAP-2 and cIAP-1 and upregulates the death receptor DR5. In vivo, we showed that the combination treatment of TRAIL plus EA did not potentiate the ability of TRAIL to inhibit the growth of established A375 tumors. TRAIL alone inhibited tumor growth by ∼80% and H&E staining of TRAIL-treated tumors presented evidence of apoptotic cells. EA was not able to stop the growth of established A375 tumors and did not augment the in vivo antiproliferative effects of TRAIL. Lastly, we showed that TRAIL is cytotoxic towards WM164 cells and can reduce the viability of A375 as determined by Trypan Blue exclusion assays. Whereas EA, although it can inhibit the growth of WM164, does not reduce the viability of the cells. Additional analysis of the potential of EA to sensitize WM164 to the effects of TRAIL is ongoing. These data demonstrate that EA can enhance the cytostatic but not the cytotoxic or proapoptotic effects of TRAIL in vitro. In vivo, TRAIL can inhibit the growth of established tumors as a single-agent but is not augmented by cotreatment with EA. Citation Format: Katherine A. Turner, Daniel Lindner, Michael Kalafatis. Sensitization of TRAIL-resistant malignant melanomas by ellagic acid. [abstract]. In: Proceedings of the AACR Special Conference on Advances in Melanoma: From Biology to Therapy; Sep 20-23, 2014; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(14 Suppl):Abstract nr B34.

Combination treatment of TRAIL, DFMO and radiation for malignant glioma cells.

Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) has shown potent and cancer-selective killing activity and drawn considerable attention as a promising therapy for cancer. Another promising cancer therapy is difluoromethylornithine (DFMO), an inhibitor of ornithine decarboxylase, which is oraly administered and well tolerated. Nevertheless, many types of cancer, including gliomas, have exhibited resistance to TRAIL-induced apoptosis and similarly the potency of DFMO should be enhanced to optimize therapeutic efficacy. In this study we sought to determine whether DFMO, in combination with TRAIL and radiation, could result in an enhanced anti-glioma effect in vitro. We investigated the effect of DFMO, TRAIL and radiation in various combinations on a panel of glioblastoma cell lines (A172, T98G, D54, U251MG). Viability and proliferation of the cells were examined with trypan blue exclusion assay, crystal violet and xCELLigence system. Apoptosis (Annexin-PI), cell cycle and activation of caspase-8 were tested with flow cytometry. BAD protein levels were determined by Western blot analysis. DFMO induced BAD overexpression. Combination treatment with DFMO, TRAIL and radiation significantly reduced cell viability in all cell lines tested. Increased induction of cell death and cell cycle arrest was confirmed with flow cytometry in A172 and D54 cell lines, while enhanced activation of annexin and caspase-8 was revealed in U251MG and T98G cells. The treatment of glioblastoma cell lines with combination of DFMO, TRAIL and radiation showed an enhanced effect. This combination treatment may represent a novel strategy for targeting glioblastoma.

Icaritin Sensitizes Human Glioblastoma Cells to TRAIL-Induced Apoptosis.

Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) has been considered to be one of the most promising candidates in research on treatments for cancer, because it induces apoptosis in a wide variety of cancer cells but not in most normal human cell types. However, many cells including glioblastoma (GBM) cells are resistant to TRAIL-induced apoptosis, which limits the potential application of TRAIL in cancer therapy. Icaritin, a hydrolytic product of icariin from Epimedium Genus, has been identified as a potential therapeutic and preventive agent in renal cell carcinoma and breast cancer. In this study, we investigated whether Icaritin treatment could modulate TRAIL-induced apoptosis in GBM. The effect of icaritin on TRAIL sensitivity was assessed in human GBM U87 and U373 cells. The underlying regulatory cascades were approached by biochemical and pharmacological strategies. We found that nontoxic concentration of icaritin alone had no significant effect on the level of apoptosis, but a combination treatment of TRAIL and icaritin caused a significantly more profound apoptosis. The sensitization was accompanied by c-FLIP down-regulation and inhibition of NF-κB activity. Studies have further demonstrated that silencing NF-κB alone was sufficient to down-regulate c-FLIP expression and sensitized both tested cells to TRAIL-induced apoptosis. These data suggest that icaritin sensitizes TRAIL-induced tumor cell apoptosis via suppression of NF-κB-dependent c-FLIP expression, providing in vitro evidence supporting the notion that icaritin is a potential sensitizer of TRAIL in anticancer therapy against human GBM.

Abstract 2289: Effects of TRAIL and ellagic acid on malignant melanoma A375 in vitro and in vivo

Abstract Cancer is a widespread disease characterized by unregulated cell growth and evasion of apoptotic stimuli. Most conventional therapies lack cancer cell specificity and induce cell death indirectly through cellular damage in a p53-dependent manner. A direct and selective p53-independent cancer therapy is the application of Tumor necrosis factor-Related Apoptosis-Inducing Ligand (TRAIL). Recombinant human TRAIL (rhTRAIL) can induce apoptosis in a broad range of transformed human cells while showing minimal toxicity towards non-malignant cells. However, some cancers are resistant to rhTRAIL-induced apoptosis as a direct result of overexpressed antiapoptotic proteins. One key driver mediating the expression of such antiapoptotic proteins is Casein Kinase 2 (CK2). CK2 is upregulated in the nucleus of all cancer cells and promotes the expression of inhibitors of apoptosis resulting in resistance to cell death. Therefore, by inhibiting CK2, TRAIL-resistant cells may become more sensitive to TRAIL-induced apoptosis. We utilized cotreatment with Ellagic Acid (EA), a natural CK2 inhibitor, and TRAIL against the malignant melanoma cell line A375, which is resistant to TRAIL. Our in vitro analysis showed that TRAIL and EA, as single-agents, can inhibit cell growth in a dose-dependent manner, and the combination treatment of TRAIL plus EA synergistically enhanced the cytostatic effects of the single-agent treatments. We show that TRAIL reduces the viability of A375 cells; whereas, EA although it can inhibit growth, cannot reduce the viability of A375 as determined by Trypan Blue exclusion assays. In combination treatments, the cytotoxic effects of TRAIL were not enhanced. Annexin-V Apoptosis assays and Western Blot analysis show that TRAIL can induce apoptosis in A375 cells characterized by the formation of Annexin-V positive cells, PARP fragmentation and the activation of caspase 3. Additionally, TRAIL initiated both the extrinsic and intrinsic pathways of apoptosis marked by caspase 8 activation and the mitochondrial release of cytochrome c, respectively. EA treatment alone did not induce apoptosis and did not induce formation of Annexin-V positive cells or PARP fragmentation. Combination treatments show that EA does not potentiate TRAIL-induced apoptosis. In vivo, TRAIL-treated nude mice bearing A375 xenografts exhibited ∼80% tumor growth inhibition and a prolonged survival rate compared to the untreated control group. EA was not able to inhibit the growth of established A375 tumors and did not augment the in vivo antiproliferative effects of TRAIL. These data demonstrate that EA can enhance the cytostatic but not the cytotoxic or proapoptotic effects of TRAIL in vitro. In vivo, TRAIL can inhibit the growth of established tumors as a single-agent but is not augmented by cotreatment with EA. Citation Format: Katherine Turner, Daniel Lindner, Michael Kalafatis. Effects of TRAIL and ellagic acid on malignant melanoma A375 in vitro and in vivo. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 2289. doi:10.1158/1538-7445.AM2014-2289

P-0020 Evodiamine Augments TNF-α- and Trail-Induced Apoptosis in Human Gastric Cancer Cells Via Inhibiting The NF-κB and PI3K/Akt Signaling Pathways

ABSTRACT Introduction Traditional medicines not only provide important notion for finding novel drugs, also may supportive to reallocate drug discovery paradigm from ‘finding new-entity drugs’ to ‘combining existing agents and might even the combinations between such agents. Previously, we found that Evodiamine which is isolated from, Chinese traditional medicine, Evodia rutaecarpa significantly induces apoptosis in gastric cancer cells. Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) selectively induces apoptosis and kills cancer cells with little or no adverse effects on normal cells. However, gastric cancer cells are less sensitive than other cancer cells to TRAIL-induced apoptosis and effect of Evodiamine on TNF-a- and TRAIL-induced apoptosis in human gastric cancer cells remain unexplored. This study was designed to investigate whether Evodiamine augments TNF-a- and TRAIL-induced apoptosis in gastric cancer cells. Methods The growth inhibitory effect of the Evodiamine, TNF-a, TRAIL, and their combinations on the viability of cells was determined by the MTT assay. Apoptosis and mitochondrial membrane potential was evaluated by flow cytometry and expression of pro-apoptotic and pro-survival proteins were analyzed by Western blotting. Results In the present study, we found that Evodiamine enhanced TNF-a- and TRAIL-induced apoptosis in human gastric cancer cells. As indicated by assays that measure mitochondrial membrane integrity, pro-survival proteins, and activation of caspase-8, caspase-9, and caspase-3, Evodiamine potentiated the TRAIL-induced apoptosis in human gastric cancer cells and converted TRAIL-resistant cells to TRAIL-sensitive cells. Here, we demonstrate the first evidence that Evodiamine effectively enhances TNF-a- and TRAIL-mediated cytotoxicity by suppressing pro-survival proteins including cFLIP, Bcl-2, and survivin and up-regulates Bax expression. Upon treatment with Evodiamine, the levels of survival proteins were strongly suppressed in gastric cancer cells. The down-regulation of these survival proteins could be regulated by repressing activation of NF-kB and Akt. Evodiamine also inhibited TNF-a- and TRAIL-induced transcriptional activity of NF-kB. In addition, this substance significantly enhanced both extrinsic and intrinsic apoptosis, which were induced by TRAIL. Conclusion Taken together, the results of the present study suggest that Evodiamine exhibits an ability to overcome TRAIL resistance and combination treatment of TRAIL and TNF-a together with Evodiamine may be an effective regimen for the treatment of advanced gastric cancer.

P2.18 Augmentation of Trail-Induced Apoptosis by Tubeimoside-1 in Human Gastric Cancer Cells Via Inhibition Of The Nf-ΚB and Pi3K/Akt Signaling Pathways

ABSTRACT Traditional medicines not only provide important notion for finding novel drugs, also may supportive to reallocate drug discovery paradigm from ‘finding new-entity drugs’ to ‘combining existing agents and may even the combinations between such agents. Previously, we found that Tubeimoside-1 (TBMS-1), a triterpenoid saponin from traditional Chinese medicine, significantly induces apoptosis in gastric cancer cells. Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) selectively induces apoptosis and kills cancer cells with little or no adverse effects on normal cells. However, gastric cancer cells are less sensitive than other cancer cells to TRAIL-induced apoptosis. In the present study, we found that Tubeimoside-1 enhanced TRAIL-induced apoptosis in human gastric adenocarcinoma SGC-7901 cells. As indicated by assays that measure mitochondrial membrane integrity, pro-survival proteins, and activation of caspase-8, caspase-9, and caspase-3, TBMS-1 potentiated the TRAIL-induced apoptosis in human gastric cancer cells, and converted TRAIL-resistant cells to TRAIL-sensitive cells. Here, we demonstrate the first evidence that TBMS-1 effectively enhances TRAIL-mediated cytotoxicity by suppressing pro-survival proteins including survivin and XIAP. Upon treatment with TBMS-1, the levels of survival proteins were strongly suppressed in SGC-7901 cells. The down-regulation of these survival proteins could be regulated by repressing activation of NF-κB and Akt. TBMS-1 also inhibited TRAIL-induced transcriptional activity of NF-κB. In addition, this substance significantly enhanced both extrinsic and intrinsic apoptosis, which were induced by TRAIL. Taken together, the results of the present study suggest that TBMS-1 exhibits an ability to overcome TRAIL resistance and combination treatment of TRAIL together with TBMS-1 may be an effective regimen for the treatment of advanced gastric cancer.

Inhibition of livin expression suppresses cell proliferation and enhances chemosensitivity to cisplatin in human lung adenocarcinoma cells

Livin is a novel member of the inhibitor of apoptosis protein family that has been reported to be overexpressed in various types of human malignancy. Although several studies have demonstrated that livin may be used as an effective target for tumor therapy, few studies have investigated its role in human lung adenocarcinoma. In the present study, two different methods were used in order to investigate the tumor‑suppressing effect of livin in human lung adenocarcinoma cells. Firstly, small interfering (si)RNA technology was used to down regulate livin expression; siRNA-mediated knockdown of livin was confirmed using reverse transcription quantitative polymerase chain reaction and western blot analysis, and cell proliferations was assessed using an MTT assay in vitro. Secondly, inhibition of livin expression was induced through the synergistic inhibitory effect between flavopiridol and tumor necrosis factor‑related apoptosis-inducing ligand (TRAIL). Experimental results revealed that, following transfection of the livin gene-silencing vector, the gene expression of livin was markedly decreased, SPC-A1 cell proliferation was significantly reduced and the therapeutic effect of the chemotherapy drug cisplatin was markedly improved. This growth inhibitory effect was also observed in the flavopiridol and TRAIL combination treatment group. In the flavopiridol and TRAIL combination treatment group, the protein expression of livin was significantly reduced and the survival rate of SPC‑A1 cells was significantly lower than the flavopiridol and TRAIL single operation group. In conclusion, the RNA silencing and the synergistic inhibitory effect between flavopiridol with TRAIL was able to effectively inhibit the expression of livin, significantly decrease SPC-A1 tumor cell proliferation and significantly enhance sensitivity to the chemotherapy drug cisplatin. These findings suggest that livin may be used as a novel target for tumor gene therapy.

Artesunate enhances TRAIL-induced apoptosis in human cervical carcinoma cells through inhibition of the NF-κB and PI3K/Akt signaling pathways

Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) selectively induces apoptosis and kills cancer cells with little or no adverse effects on normal cells. TRAIL is relatively safe for clinical applications. However, TRAIL resistance is widely found in cancer cells leading to limitations in utilizing TRAIL as a therapeutic agent for cancer treatment. Recently, artesunate, an effective and safe anti-malarial drug, was also described as a promising candidate for cancer therapy. It would be of importance to determine whether combination treatment of TRAIL together with artesunate could overcome drug-resistance of tumors. Here, we demonstrate the first evidence that artesunate effectively enhances TRAIL-mediated cytotoxicity by suppressing pro-survival proteins, such as survivin, XIAP and Bcl-XL. Upon treatment with artesunate, the levels of survival proteins were strongly suppressed in HeLa cells. The down-regulation of these survival proteins could be regulated by repressing activation of NF-κB and Akt. Artesunate also inhibited TRAIL-induced transcriptional activity of NF-κB. In addition, this substance significantly enhanced both extrinsic and intrinsic apoptosis, which were induced by TRAIL. Taken together, the results of the present study suggest that artesunate exhibits an ability to overcome TRAIL resistance and combination treatment of TRAIL together with artesunate may be an effective strategy for cancer therapy.

Differential responsiveness of human hepatoma cells versus normal hepatocytes to TRAIL in combination with either histone deacetylase inhibitors or conventional cytostatics

Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) is a promising candidate for the treatment of cancer because it elicits cell death in many tumor cells while sparing most normal cells. Liver cancer, however, is largely resistant to TRAIL and, thus, requires sensitization for TRAIL-mediated cytotoxicity. Sensitization may be achieved by cotreatment with chemotherapeutic agents. In this study, we comparatively investigated the treatment efficacy of TRAIL in combination with histone deacetylase inhibitors (HDI) versus TRAIL in combination with conventional cytostatics in the hepatocellular carcinoma cell line HepG2 and in the childhood hepatoblastoma cell line Huh6. We found that TRAIL resistance could be overcome by cotreatment with the HDI vorinostat, sodium butyrate and MS-275, but not by cotreatment with the cytostatics carboplatin and etoposide. However, TRAIL combination treatment bears the risk of sensitizing otherwise TRAIL-resistant normal cells. We thus explored a potential cytotoxic effect of combined HDI/TRAIL treatment in normal hepatocytes: TRAIL in conjunction with HDI did not impose any cytotoxicity on the non-malignant cells. In searching for the determinants of HDI-mediated TRAIL sensitization in hepatoma cells, we observed that HDI treatment did not increase cell-surface expression of proapoptotic TRAIL receptors. Instead, HDI treatment enhanced TRAIL-induced cleavage of Bid. In conclusion, our data suggest that HDI are potent sensitizers to TRAIL in hepatoma cells and that the combination of HDI and TRAIL is selectively active in hepatoma cells without affecting normal hepatocytes, indicating that the combination of HDI and TRAIL may be an effective approach for the treatment of advanced liver cancer.