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A Novel Recombinant Fcγ Receptor-Targeted Survivin Combines with Chemotherapy for Efficient Cancer Treatment. Biomedicines 2021; 9:biomedicines9070806. [PMID: 34356870 PMCID: PMC8301409 DOI: 10.3390/biomedicines9070806] [Citation(s) in RCA: 4] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/31/2021] [Revised: 07/03/2021] [Accepted: 07/09/2021] [Indexed: 01/03/2023] Open
Abstract
Formyl peptide receptor-like 1 inhibitor (FLIPr), an Fcγ receptor (FcγR) antagonist, can be used as a carrier to guide antigen-FLIPr fusion protein to FcγR then enhances antigen-specific immune responses. Survivin, a tumor-associated antigen, is over-expressed in various types of human cancer. In this study, we demonstrate that recombinant survivin-FLIPr fusion protein (rSur-FLIPr) binds to FcγRs, and efficient uptake by dendritic cells in vivo. In addition, rSur-FLIPr alone stimulates survivin-specific immune responses, which effectively suppresses the tumor growth. The antitumor immunities are through TAP-mediated and CD8-dependent pathways. Furthermore, preexisting anti-FLIPr antibody does not abolish antitumor responses induced by rSur-FLIPr immunization. These results suggest that FLIPr is an effective antigen delivery vector and can be repeatedly used. Combination of chemotherapy with rSur-FLIPr treatment reveals a great benefit to tumor-bearing mice. Altogether, these findings suggest that rSur-FLIPr is a potential candidate for efficient cancer therapy.
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2
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Wu M, Tong CWS, Yan W, To KKW, Cho WCS. The RNA Binding Protein HuR: A Promising Drug Target for Anticancer Therapy. Curr Cancer Drug Targets 2020; 19:382-399. [PMID: 30381077 DOI: 10.2174/1568009618666181031145953] [Citation(s) in RCA: 32] [Impact Index Per Article: 6.4] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/24/2018] [Revised: 08/24/2018] [Accepted: 10/18/2018] [Indexed: 02/07/2023]
Abstract
The stability of mRNA is one of the key factors governing the regulation of eukaryotic gene expression and function. Human antigen R (HuR) is an RNA-binding protein that regulates the stability, translation, and nucleus-to-cytoplasm shuttling of its target mRNAs. While HuR is normally localized within the nucleus, it has been shown that HuR binds mRNAs in the nucleus and then escorts the mRNAs to the cytoplasm where HuR protects them from degradation. It contains several RNA recognition motifs, which specifically bind to adenylate and uridylate-rich regions within the 3'-untranslated region of the target mRNA to mediate its effect. Many of the HuR target mRNAs encode proteins important for cell growth, tumorigenesis, angiogenesis, tumor inflammation, invasion and metastasis. HuR overexpression is known to correlate well with high-grade malignancy and poor prognosis in many tumor types. Thus, HuR has emerged as an attractive drug target for cancer therapy. Novel small molecule HuR inhibitors have been identified by high throughput screening and new formulations for targeted delivery of HuR siRNA to tumor cells have been developed with promising anticancer activity. This review summarizes the significant role of HuR in cancer development, progression, and poor treatment response. We will discuss the potential and challenges of targeting HuR therapeutically.
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Affiliation(s)
- Mingxia Wu
- School of Pharmacy, Faculty of Medicine, The Chinese University of Hong Kong, Hong Kong
| | - Christy W S Tong
- School of Pharmacy, Faculty of Medicine, The Chinese University of Hong Kong, Hong Kong
| | - Wei Yan
- School of Pharmacy, Faculty of Medicine, The Chinese University of Hong Kong, Hong Kong
| | - Kenneth K W To
- School of Pharmacy, Faculty of Medicine, The Chinese University of Hong Kong, Hong Kong
| | - William C S Cho
- Department of Clinical Oncology, Queen Elizabeth Hospital, Hong Kong
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Triggered Immune Response Induced by Antigenic Epitopes Covalently Linked with Immunoadjuvant-Pulsed Dendritic Cells as a Promising Cancer Vaccine. J Immunol Res 2020; 2020:3965061. [PMID: 32322595 PMCID: PMC7160722 DOI: 10.1155/2020/3965061] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.2] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/21/2019] [Revised: 03/03/2020] [Accepted: 03/11/2020] [Indexed: 12/19/2022] Open
Abstract
The success of peptide-based dendritic cell (DC) cancer vaccines mainly depends on the utilized peptides and selection of an appropriate adjuvant. Herein, we aimed to evoke a broad immune response against multiple epitopes concurrently in the presence of immunoadjuvant. Three synthetic HLA-A∗0201-restricted peptides were separately linked with HMGB1-derived peptide (SAFFLFCSE, denoted as HB100-108) as immunoadjuvant via double arginine (RR) linker and loaded onto human monocyte-derived DCs. Peptide uptake was detected by immunofluorescence microscopy and flow cytometry. The maturation and activation status of pulsed DCs were monitored by detection of the expression of specific markers and released cytokines. The ability of peptide-pulsed DCs to activate allogeneic T cells has been assessed by a degranulation assay and detection of secreted cytokines. The lytic activity of effector T cells against cancer cells in vitro was analyzed by a lactate dehydrogenase (LDH) assay. Results revealed that DCs efficiently take up peptides+HB100-108 and expressed higher levels of surface markers (HLA-ABC, HLA-DR, CD80, CD86, CD83, CD40, and CCR7) and proinflammatory cytokines (IL-6, IFN-γ, TNF-α, and IL-12) than control DCs, free peptide-pulsed DCs, and free HB100-108-pulsed DC groups. Moreover, peptides+HB100-108/pulsed DCs were capable of activating allogeneic T cells and enhance their lytic activity against a pancreatic cancer cell line (PANC-1) in vitro. These findings suggest that antigenic peptides covalently linked with HB100-108/pulsed DCs could be a promising strategy to improve the current DC-based cancer vaccines.
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Xu F, Sun Y, Yang SZ, Zhou T, Jhala N, McDonald J, Chen Y. Cytoplasmic PARP-1 promotes pancreatic cancer tumorigenesis and resistance. Int J Cancer 2019; 145:474-483. [PMID: 30614530 DOI: 10.1002/ijc.32108] [Citation(s) in RCA: 23] [Impact Index Per Article: 3.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/27/2018] [Revised: 11/26/2018] [Accepted: 12/20/2018] [Indexed: 12/16/2022]
Abstract
The poly(ADP-ribose) polymerases (PARP) play important roles in repairing damaged DNA during intrinsic cell death. We recently linked PARP-1 to death receptor (DR)-activated extrinsic apoptosis, the present studies sought to elucidate the function of cytoplasmic PARP-1 in pancreatic cancer tumorigenesis and therapy. Using human normal and pancreatic cancer tissues, we analyzed the prevalence of cytoplasmic PARP-1 expression. In normal human pancreatic tissues, PARP-1 expression was present in the nucleus; however, cytoplasmic PARP-1 expression was identified in pancreatic cancers. Therefore, cytoplasmic PARP-1 mutants were generated by site-direct mutagenesis, to determine a causative effect of cytoplasmic PARP-1 on pancreatic cancer tumorigenesis and sensitivity to therapy with TRA-8, a humanized DR5 antibody. PARP-1 cytoplasmic mutants rendered TRA-8 sensitive pancreatic cancer cells, BxPc-3 and MiaPaCa-2, more resistant to TRA-8-induced apoptosis; whereas wild-type PARP-1, localizing mainly in the nucleus, had no effects. Additionally, cytoplasmic PARP-1, but not wild-type PARP-1, increased resistance of BxPc-3 cells to TRA-8 therapy in a mouse xenograft model in vivo. Inhibition of PARP enzymatic activity attenuated cytoplasmic PARP-1-mediated TRA-8 resistance. Furthermore, increased cytoplasmic PARP-1, but not wild-type PARP-1, was recruited into the TRA-8-activated death-inducing signaling complex and associated with increased and sustained activation of Src-mediated survival signals. In contrast, PARP-1 knockdown inhibited Src activation. Taken together, we have identified a novel function and mechanism underlying cytoplasmic PARP-1, distinct from nuclear PARP-1, in regulating DR5-activated apoptosis. Our studies support an innovative application of available PARP inhibitors or new cytoplasmic PARP-1 antagonists to enhance TRAIL therapy for TRAIL-resistant pancreatic cancers.
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Affiliation(s)
- Fei Xu
- Department of Pathology, University of Alabama at Birmingham, Birmingham, AL.,Department of Gastrointestinal Surgery, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, China
| | - Yong Sun
- Department of Pathology, University of Alabama at Birmingham, Birmingham, AL
| | - Shan-Zhong Yang
- Department of Pathology, University of Alabama at Birmingham, Birmingham, AL
| | - Tong Zhou
- Department of Medicine, University of Alabama at Birmingham, Birmingham, AL
| | - Nirag Jhala
- Department of Pathology, Temple University, Philadelphia, PA
| | - Jay McDonald
- Department of Pathology, University of Alabama at Birmingham, Birmingham, AL.,Birmingham Veterans Affairs Medical Center, Research Department, Birmingham, AL
| | - Yabing Chen
- Department of Pathology, University of Alabama at Birmingham, Birmingham, AL.,Birmingham Veterans Affairs Medical Center, Research Department, Birmingham, AL
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5
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The application of mRNA-based gene transfer in mesenchymal stem cell-mediated cytotoxicity of glioma cells. Oncotarget 2018; 7:55529-55542. [PMID: 27487125 PMCID: PMC5342434 DOI: 10.18632/oncotarget.10835] [Citation(s) in RCA: 12] [Impact Index Per Article: 1.7] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/14/2015] [Accepted: 07/14/2016] [Indexed: 12/15/2022] Open
Abstract
Since the tumor-oriented homing capacity of mesenchymal stem cells (MSCs) was discovered, MSCs have attracted great interest in the research field of cancer therapy mainly focused on their use as carries for anticancer agents. Differing from DNA-based vectors, the use of mRNA-based antituor gene delivery benefits from readily transfection and mutagenesis-free. However, it is essential to verify if mRNA transfection interferes with MSCs' tropism and their antitumor properties. TRAIL- and PTEN-mRNAs were synthesized and studied in an in vitro model of MSC-mediated indirect co-culture with DBTRG human glioma cells. The expression of TRAIL and PTEN in transfected MSCs was verified by immunoblotting analysis, and the migration ability of MSCs after anticancer gene transfection was demonstrated using transwell co-cultures. The viability of DBTRG cells was determined with bioluminescence, live/dead staining and real time cell analyzer. An in vivo model of DBTRG cell-derived xenografted tumors was used to verify the antitumor effects of TRAIL- and PTEN-engineered MSCs. With regard to the effect of mRNA transfection on MSCs' migration toward glioma cells, an enhanced migration rate was observed with MSCs transfected with all tested mRNAs compared to non-transfected MSCs (p<0.05). TRAIL- and PTEN-mRNA-induced cytotoxicity of DBTRG glioma cells was proportionally correlated with the ratio of conditioned medium from transfected MSCs. A synergistic action of TRAIL and PTEN was demonstrated in the current co-culture model. The immunoblotting analysis revealed the apoptotic nature of the cells death in the present study. The growth of the xenografted tumors was significantly inhibited by the application of MSCPTEN or MSCTRAIL/PTEN on day 14 and MSCTRAIL on day 28 (p<0.05). The results suggested that anticancer gene-bearing mRNAs synthesized in vitro are capable of being applied for MSC-mediated anticancer modality. This study provides an experimental base for further clinical anticancer studies using synthesized mRNAs.
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Romeo C, Weber MC, Zarei M, DeCicco D, Chand SN, Lobo AD, Winter JM, Sawicki JA, Sachs JN, Meisner-Kober N, Yeo CJ, Vadigepalli R, Tykocinski ML, Brody JR. HuR Contributes to TRAIL Resistance by Restricting Death Receptor 4 Expression in Pancreatic Cancer Cells. Mol Cancer Res 2016; 14:599-611. [PMID: 27053682 PMCID: PMC5312260 DOI: 10.1158/1541-7786.mcr-15-0448] [Citation(s) in RCA: 44] [Impact Index Per Article: 4.9] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/12/2015] [Accepted: 03/22/2016] [Indexed: 12/11/2022]
Abstract
UNLABELLED Pancreatic ductal adenocarcinoma (PDA) is one of the most lethal cancers, in part, due to resistance to both conventional and targeted therapeutics. TRAIL directly induces apoptosis through engagement of cell surface Death Receptors (DR4 and DR5), and has been explored as a molecular target for cancer treatment. Clinical trials with recombinant TRAIL and DR-targeting agents, however, have failed to show overall positive outcomes. Herein, we identify a novel TRAIL resistance mechanism governed by Hu antigen R (HuR, ELAV1), a stress-response protein abundant and functional in PDA cells. Exogenous HuR overexpression in TRAIL-sensitive PDA cell lines increases TRAIL resistance whereas silencing HuR in TRAIL-resistant PDA cells, by siRNA oligo-transfection, decreases TRAIL resistance. PDA cell exposure to soluble TRAIL induces HuR translocation from the nucleus to the cytoplasm. Furthermore, it is demonstrated that HuR interacts with the 3'-untranslated region (UTR) of DR4 mRNA. Pre-treatment of PDA cells with MS-444 (Novartis), an established small molecule inhibitor of HuR, substantially increased DR4 and DR5 cell surface levels and enhanced TRAIL sensitivity, further validating HuR's role in affecting TRAIL apoptotic resistance. NanoString analyses on the transcriptome of TRAIL-exposed PDA cells identified global HuR-mediated increases in antiapoptotic processes. Taken together, these data extend HuR's role as a key regulator of TRAIL-induced apoptosis. IMPLICATIONS Discovery of an important new HuR-mediated TRAIL resistance mechanism suggests that tumor-targeted HuR inhibition increases sensitivity to TRAIL-based therapeutics and supports their re-evaluation as an effective treatment for PDA patients. Mol Cancer Res; 14(7); 599-611. ©2016 AACR.
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Affiliation(s)
- Carmella Romeo
- Division of Surgical Research, Department of Surgery, Jefferson Pancreas, Biliary, and Related Cancer Center, Sidney Kimmel Medical College, Thomas Jefferson University, Philadelphia, Pennsylvania. Department of Pathology, Anatomy, and Cell Biology, Sidney Kimmel Medical College, Thomas Jefferson University, Philadelphia, Pennsylvania
| | - Matthew C Weber
- Department of Pathology, Anatomy, and Cell Biology, Sidney Kimmel Medical College, Thomas Jefferson University, Philadelphia, Pennsylvania
| | - Mahsa Zarei
- Division of Surgical Research, Department of Surgery, Jefferson Pancreas, Biliary, and Related Cancer Center, Sidney Kimmel Medical College, Thomas Jefferson University, Philadelphia, Pennsylvania
| | - Danielle DeCicco
- Department of Pathology, Anatomy, and Cell Biology, Daniel Baugh Institute, Thomas Jefferson University, Philadelphia, Pennsylvania
| | - Saswati N Chand
- Division of Surgical Research, Department of Surgery, Jefferson Pancreas, Biliary, and Related Cancer Center, Sidney Kimmel Medical College, Thomas Jefferson University, Philadelphia, Pennsylvania
| | - Angie D Lobo
- Department of Pathology, Anatomy, and Cell Biology, Sidney Kimmel Medical College, Thomas Jefferson University, Philadelphia, Pennsylvania
| | - Jordan M Winter
- Division of Surgical Research, Department of Surgery, Jefferson Pancreas, Biliary, and Related Cancer Center, Sidney Kimmel Medical College, Thomas Jefferson University, Philadelphia, Pennsylvania
| | - Janet A Sawicki
- Lankenau Institute for Medical Research, Wynnewood, Pennsylvania
| | - Jonathan N Sachs
- Department of Biomedical Engineering, University of Minnesota, Minneapolis, Minnesota
| | | | - Charles J Yeo
- Division of Surgical Research, Department of Surgery, Jefferson Pancreas, Biliary, and Related Cancer Center, Sidney Kimmel Medical College, Thomas Jefferson University, Philadelphia, Pennsylvania
| | - Rajanikanth Vadigepalli
- Department of Pathology, Anatomy, and Cell Biology, Daniel Baugh Institute, Thomas Jefferson University, Philadelphia, Pennsylvania
| | - Mark L Tykocinski
- Department of Pathology, Anatomy, and Cell Biology, Sidney Kimmel Medical College, Thomas Jefferson University, Philadelphia, Pennsylvania
| | - Jonathan R Brody
- Division of Surgical Research, Department of Surgery, Jefferson Pancreas, Biliary, and Related Cancer Center, Sidney Kimmel Medical College, Thomas Jefferson University, Philadelphia, Pennsylvania.
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Common Variants in CLDN2 and MORC4 Genes Confer Disease Susceptibility in Patients with Chronic Pancreatitis. PLoS One 2016; 11:e0147345. [PMID: 26820620 PMCID: PMC4731142 DOI: 10.1371/journal.pone.0147345] [Citation(s) in RCA: 26] [Impact Index Per Article: 2.9] [Reference Citation Analysis] [Abstract] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/22/2015] [Accepted: 12/31/2015] [Indexed: 01/09/2023] Open
Abstract
A recent genome-wide association study (GWAS) identified association with variants in X-linked CLDN2 and MORC4, and PRSS1-PRSS2 loci with chronic pancreatitis (CP) in North American patients of European ancestry. We selected 9 variants from the reported GWAS and replicated the association with CP in Indian patients by genotyping 1807 unrelated Indians of Indo-European ethnicity, including 519 patients with CP and 1288 controls. The etiology of CP was idiopathic in 83.62% and alcoholic in 16.38% of 519 patients. Our study confirmed a significant association of 2 variants in CLDN2 gene (rs4409525-OR 1.71, P = 1.38 x 10-09; rs12008279-OR 1.56, P = 1.53 x 10-04) and 2 variants in MORC4 gene (rs12688220-OR 1.72, P = 9.20 x 10-09; rs6622126-OR 1.75, P = 4.04x10-05) in Indian patients with CP. We also found significant association at PRSS1-PRSS2 locus (OR 0.60; P = 9.92 x 10-06) and SAMD12-TNFRSF11B (OR 0.49, 95% CI [0.31-0.78], P = 0.0027). A variant in the gene MORC4 (rs12688220) showed significant interaction with alcohol (OR for homozygous and heterozygous risk allele -14.62 and 1.51 respectively, P = 0.0068) suggesting gene-environment interaction. A combined analysis of the genes CLDN2 and MORC4 based on an effective risk allele score revealed a higher percentage of individuals homozygous for the risk allele in CP cases with 5.09 fold enhanced risk in individuals with 7 or more effective risk alleles compared with individuals with 3 or less risk alleles (P = 1.88 x 10-14). Genetic variants in CLDN2 and MORC4 genes were associated with CP in Indian patients.
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Yuan K, Yong S, Xu F, Zhou T, McDonald JM, Chen Y. Calmodulin antagonists promote TRA-8 therapy of resistant pancreatic cancer. Oncotarget 2015; 6:25308-19. [PMID: 26320171 PMCID: PMC4694833 DOI: 10.18632/oncotarget.4490] [Citation(s) in RCA: 27] [Impact Index Per Article: 2.7] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/06/2015] [Accepted: 06/30/2015] [Indexed: 12/20/2022] Open
Abstract
Pancreatic cancer is highly malignant with limited therapy and a poor prognosis. TRAIL-activating therapy has been promising, however, clinical trials have shown resistance and limited responses of pancreatic cancers. We investigated the effects of calmodulin(CaM) antagonists, trifluoperazine(TFP) and tamoxifen(TMX), on TRA-8-induced apoptosis and tumorigenesis of TRA-8-resistant pancreatic cancer cells, and underlying mechanisms. TFP or TMX alone did not induce apoptosis of resistant PANC-1 cells, while they dose-dependently enhanced TRA-8-induced apoptosis. TMX treatment enhanced efficacy of TRA-8 therapy on tumorigenesis in vivo. Analysis of TRA-8-induced death-inducing-signaling-complex (DISC) identified recruitment of survival signals, CaM/Src, into DR5-associated DISC, which was inhibited by TMX/TFP. In contrast, TMX/TFP increased TRA-8-induced DISC recruitment/activation of caspase-8. Consistently, caspase-8 inhibition blocked the effects of TFP/TMX on TRA-8-induced apoptosis. Moreover, TFP/TMX induced DR5 expression. With a series of deletion/point mutants, we identified CaM antagonist-responsive region in the putative Sp1-binding domain between -295 to -300 base pairs of DR5 gene. Altogether, we have demonstrated that CaM antagonists enhance TRA-8-induced apoptosis of TRA-8-resistant pancreatic cancer cells by increasing DR5 expression and enhancing recruitment of apoptotic signal while decreasing survival signals in DR5-associated DISC. Our studies support the use of these readily available CaM antagonists combined with TRAIL-activating agents for pancreatic cancer therapy.
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Affiliation(s)
- Kaiyu Yuan
- Department of Pathology, University of Alabama at Birmingham, Alabama 35294, Birmingham, USA
| | - Sun Yong
- Department of Pathology, University of Alabama at Birmingham, Alabama 35294, Birmingham, USA
| | - Fei Xu
- Department of Pathology, University of Alabama at Birmingham, Alabama 35294, Birmingham, USA
| | - Tong Zhou
- Department of Medicine, University of Alabama at Birmingham, Alabama 35294, Birmingham, USA
| | - Jay M McDonald
- Department of Pathology, University of Alabama at Birmingham, Alabama 35294, Birmingham, USA
- Birmingham Veterans Affairs Medical Center, Alabama 35294, Birmingham, USA
| | - Yabing Chen
- Department of Pathology, University of Alabama at Birmingham, Alabama 35294, Birmingham, USA
- Birmingham Veterans Affairs Medical Center, Alabama 35294, Birmingham, USA
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Xiao M, Li W. Recent Advances on Small-Molecule Survivin Inhibitors. Curr Med Chem 2015; 22:1136 - 1146. [PMID: 25613234 DOI: 10.2174/0929867322666150114102146] [Citation(s) in RCA: 26] [Impact Index Per Article: 2.6] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/14/2014] [Revised: 12/07/2014] [Accepted: 12/09/2014] [Indexed: 12/18/2022]
Abstract
Survivin, a member of the inhibitor of apoptosisproteins family, is highly expressed in most human neoplasms, but its expression is very low or undetectable in terminally differentiated normal tissues. Survivin has been shown to inhibit cancer cell apoptosis and promote cell proliferation. The overexpression of survivin closely correlates with tumor progression and drug resistance. Because of its key role in tumor formation and maintenance, survivin is considered as an ideal target for anticancer treatment. However, the development of small-molecule survivin inhibitors has been challenging due to the requirement to disrupt the protein-protein interactions. Currently only a limited number of survivin inhibitors have been developed in recent years, and most of these inhibitors reduce survivin levels by interacting with other biomolecules instead of directly interacting with survivin protein. Despite these challenges, developing potent and selective small-molecule survivin inhibitors will be important in both basic science to better understand survivin biology and in translational research to develop potentially more effective, broad-spectrum anticancer agents. In this review, the functions of survivin and its role in cancer are summarized. Recent developments, challenges, and future direction of small-molecule survivin inhibitors are also discussed in detail.
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Affiliation(s)
| | - Wei Li
- Department of Pharmaceutical Sciences, University of Tennessee Health Science Center, Memphis, Tennessee 38163, United States.
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Hwang KE, Kwon SJ, Kim YS, Park DS, Kim BR, Yoon KH, Jeong ET, Kim HR. Effect of simvastatin on the resistance to EGFR tyrosine kinase inhibitors in a non-small cell lung cancer with the T790M mutation of EGFR. Exp Cell Res 2014; 323:288-96. [PMID: 24631288 DOI: 10.1016/j.yexcr.2014.02.026] [Citation(s) in RCA: 43] [Impact Index Per Article: 3.9] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/15/2013] [Revised: 02/21/2014] [Accepted: 02/22/2014] [Indexed: 10/25/2022]
Abstract
Although non-small cell lung cancer (NSCLC) tumors with activating mutations in the epidermal growth factor receptor (EGFR) are highly responsive to EGFR tyrosine kinase inhibitors (TKIs) including gefitinib and erlotinib, development of acquired resistance is almost inevitable. Statins show antitumor activity, but it is unknown whether they can reverse EGFR-TKIs resistance in NSCLC with the T790M mutation of EGFR. This study investigated overcoming resistance to EGFR-TKI using simvastatin. We demonstrated that addition of simvastatin to gefitinib enhanced caspase-dependent apoptosis in T790M mutant NSCLC cells. Simvastatin also strongly inhibited AKT activation, leading to suppression of β-catenin activity and the expression of its targets, survivin and cyclin D1. Both insulin treatment and AKT overexpression markedly increased p-β-catenin and survivin levels, even in the presence of gefitinib and simvastatin. However, inhibition of AKT by siRNA or LY294002 treatment decreased p-β-catenin and survivin levels. To determine the role of survivin in simvastatin-induced apoptosis of gefitinib-resistant NSCLC, we showed that the proportion of apoptotic cells following treatment with survivin siRNA and the gefitinib-simvastatin combination was greater than the theoretical additive effects, whereas survivin up-regulation could confer protection against gefitinib and simvastatin-induced apoptosis. Similar results were obtained in erlotinib and simvastatin-treated HCC827/ER cells. These findings suggest that survivin is a key molecule that renders T790M mutant NSCLC cells resistant to apoptosis induced by EGFR-TKIs and simvastatin. Overall, these data indicate that simvastatin may overcome EGFR-TKI resistance in T790M mutant NSCLCs via an AKT/β-catenin signaling-dependent down-regulation of survivin and apoptosis induction.
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Affiliation(s)
- Ki-Eun Hwang
- Department of Internal Medicine, Institute of Wonkwang Medical Science, Wonkwang University, School of Medicine, 344-2 shinyong-dong, Iksan, Jeonbuk 570-749, Republic of Korea
| | - Su-Jin Kwon
- Department of Internal Medicine, Institute of Wonkwang Medical Science, Wonkwang University, School of Medicine, 344-2 shinyong-dong, Iksan, Jeonbuk 570-749, Republic of Korea
| | - Young-Suk Kim
- Department of Internal Medicine, Institute of Wonkwang Medical Science, Wonkwang University, School of Medicine, 344-2 shinyong-dong, Iksan, Jeonbuk 570-749, Republic of Korea
| | - Do-Sim Park
- Department of Laboratory Medicine, Wonkwang University, School of Medicine, Iksan, Republic of Korea
| | - Byoung-Ryun Kim
- Department of Obstetrics & Gynecology, Wonkwang University, School of Medicine, Iksan, Republic of Korea
| | - Kwon-Ha Yoon
- Department of Radiology, Wonkwang University, School of Medicine, Iksan, Republic of Korea
| | - Eun-Taik Jeong
- Department of Internal Medicine, Institute of Wonkwang Medical Science, Wonkwang University, School of Medicine, 344-2 shinyong-dong, Iksan, Jeonbuk 570-749, Republic of Korea
| | - Hak-Ryul Kim
- Department of Internal Medicine, Institute of Wonkwang Medical Science, Wonkwang University, School of Medicine, 344-2 shinyong-dong, Iksan, Jeonbuk 570-749, Republic of Korea.
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Byeon HJ, Choi SH, Choi JS, Kim TH, Lee ES, Lee KC, Youn YS. Apoptotic activity and antitumor efficacy of PEGylated TNF-related apoptosis-inducing ligand (TRAIL) in a Mia Paca-2 cell-xenografted mouse model. Biomed Pharmacother 2013; 68:65-9. [PMID: 24268811 DOI: 10.1016/j.biopha.2013.10.010] [Citation(s) in RCA: 5] [Impact Index Per Article: 0.4] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/24/2013] [Accepted: 10/24/2013] [Indexed: 01/29/2023] Open
Abstract
The purpose of this study was to demonstrate the apoptotic activity and antitumor effect of PEGylated tumor necrosis factor-related apoptosis-inducing ligand (PEG-TRAIL) in pancreatic carcinoma Mia Paca-2 cells and in Mia Paca-2 cell-xenografted mice. PEG-TRAIL was prepared using mPEG-aldehyde (Mw 5 kDa). The apoptosis induced by PEG-TRAIL in Mia Paca-2 cells and in the tumors of Mia Paca-2 cell-xenografted mice was quantified by FACS analysis and using a TUNEL assay. Mia Paca-2 cell-xenografted BALB/c nu/nu mice were administered intratumoral injections of PEG-TRAIL (50 μg/mouse/injection) every 3 days from day 0 (~4 weeks after xenografting) to day 15. Tumor volumes were measured every 3 days from day 0 to day 27. PEG-TRAIL displayed obvious apoptotic activity in Mia Paca-2 cells; the FACS signal was shifted to the apoptotic area and the cells exhibited green fluorescence indicating apoptosis in the TUNEL assay. Furthermore, PEG-TRAIL was found to suppress tumors in Mia Paca-2 cell-xenografted mice (tumor volumes: 183.9±134.1 for PEG-TRAIL vs. 1827.3±264.5 mm(3) for saline control). In addition, in vivo TUNEL assays of tumor tissues showed that the antitumor effect of PEG-TRAIL was due apoptosis. Our findings provide clear in vivo evidence of the antitumor potential of PEG-TRAIL in a Mia Paca-2 cell-xenografted mouse model based of pancreatic cancer.
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Affiliation(s)
- Hyeong Jun Byeon
- School of Pharmacy, Sungkyunkwan University, 300, Cheoncheon-dong, Jangan-gu, 440-746 Suwon, Republic of Korea
| | - Seong Ho Choi
- School of Pharmacy, Sungkyunkwan University, 300, Cheoncheon-dong, Jangan-gu, 440-746 Suwon, Republic of Korea
| | - Ji Su Choi
- School of Pharmacy, Sungkyunkwan University, 300, Cheoncheon-dong, Jangan-gu, 440-746 Suwon, Republic of Korea
| | - Tae Hyung Kim
- School of Pharmacy, Sungkyunkwan University, 300, Cheoncheon-dong, Jangan-gu, 440-746 Suwon, Republic of Korea
| | - Eun Seong Lee
- Division of Biotechnology, The Catholic University of Korea, 43-1, Yeokgok 2-dong, Wonmi-gu, Bucheon-si, 420-743 Gyeonggi-do, Republic of Korea
| | - Kang Choon Lee
- School of Pharmacy, Sungkyunkwan University, 300, Cheoncheon-dong, Jangan-gu, 440-746 Suwon, Republic of Korea
| | - Yu Seok Youn
- School of Pharmacy, Sungkyunkwan University, 300, Cheoncheon-dong, Jangan-gu, 440-746 Suwon, Republic of Korea.
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Zhang G, He P, Gaedcke J, Ghadimi BM, Ried T, Yfantis HG, Lee DH, Hanna N, Alexander HR, Hussain SP. FOXL1, a novel candidate tumor suppressor, inhibits tumor aggressiveness and predicts outcome in human pancreatic cancer. Cancer Res 2013; 73:5416-25. [PMID: 23801748 DOI: 10.1158/0008-5472.can-13-0362] [Citation(s) in RCA: 34] [Impact Index Per Article: 2.8] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/08/2023]
Abstract
The forkhead box L1 (FOXL1) transcription factor regulates epithelial proliferation and development of gastrointestinal tract and has been implicated in gastrointestinal tumorigenesis in mouse models. However, the role of FOXL1 in pancreatic cancer development and progression remains to be elucidated. Here, we report that higher expression of FOXL1 is significantly associated with better clinical outcome in human pancreatic ductal adenocarcinoma (PDAC). A lower FOXL1 expression is correlated with metastasis and advanced pathologic stage of pancreatic cancer. Mechanistic analyses showed that overexpression of FOXL1 induces apoptosis and inhibits proliferation and invasion in pancreatic cancer cells, whereas silencing of FOXL1 by siRNA inhibits apoptosis and enhances tumor cell growth and invasion. Furthermore, FOXL1 overexpression significantly suppressed the growth of tumor xenografts in nude mice. FOXL1 promoted apoptosis partly through the induction of TNF-related apoptosis-inducing ligand (TRAIL) in pancreatic cancer cells. In addition, FOXL1 suppressed the transcription of zinc finger E-box-binding homeobox 1 (ZEB1), an activator of epithelial-mesenchymal transition, and the negative regulation of ZEB1 contributed to the inhibitory effect of FOXL1 on tumor cell invasion. Taken together, our findings suggest that FOXL1 expression is a candidate predictor of clinical outcome in patients with resected PDAC and it plays an inhibitory role in pancreatic tumor progression.
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Affiliation(s)
- Geng Zhang
- Pancreatic Cancer Unit, Laboratory of Human Carcinogenesis, Center for Cancer Research and Genetics Branch, National Cancer Institute, NIH, Bethesda, MD 20892, USA
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Hung CS, Liu HH, Huang MT, Cheng CW, Kuo LJ, Ho YS, Wu CH, Chen CM, Wei PL, Chang YJ. Knockdown survivin expression reduces the efficacy of curcumin treatment in hepatocellular carcinoma cells. Ann Surg Oncol 2012; 19:3547-3555. [PMID: 22711176 DOI: 10.1245/s10434-012-2393-4] [Citation(s) in RCA: 13] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/08/2011] [Indexed: 01/04/2025]
Abstract
BACKGROUND Survivin is a potential therapeutic target for cancer. Increased survivin expression promotes cell survival and therapeutic resistance. However, there is little information regarding whether the expression level of survivin affects curcumin treatment in hepatocellular carcinoma (HCC). METHODS Survivin expression was suppressed in HCC cells using a short interfering RNA (siRNA) technique. The anticancer effects of curcumin were examined using a biosensor system, MTT assay, TUNEL assay, and cell cycle analysis. RESULTS Curcumin resistance developed in cells with suppressed survivin, in contrast to the parental cells, as determined by survival assays. Cell cycle analysis and TUNEL assays revealed that the apoptotic cell population was increased in the scrambled-siRNA cells treated with curcumin compared with the survivin-siRNA cells. Suppression of survivin expression resulted in curcumin resistance via the modulation of Bcl-2 and Bax expression. CONCLUSIONS We conclude that the expression levels of survivin may mediate the therapeutic efficacy of curcumin in HCC cells.
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Affiliation(s)
- Chin-Sheng Hung
- Graduate Institute of Clinical Medicine, College of Medicine, Taipei Medical University, Taipei, Taiwan
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14
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Kita A, Nakahara T, Yamanaka K, Nakano K, Nakata M, Mori M, Kaneko N, Koutoku H, Izumisawa N, Sasamata M. Antitumor effects of YM155, a novel survivin suppressant, against human aggressive non-Hodgkin lymphoma. Leuk Res 2011; 35:787-92. [DOI: 10.1016/j.leukres.2010.11.016] [Citation(s) in RCA: 35] [Impact Index Per Article: 2.5] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/23/2010] [Revised: 10/16/2010] [Accepted: 11/22/2010] [Indexed: 10/18/2022]
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Prognostic significance of survivin polymorphisms on non-small cell lung cancer survival. J Thorac Oncol 2011; 5:1748-54. [PMID: 20881643 DOI: 10.1097/jto.0b013e3181f18cb9] [Citation(s) in RCA: 22] [Impact Index Per Article: 1.6] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/25/2022]
Abstract
INTRODUCTION Survivin is an apoptotic inhibitor, involves in regulation of apoptosis and cell cycle progression, and its polymorphisms may influence the development and progression of cancer. This study evaluated the impact of the survivin gene polymorphisms on survival of non-small cell lung cancer (NSCLC) patients. METHODS In this case-cohort follow-up study, a total of 568 NSCLC patients were investigated and 12 single nucleotide polymorphisms in survivin gene were genotyped by using the Illumina GoldenGate platform. RESULTS During the maximum of 72 months of follow-up, 314 (55.11%) deaths were observed. After adjusting for age, gender, smoking status, histology, stage, surgical operation, and chemotherapy or radiotherapy status, Cox hazard proportional model suggested that four single nucleotide polymorphisms had statistically significant impacts on NSCLC survival (rs3764383, AG/GG versus AA, hazard ratio [HR] = 0.78, 95% confidence interval [CI]: 0.62-0.99; rs8073069, GG versus CG/CC, HR = 1.76, 95% CI: 1.16-2.67; rs4789551, GG versus AG/AA, HR = 2.04, 95% CI: 1.08-3.86; rs1042489, GG versus AG/AA, HR = 1.37, 95% CI: 1.03-1.83). Further combined analysis showed that the high risk group (3-4 unfavorable loci) presented a 1.84-fold (95% CI: 1.22-2.77) increased risk compared with low risk group (0-2 unfavorable loci). Among 185 stage III to IV patients who received only chemotherapy, only the potentially functional rs8073069 still had a significantly increased risk on the prognosis of NSCLC (GG versus CG/CC, HR = 2.06, 95% CI: 1.10-3.87). CONCLUSIONS Our results suggest that polymorphisms in survivin may be a genetic modifier for NSCLC prognosis in this Chinese population.
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Fulda S. Targeting apoptosis signaling in pancreatic cancer. Cancers (Basel) 2011; 3:241-51. [PMID: 24212616 PMCID: PMC3756359 DOI: 10.3390/cancers3010241] [Citation(s) in RCA: 7] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/24/2010] [Revised: 01/05/2011] [Accepted: 01/06/2011] [Indexed: 12/14/2022] Open
Abstract
The ability to escape apoptosis or programmed cell death is a hallmark of human cancers, for example pancreatic cancer. This can promote tumorigenesis, since too little cell death by apoptosis disturbs tissue homeostasis. Additionally, defective apoptosis signaling is the underlying cause of failure to respond to current treatment approaches, since therapy-mediated antitumor activity requires the intactness of apoptosis signaling pathways in cancer cells. Thus, the elucidation of defects in the regulation of apoptosis in pancreatic carcinoma can result in the identification of novel targets for therapeutic interference and for exploitation for cancer drug discovery.
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Affiliation(s)
- Simone Fulda
- Institute for Experimental Cancer Research in Pediatrics, Goethe-University Frankfurt, Komturstr. 3a, 60528 Frankfurt, Germany.
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Samm N, Werner K, Rückert F, Saeger HD, Grützmann R, Pilarsky C. The role of apoptosis in the pathology of pancreatic cancer. Cancers (Basel) 2010; 3:1-16. [PMID: 24212603 PMCID: PMC3756346 DOI: 10.3390/cancers3010001] [Citation(s) in RCA: 17] [Impact Index Per Article: 1.1] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/11/2010] [Revised: 12/14/2010] [Accepted: 12/21/2010] [Indexed: 01/27/2023] Open
Abstract
Pancreatic cancer is a disease with high resistance to most common therapies and therefore has a poor prognosis, which is partly due to a lack of reaction to apoptotic stimuli. Signal transduction of such stimuli includes a death receptor-mediated extrinsic pathway as well as an intrinsic pathway linked to the mitochondria. Defects in apoptotic pathways and the deregulation of apoptotic proteins, such as Survivin, Bcl-2, Bcl-xL and Mcl-1, play decisive roles in the development of pancreatic cancer. Investigation of the molecular mechanism allowing tumors to resist apoptotic cell death would lead to an improved understanding of the physiology and the development of new molecular strategies in pancreatic cancer.
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Affiliation(s)
- Nicole Samm
- Department of Visceral-, Thoracic-and Vascular-Surgery, University Hospital Dresden, Dresden, Germany; E-Mails: (N.S.); (K.W.); (F.R.); (H.D.S.); (R.G.)
| | - Kristin Werner
- Department of Visceral-, Thoracic-and Vascular-Surgery, University Hospital Dresden, Dresden, Germany; E-Mails: (N.S.); (K.W.); (F.R.); (H.D.S.); (R.G.)
| | - Felix Rückert
- Department of Visceral-, Thoracic-and Vascular-Surgery, University Hospital Dresden, Dresden, Germany; E-Mails: (N.S.); (K.W.); (F.R.); (H.D.S.); (R.G.)
| | - Hans Detlev Saeger
- Department of Visceral-, Thoracic-and Vascular-Surgery, University Hospital Dresden, Dresden, Germany; E-Mails: (N.S.); (K.W.); (F.R.); (H.D.S.); (R.G.)
| | - Robert Grützmann
- Department of Visceral-, Thoracic-and Vascular-Surgery, University Hospital Dresden, Dresden, Germany; E-Mails: (N.S.); (K.W.); (F.R.); (H.D.S.); (R.G.)
| | - Christian Pilarsky
- Department of Visceral-, Thoracic-and Vascular-Surgery, University Hospital Dresden, Dresden, Germany; E-Mails: (N.S.); (K.W.); (F.R.); (H.D.S.); (R.G.)
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Hwang KE, Na KS, Park DS, Choi KH, Kim BR, Shim H, Jeong ET, Kim HR. Apoptotic induction by simvastatin in human lung cancer A549 cells via Akt signaling dependent down-regulation of survivin. Invest New Drugs 2010; 29:945-52. [PMID: 20464445 DOI: 10.1007/s10637-010-9450-2] [Citation(s) in RCA: 50] [Impact Index Per Article: 3.3] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/25/2010] [Accepted: 05/02/2010] [Indexed: 02/06/2023]
Abstract
Statins, HMG-CoA reductase inhibitors have been studied for their antiproliferative and proapototic effects. Recently, statin-induced apoptosis has been associated with down-regulation of survivin expression in cancer cells. However, the mechanism of deregulated survivin by simvastatin on lung cancer is still unclear. Herein, we demonstrated that simvastatin induced caspase-dependent apoptosis in A549 lung cancer cells. Simvastatin also resulted in a decrease in the expression of phosphorylated Akt. In addition, simvastatin effectively down-regulated survivin mRNA and protein, but not cIAP-1 and cIAP-2. The combination of simvastatin and 10 μM LY294002 (non-toxic dose) augmented apoptosis significantly, as evidenced by cleavage of PARP. The immunoreactive band of survivin was markedly decreased in cells treated with 50 μM LY294002 (toxic dose) as well as by the combination of simvastatin and 10 μM LY294002. Moreover, survivin down-regulation by RNA interference induced apoptosis accompanied by an increase in hypodiploid DNA content. Taken together, these data suggest that the anti-cancer effect of simvastatin via induction of apoptosis is related to Akt signaling dependent down-regulation of survivin in lung cancer A549 cells.
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Affiliation(s)
- Ki-Eun Hwang
- Department of Internal Medicine, Institute of Wonkwang Medical Science, School of Medicine, Wonkwang University, 344-2 shinyong-dong, Iksan, Jeonbuk 570-749, South Korea
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Kang S, Park SY, Lee HJ, Yoo YH. TRAIL upregulates decoy receptor 1 and mediates resistance to apoptosis in insulin-secreting INS-1 cells. Biochem Biophys Res Commun 2010; 396:731-5. [PMID: 20451496 DOI: 10.1016/j.bbrc.2010.05.004] [Citation(s) in RCA: 14] [Impact Index Per Article: 0.9] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/27/2010] [Accepted: 05/01/2010] [Indexed: 11/16/2022]
Abstract
TRAIL/Apo2L (tumor necrosis factor-related apoptosis-inducing ligand) is a multifunctional protein regulating the homeostasis of the immune system, infection, autoimmune diseases, and apoptosis. In particular, the potential role of TRAIL in type 1 diabetes (T1D) has been studied by several research groups. A previous study found that TRAIL did not have significant cytotoxic effects on the insulin-secreting pancreatic beta cell line, INS-1. However, the mechanism was not clear. Here we demonstrate that INS-1 cells are resistant to TRAIL-induced apoptosis and show alteration in the expression of death and decoy receptors upon TRAIL treatment. To compare TRAIL-resistant INS-1 cells with TRAIL-sensitive cells, we utilized U87MG cells, which are known to be TRAIL-sensitive. TRAIL treatment showed NF-kappaB translocation to the nucleus in TRAIL-resistant INS-1 cells, and TRAIL-induced NF-kappaB activation was preceded by IkappaBalpha degradation. A pharmacological inhibitor of NF-kappaB, Bay 11-7082, blocked TRAIL-induced NF-kappaB translocation to the nucleus and IkappaBalpha degradation. Four related receptors bind TRAIL: two death receptors (DR4 and DR5) that promote apoptosis, and two decoy receptors (DcR1 and DcR2) that act as dominant-negative inhibitors of TRAIL-mediated apoptosis. In the present study, TRAIL treatment in INS-1 cells upregulated DcR1 and downregulated DR5 without altering the expression of DcR2 and DR4. The resistance to apoptosis in INS-1 cells might therefore, be a consequence of DcR1 upregulation and DR5 downregulation, and the transcription factor, NF-kappaB, could regulate the sensitivity of cells to TRAIL by controlling the ratio of decoy to death receptors. Thus, TRAIL may play an important role in the survival of pancreatic beta cells by regulating receptor expression in an NF-kappaB-dependent manner.
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Affiliation(s)
- Soojeong Kang
- Department of Pharmacology, Dong-A University College of Medicine, Medical Science Research Center, Busan 602-714, Republic of Korea
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20
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Prognostic significance of tumour necrosis factor-related apoptosis-inducing ligand (TRAIL) receptor expression in patients with breast cancer. J Mol Med (Berl) 2009; 87:995-1007. [PMID: 19680616 DOI: 10.1007/s00109-009-0510-z] [Citation(s) in RCA: 65] [Impact Index Per Article: 4.1] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/14/2007] [Revised: 06/24/2009] [Accepted: 07/17/2009] [Indexed: 01/02/2023]
Abstract
TNF-related apoptosis-inducing ligand (TRAIL) induces apoptosis upon binding to TRAIL receptors 1 and 2 (TRAIL-R1/DR4 and TRAIL-R2/DR5). TRAIL-R3 (DcR1) and TRAIL-R4 (DcR2) have no or only a truncated cytoplasmic death domain. Consequently, they cannot induce apoptosis and instead have been proposed to inhibit apoptosis induction by TRAIL. Agonists for the apoptosis-inducing TRAIL-R1 and TRAIL-R2 are currently tested in clinical trials. To determine the expression pattern of all surface-bound TRAIL receptors and their prognostic clinical value, we investigated tumour samples of 311 patients with breast cancer by immunohistochemistry. TRAIL receptor expression profiles were correlated with clinico-pathological data, disease-free survival and overall survival. TRAIL-R1 was more strongly expressed in better differentiated tumours, and correlated positively with surrogate markers of a better prognosis (hormone receptor status, Bcl-2, negative nodal status), but negatively with the expression of Her2/neu and the proliferation marker Ki67. In contrast, TRAIL-R2 and TRAIL-R4 expression correlated with higher tumour grades, higher Ki67 index, higher Her2/neu expression and a positive nodal status at the time of diagnosis, but with lower expression of Bcl-2. Thus, the TRAIL receptor expression pattern was predictive of nodal status. Patients with grade 1 and 2 tumours, who had TRAIL-R2 but no TRAIL-R1, showed a positive lymph node status in 47% of the cases. Vice versa, only 19% had a positive nodal status with high TRAIL-R1 but low TRAIL-R2. Most strikingly, TRAIL-R4 and -R2 expression negatively correlated with overall survival of breast cancer patients. Although TRAIL-R2 correlated with more aggressive tumour behaviour, mammary carcinoma could be sensitised to TRAIL-R2-induced apoptosis, suggesting that TRAIL-R2 might therefore be used to therapeutically target such tumours. Hence, determination of the TRAIL receptor expression profile may aid in defining which breast cancer patients have a higher risk of lymph node metastasis and worse overall survival and on the other hand will help to guide TRAIL-based tumour therapy.
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Abstract
Resistance to apoptosis (programmed cell death) is a characteristic feature of human malignancies including pancreatic cancer, which is one of the leading causes of cancer deaths in the western world. Defects in this intrinsic cell death program can contribute to the multistep process of tumorigenesis, because too little cell death can disturb tissue homeostasis. Further, blockade of apoptosis pathways can cause treatment failure, because intact apoptosis signalling cascades largely mediate therapy-induced cytotoxicity. The elucidation of apoptosis pathways in pancreatic carcinoma over the last decade has resulted in the identification of various molecular defects. How apoptosis pathways can be exploited for the treatment of pancreatic cancer will be discussed in this review.
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Affiliation(s)
- Simone Fulda
- University Children's Hospital, Eythstr., Ulm, Germany.
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22
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Sanlioglu AD, Dirice E, Elpek O, Korcum AF, Ozdogan M, Suleymanlar I, Balci MK, Griffith TS, Sanlioglu S. High TRAIL death receptor 4 and decoy receptor 2 expression correlates with significant cell death in pancreatic ductal adenocarcinoma patients. Pancreas 2009; 38:154-60. [PMID: 18981952 DOI: 10.1097/mpa.0b013e31818db9e3] [Citation(s) in RCA: 25] [Impact Index Per Article: 1.6] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 02/05/2023]
Abstract
OBJECTIVES The importance of tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) and TRAIL receptor expression in pancreatic carcinoma development is not known. To reveal the putative connection of TRAIL and TRAIL receptor expression profile to this process, we analyzed and compared the expression profile of TRAIL and its receptors in pancreatic tissues of both noncancer patients and patients with pancreatic ductal adenocarcinoma (PDAC). METHODS Thirty-one noncancer patients and 34 PDAC patients were included in the study. TRAIL and TRAIL receptor expression profiles were determined by immunohistochemistry. Annexin V binding revealed the apoptotic index in pancreas. Lastly, the tumor grade, tumor stage, tumor diameter, perineural invasion, and number of lymph node metastasis were used for comparison purposes. RESULTS TRAIL decoy receptor 2 (DcR2) and death receptor 4 expression were up-regulated in PDAC patients compared with noncancer patients, and the ductal cells of PDAC patients displayed significant levels of apoptosis. In addition, acinar cells from PDAC patients had higher DcR2 expression but lower death receptor 4 expression. Increased DcR2 expression was also observed in Langerhans islets of PDAC patients. CONCLUSIONS Differential alteration of TRAIL and TRAIL receptor expression profiles in PDAC patients suggest that the TRAIL/TRAIL receptor system may play a pivotal role during pancreatic carcinoma development.
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Affiliation(s)
- Ahter Dilsad Sanlioglu
- Department of Medical Biology and Genetics, Faculty of Medicine, Human Gene Therapy Unit, Akdeniz University, Antalya, Turkey.
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Mita AC, Mita MM, Nawrocki ST, Giles FJ. Survivin: key regulator of mitosis and apoptosis and novel target for cancer therapeutics. Clin Cancer Res 2008; 14:5000-5. [PMID: 18698017 DOI: 10.1158/1078-0432.ccr-08-0746] [Citation(s) in RCA: 579] [Impact Index Per Article: 34.1] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/17/2022]
Abstract
Survivin, a member of the family of inhibitor of apoptosis proteins, functions as a key regulator of mitosis and programmed cell death. Initially, survivin was described as an inhibitor of caspase-9. However, over the last years, research studies have shown that the role of survivin in cancer pathogenesis is not limited to apoptosis inhibition but also involves the regulation of the mitotic spindle checkpoint and the promotion of angiogenesis and chemoresistance. Survivin gene expression is transcriptionally repressed by wild-type p53 and can be deregulated in cancer by several mechanisms, including gene amplification, hypomethylation, increased promoter activity, and loss of p53 function. This article reviews the multiple functions of survivin in the regulation of apoptosis, the promotion of tumorigenesis, and the development of survivin inhibitors as a novel anticancer therapeutic strategy.
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Affiliation(s)
- Alain C Mita
- Institute for Drug Development, Cancer Therapy and Research Center at the University of Texas Health Science Center, 7979 Wurzbach Road, San Antonio, TX 78229, USA.
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Sanlioglu AD, Griffith TS, Omer A, Dirice E, Sari R, Altunbas HA, Balci MK, Sanlioglu S. Molecular mechanisms of death ligand-mediated immune modulation: a gene therapy model to prolong islet survival in type 1 diabetes. J Cell Biochem 2008; 104:710-20. [PMID: 18247339 DOI: 10.1002/jcb.21677] [Citation(s) in RCA: 24] [Impact Index Per Article: 1.4] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/12/2022]
Abstract
Type 1 diabetes results from the T cell-mediated destruction of pancreatic beta cells. Islet transplantation has recently become a potential therapeutic approach for patients with type 1 diabetes. However, islet-graft failure appears to be a challenging issue to overcome. Thus, complementary gene therapy strategies are needed to improve the islet-graft survival following transplantation. Immune modulation through gene therapy represents a novel way of attacking cytotoxic T cells targeting pancreatic islets. Various death ligands of the TNF family such as FasL, TNF, and TNF-Related Apoptosis-Inducing Ligand (TRAIL) have been studied for this purpose. The over-expression of TNF or FasL in pancreatic islets exacerbates the onset of type 1 diabetes generating lymphocyte infiltrates responsible for the inflammation. Conversely, the lack of TRAIL expression results in higher degree of islet inflammation in the pancreas. In addition, blocking of TRAIL function using soluble TRAIL receptors facilitates the onset of diabetes. These results suggested that contrary to what was observed with TNF or FasL, adenovirus mediated TRAIL gene delivery into pancreatic islets is expected to be therapeutically beneficial in the setting of experimental models of type 1 diabetes. In conclusion; this study mainly reveals the fundamental principles of death ligand-mediated immune evasion in diabetes mellitus.
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Affiliation(s)
- Ahter Dilsad Sanlioglu
- Human Gene Therapy Unit and the Department of Medical Biology and Genetics, Akdeniz University, Faculty of Medicine, 07070 Antalya, Turkey
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25
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Hamacher R, Schmid RM, Saur D, Schneider G. Apoptotic pathways in pancreatic ductal adenocarcinoma. Mol Cancer 2008; 7:64. [PMID: 18652674 PMCID: PMC2515336 DOI: 10.1186/1476-4598-7-64] [Citation(s) in RCA: 90] [Impact Index Per Article: 5.3] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/21/2008] [Accepted: 07/24/2008] [Indexed: 02/08/2023] Open
Abstract
Pancreatic ductal adenocarcinoma (PDAC) is one of the most common causes of cancer related death. Despite the advances in understanding of the molecular pathogenesis, pancreatic cancer remains a major unsolved health problem. Overall, the 5-year survival rate is less than 5% demonstrating the insufficiency of current therapies. Most cytotoxic therapies induce apoptosis and PDAC cells have evolved a plethora of molecular mechanisms to assure survival. We will present anti-apoptotic strategies working at the level of the death receptors, the mitochondria or involving the caspase inhibitors of the IAP family. Furthermore, the survival function of the phosphotidylinositol-3' kinase (PI3K)/AKT- and NF-kappaB-pathways are illustrated. A detailed molecular knowledge of the anti-apoptotic mechanisms of PDAC cells will help to improve therapies for this dismal disease and therapeutic strategies targeting the programmed cell death machinery are in early preclinical and clinical development.
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Affiliation(s)
- Rainer Hamacher
- II. Medizinische Klinik, Klinikum rechts der Isar, Technische Universität München, München, Germany
| | - Roland M Schmid
- II. Medizinische Klinik, Klinikum rechts der Isar, Technische Universität München, München, Germany
| | - Dieter Saur
- II. Medizinische Klinik, Klinikum rechts der Isar, Technische Universität München, München, Germany
| | - Günter Schneider
- II. Medizinische Klinik, Klinikum rechts der Isar, Technische Universität München, München, Germany
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Sanlioglu AD, Dirice E, Elpek O, Korcum AF, Balci MK, Omer A, Griffith TS, Sanlioglu S. High levels of endogenous tumor necrosis factor-related apoptosis-inducing ligand expression correlate with increased cell death in human pancreas. Pancreas 2008; 36:385-93. [PMID: 18437085 DOI: 10.1097/mpa.0b013e318158a4e5] [Citation(s) in RCA: 25] [Impact Index Per Article: 1.5] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 02/05/2023]
Abstract
OBJECTIVES Type 1 diabetes (T1D) has been characterized by the T cell-mediated destruction of pancreatic beta cells. Although various members of the tumor necrosis factor (TNF) family, such as Fas ligand or TNF, have recently been implicated in the development of T1D, the lack of TNF-related apoptosis-inducing ligand (TRAIL) expression or function facilitates the onset of T1D. Thus, the goal of the present study was to investigate the expression profiles of TRAIL and its receptors in human pancreas. METHODS Pancreata of 31 patients were analyzed by immunohistochemistry using antibodies developed against TRAIL and its receptors. Apoptosis was confirmed by Annexin V-fluorescein isothiocyanate binding and terminal deoxynucleotidyl transferase-mediated 2'-deoxyuridine 5'-triphosphate nick end labeling assays. RESULTS Acinar cells displayed high levels of TRAIL and death receptor 4, but only low levels of death receptor 5. In contrast, only TRAIL and TRAIL decoy receptors (DcR1, DcR2) were detected in ductal cells. Similarly, Langerhans islets expressed only TRAIL and TRAIL decoy receptor. High levels of TRAIL expression in pancreas correlated with increased number of apoptotic cells. CONCLUSIONS Although the expression of TRAIL decoy receptors might be necessary for defense from TRAIL-induced apoptosis, high levels of TRAIL may provide protection for Langerhans islets from the immunological attack of cytotoxic T cells.
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Affiliation(s)
- Ahter Dilsad Sanlioglu
- Human Gene Therapy Unit, Departments of Medical Biology and Genetics, Akdeniz University, Faculty of Medicine, Antalya, Turkey.
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Derosier LC, Vickers SM, Zinn KR, Huang Z, Wang W, Grizzle WE, Sellers J, Stockard CR, Zhou T, Oliver PG, Arnoletti P, Lobuglio AF, Buchsbaum DJ. TRA-8 anti-DR5 monoclonal antibody and gemcitabine induce apoptosis and inhibit radiologically validated orthotopic pancreatic tumor growth. Mol Cancer Ther 2008; 6:3198-207. [PMID: 18089714 DOI: 10.1158/1535-7163.mct-07-0299] [Citation(s) in RCA: 38] [Impact Index Per Article: 2.2] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/16/2022]
Abstract
PURPOSE To evaluate agonistic TRA-8 monoclonal antibody to human death receptor 5 (DR5) and gemcitabine in vitro and in an orthotopic pancreatic cancer model. EXPERIMENTAL DESIGN Pancreatic cancer cell lines were screened for DR5 expression, cytotoxicity, and apoptosis induced by TRA-8, gemcitabine, or gemcitabine and TRA-8. An orthotopic model of pancreatic cancer was established in severe combined immunodeficient mice. Mice were treated with TRA-8, gemcitabine, or a combination for one or two cycles of therapy. Tumor growth (ultrasound) and survival were analyzed. RESULTS All five pancreatic cancer cell lines showed DR5 protein expression and varying sensitivity to TRA-8-mediated cytotoxicity. MIA PaCa-2 cells were very sensitive to TRA-8, moderately resistant to gemcitabine, with additive cytotoxicity to the combination. S2-VP10 cells were resistant to TRA-8 and sensitive to gemcitabine with synergistic sensitivity to the combination. Combination treatment in vitro produced enhanced caspase-3 and caspase-8 activation. A single cycle of therapy produced comparable efficacy for single-agent TRA-8 and the combination of TRA-8 and gemcitabine, with significant reduction in tumor size and prolonged survival compared with gemcitabine alone or control animals. With two cycles of therapy, TRA-8 and combination therapy produced enhanced inhibition of tumor growth compared with single-agent gemcitabine or untreated animals. However, the combination regimen showed enhanced survival as compared with single-agent TRA-8. CONCLUSIONS Pancreatic cancer cell lines express varying levels of DR5 and differ in their sensitivity to TRA-8 and gemcitabine-induced cytotoxicity. TRA-8 with two cycles of gemcitabine therapy produced the best overall survival.
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Affiliation(s)
- Leo Christopher Derosier
- Departments of Surgery, University of Alabama at Birmingham, 1530 3rd Avenue South, Wallace Tumor Institute 674, Birmingham, AL 35294-6832, USA
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DeRosier LC, Buchsbaum DJ, Oliver PG, Huang ZQ, Sellers JC, Grizzle WE, Wang W, Zhou T, Zinn KR, Long JW, Vickers SM. Combination treatment with TRA-8 anti death receptor 5 antibody and CPT-11 induces tumor regression in an orthotopic model of pancreatic cancer. Clin Cancer Res 2007; 13:5535s-5543s. [PMID: 17875786 PMCID: PMC3045836 DOI: 10.1158/1078-0432.ccr-07-1075] [Citation(s) in RCA: 36] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/26/2023]
Abstract
PURPOSE Evaluate the response of human pancreatic cancer cell lines and orthotopic tumors to TRA-8, an agonistic antibody to death receptor 5, in combination with irinotecan (CPT-11). EXPERIMENTAL DESIGN MIA PaCa-2 and S2VP10 cells were treated with TRA-8 and/or CPT 11. Cell viability was determined by ATP assay. JC-1 mitochondrial depolarization and Annexin V assays confirmed cell death by apoptosis. Immunoblotting was used to evaluate protein changes. MIA PaCa-2 cells were injected into the pancreas of severe combined immunodeficient mice. Mice underwent abdominal ultrasound to quantitate tumor size before and after treatment with twice weekly injections of 200 microg TRA-8 and/or 25 mg/kg CPT-11 for one or two treatment cycles, each lasting 2 weeks. RESULTS MIA PaCa-2 cells were more sensitive to TRA-8 and showed additive cytotoxicity, whereas S2VP10 cells showed synergistic cytotoxicity when treated with TRA-8 and CPT-11. Cell death occurred via apoptosis with increased cleavage of caspase-3, caspase-8, and caspase-9 and proapoptotic proteins Bid and poly(ADP)ribose polymerase after combination treatment compared with either agent alone. XIAP and Bcl-XL inhibitors of apoptosis were down-regulated. After a single cycle of in vivo combination therapy, tumor sizes had diminished significantly (P<0.001) at 8 days posttreatment compared with no treatment, CPT-11, and TRA-8; and there was a 50-day increase in survival with combination treatment over untreated controls (P=0.0002), 30 days over TRA-8, and a 36-day increase over CPT-11 monotherapy (P=0.0003). With two cycles of TRA-8/CPT-11 treatment, mean survival time increased significantly (P<0.001) to 169 days versus untreated controls, TRA-8 or CPT-11 (76, 121, or 108 days, respectively). CONCLUSIONS Combination TRA-8 and CPT-11 therapy produced enhanced cytotoxicity and survival in the MIA PaCa-2 orthotopic model of pancreatic cancer.
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Affiliation(s)
| | - Donald J. Buchsbaum
- Department of Surgery, University of Alabama at Birmingham, Birmingham, Alabama
- Department of Radiation Oncology, University of Alabama at Birmingham, Birmingham, Alabama
| | - Patsy G. Oliver
- Department of Radiation Oncology, University of Alabama at Birmingham, Birmingham, Alabama
| | - Zhi-Qiang Huang
- Department of Surgery, University of Alabama at Birmingham, Birmingham, Alabama
| | - Jeffrey C. Sellers
- Department of Radiation Oncology, University of Alabama at Birmingham, Birmingham, Alabama
| | - William E. Grizzle
- Department of Pathology, University of Alabama at Birmingham, Birmingham, Alabama
| | - Wenquan Wang
- Department of Medicine, University of Alabama at Birmingham, Birmingham, Alabama
| | - Tong Zhou
- Department of Medicine, University of Alabama at Birmingham, Birmingham, Alabama
| | - Kurt R. Zinn
- Department of Medicine, University of Alabama at Birmingham, Birmingham, Alabama
| | - Joshua W. Long
- Department of Surgery, University of Alabama at Birmingham, Birmingham, Alabama
| | - Selwyn M. Vickers
- Department of Surgery, University of Alabama at Birmingham, Birmingham, Alabama
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Tekmen I, Ozyurt D, Pekçetin C, Buldan Z. The effect of TRAIL molecule on cell viability in in vitro beta cell culture. Acta Diabetol 2007; 44:60-4. [PMID: 17530468 DOI: 10.1007/s00592-007-0243-9] [Citation(s) in RCA: 3] [Impact Index Per Article: 0.2] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 07/30/2005] [Accepted: 02/19/2007] [Indexed: 10/23/2022]
Abstract
Insulin-dependent diabetes mellitus (IDDM) is an organ-specific autoimmune disorder triggered by autoreactive T cells directed to pancreas beta-cell antigens. In this disorder, more than 90% of beta cells are destroyed. Cell death may be mediated via soluble or membrane-bound cell death ligands. One of these ligands may be tumour necrosis factor (TNF)-related apoptosis-inducing ligand (TRAIL), a member of the TNF-alpha superfamily. In the present study, we examined whether TRAIL had cytotoxic effects on adult rat pancreas beta cell cultures and INS1-E rat insulinoma cell line cultures or not. In this study, cell destruction models were built with TRAIL concentrations of 10, 100 and 1000 ng. 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) test was used for evaluating cell viability. It was detected that cell cultures with TRAIL added showed no differences statistically when compared with control cultures containing no toxic additions. These results showed that TRAIL did not have significant cytotoxic effects on pancreas beta cell culture and INS-1E rat insulinoma cell line cultures. Detection of the expression of TRAIL receptors and natural apoptosis inhibitor proteins will be favourable to investigate the resistance mechanisms to TRAIL-induced cell death in this cell culture system.
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Affiliation(s)
- I Tekmen
- Department of Histology and Embryology, Dokuz Eylul University Medical Faculty, 35340, Inciralti, Izmir, Turkey.
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Vogler M, Dürr K, Jovanovic M, Debatin KM, Fulda S. Regulation of TRAIL-induced apoptosis by XIAP in pancreatic carcinoma cells. Oncogene 2006; 26:248-57. [PMID: 16832350 DOI: 10.1038/sj.onc.1209776] [Citation(s) in RCA: 90] [Impact Index Per Article: 4.7] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/18/2023]
Abstract
Tumor necrosis factor (TNF)-related apoptosis-inducing ligand (TRAIL) is a promising candidate for cancer therapy because of its relative tumor selectivity. However, many cancers including pancreatic cancer remain resistant towards TRAIL. To develop TRAIL for cancer therapy of pancreatic carcinoma, it will therefore be pivotal to elucidate the molecular mechanisms of TRAIL resistance. Here, we identify X-linked inhibitor of apoptosis (XIAP) as a regulator of TRAIL sensitivity in pancreatic carcinoma cells. Full activation of effector caspases, loss of mitochondrial membrane potential and cytochrome c release following TRAIL treatment were markedly impaired in pancreatic carcinoma cell lines, which poorly responded to TRAIL (PaTuII, PancTu1, ASPC1, DanG), compared to TRAIL-sensitive Colo357 pancreatic carcinoma cells. Stable downregulation of XIAP by RNA interference significantly reduced survival and enhanced TRAIL-induced apoptosis in pancreatic carcinoma cells. Also, downregulation of XIAP significantly increased CD95-induced cell death. Importantly, knockdown of XIAP strongly inhibited clonogenicity of pancreatic cancer cells treated with TRAIL indicating that XIAP promotes clonogenic survival of pancreatic carcinoma cells. Thus, our findings for the first time indicate that targeting XIAP represents a promising strategy to enhance the antitumor activity of TRAIL in pancreatic cancer, which has important clinical implications.
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Affiliation(s)
- M Vogler
- University Children's Hospital, Ulm, Germany
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Tafuku S, Matsuda T, Kawakami H, Tomita M, Yagita H, Mori N. Potential mechanism of resistance to TRAIL-induced apoptosis in Burkitt's lymphoma. Eur J Haematol 2006; 76:64-74. [PMID: 16343273 DOI: 10.1111/j.0902-4441.0000.t01-1-ejh2345.x] [Citation(s) in RCA: 18] [Impact Index Per Article: 0.9] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/30/2022]
Abstract
OBJECTIVES Members of the tumor necrosis factor family are potent inducers of apoptosis in sensitive cells and may be suitable for novel anti-cancer therapies aimed at inducing apoptosis via the activation of receptors with the death domain on malignant cells. We characterized the sensitivity of Burkitt's lymphoma (BL) cell lines to tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) and anti-Fas agonist, and investigated the mechanism of resistance of BL cell lines to TRAIL and Fas apoptotic pathways. METHODS Epstein-Barr virus (EBV) status in BL cell lines was determined by PCR. The extent of apoptosis following exposure to TRAIL and anti-Fas agonist was measured by 7A6 antigen staining. Expression of TRAIL receptors and Fas was determined by flow cytometry and reverse transcriptase-PCR. Western blot analyses were used to determine the expression of proapoptotic and antiapoptotic proteins. NF-kappaB activity was evaluated by electrophoretic mobility shift assay. RESULTS The sensitivity of BL cell lines to anti-Fas agonist depended on the expression of Fas. In contrast, the expression of TRAIL receptors did not correlate with the sensitivity to TRAIL-induced apoptosis. Interestingly, EBV-infected BL cell lines which showed constitutive levels of NF-kappaB activation, were TRAIL-resistant. NF-kappaB inhibitors reversed the resistance to TRAIL-induced apoptosis. CONCLUSIONS Our results suggest that activation of NF-kappaB by EBV infection plays an important role in resistance of BL cell lines to TRAIL-induced apoptosis, and that NF-kappaB inhibitors may be useful adjuncts in clinical use of TRAIL against BL.
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Affiliation(s)
- Senji Tafuku
- Division of Molecular Virology and Oncology, Graduate School of Medicine, University of the Ryukyus, Nishihara, Okinawa, Japan
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Affiliation(s)
- Monica Mita
- Institute for Drug Development, Cancer Therapy and Research Center, University of Texas Health Science Center, San Antonio, USA
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Hylander BL, Pitoniak R, Penetrante RB, Gibbs JF, Oktay D, Cheng J, Repasky EA. The anti-tumor effect of Apo2L/TRAIL on patient pancreatic adenocarcinomas grown as xenografts in SCID mice. J Transl Med 2005; 3:22. [PMID: 15943879 PMCID: PMC1156958 DOI: 10.1186/1479-5876-3-22] [Citation(s) in RCA: 87] [Impact Index Per Article: 4.4] [Reference Citation Analysis] [Abstract] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/10/2005] [Accepted: 05/19/2005] [Indexed: 12/22/2022] Open
Abstract
Background Apo2L/TRAIL has considerable promise for cancer therapy based on the fact that this member of the tumor necrosis factor family induces apoptosis in the majority of malignant cells, while normal cells are more resistant. Furthermore, in many cells, when Apo2L/TRAIL is combined with chemotherapy, the effect is synergistic. The majority of this work has been carried out using cell lines. Therefore, investigation of how patient tumors respond to Apo2L/TRAIL can validate and/or complement information obtained from cell lines and prove valuable in the design of future clinical trials. Methods We have investigated the Apo2L/TRAIL sensitivity of patient derived pancreatic tumors using a patient tumor xenograft/ SCID mouse model. Mice bearing engrafted tumors were treated with Apo2L/TRAIL, gemcitabine or a combination of both therapies. Results Patient tumors grown as xenografts exhibited a spectrum of sensitivity to Apo2L/TRAIL. Both Apo2L/TRAIL sensitive and resistant pancreatic tumors were found, as well as tumors that showed heterogeneity of response. Changes in apoptotic signaling molecules in a sensitive tumor were analyzed by Western blot following Apo2L/TRAIL treatment; loss of procaspase 8, Bid and procaspase 3 was observed and correlated with inhibition of tumor growth. However, in a tumor that was highly resistant to killing by Apo2L/TRAIL, although there was a partial loss of procaspase 8 and Bid in response to Apo2L/TRAIL treatment, loss of procaspase 3 was negligible. This resistant tumor also expressed a high level of the anti-apoptotic molecule Bcl-XL that, in comparison, was not detected in a sensitive tumor. Importantly, in the majority of these tumors, addition of gemcitabine to Apo2L/TRAIL resulted in a greater anti-tumor effect than either therapy used alone. Conclusion These data suggest that in a clinical setting we will see heterogeneity in the response of patients' tumors to Apo2L/TRAIL, including tumors that are highly sensitive as well as those that are resistant. While much more work is needed to understand the molecular basis for this heterogeneity, it is very encouraging, that Apo2L/TRAIL in combination with gemcitabine increased therapeutic efficacy in almost every case and therefore may be a highly effective strategy for controlling human pancreatic cancer validating and expanding upon what has been reported for cell lines.
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Affiliation(s)
- Bonnie L Hylander
- Department of Immunology, Roswell Park Cancer Institute, Buffalo, NY, 14263, USA
| | - Rose Pitoniak
- Department of Immunology, Roswell Park Cancer Institute, Buffalo, NY, 14263, USA
| | | | - John F Gibbs
- Department of Surgery, Roswell Park Cancer Institute, Buffalo, NY, 14263, USA
| | - Dilek Oktay
- Department of Immunology, Roswell Park Cancer Institute, Buffalo, NY, 14263, USA
| | - Jinrong Cheng
- Department of Experimental Therapeutics, Roswell Park Cancer Institute, Buffalo, NY, 14263, USA
| | - Elizabeth A Repasky
- Department of Immunology, Roswell Park Cancer Institute, Buffalo, NY, 14263, USA
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Matsuda T, Almasan A, Tomita M, Tamaki K, Saito M, Tadano M, Yagita H, Ohta T, Mori N. Dengue virus-induced apoptosis in hepatic cells is partly mediated by Apo2 ligand/tumour necrosis factor-related apoptosis-inducing ligand. J Gen Virol 2005; 86:1055-1065. [PMID: 15784899 PMCID: PMC2917180 DOI: 10.1099/vir.0.80531-0] [Citation(s) in RCA: 41] [Impact Index Per Article: 2.1] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/24/2022] Open
Abstract
Although hepatic injury is reported in cases with dengue haemorrhagic fever and dengue shock syndrome, its mechanism remains poorly understood. Several findings suggest that dengue virus (DEN) induces apoptosis of hepatocytes in vivo. In this work, DEN type 2 (DEN-2) strain NGC was shown to induce apoptosis in the hepatic cell line HepG2, and infection of HepG2 cells was found to induce Apo2 ligand (Apo2L, also known as tumour necrosis factor-related apoptosis-inducing ligand or TRAIL) expression. Furthermore, Apo2L/TRAIL induced apoptosis in HepG2 cells, which expressed the Apo2L/TRAIL receptor DR5/TRAIL-R2 on their surface. Analysis of the Apo2L/TRAIL promoter revealed that this gene was activated by DEN-2 infection, whose responsive element was overlapping NF-kappaB- and Sp1-binding sites located at nt -75 to -65. The proteasome inhibitor N-acetyl-L-leucinyl-L-leucinyl-L-norleucinal (LLnL) inhibited Apo2L/TRAIL mRNA expression, and LLnL and anti-Apo2L/TRAIL antibody inhibited DEN-2-induced apoptosis. It was proposed that DEN infection promotes apoptosis partly through the induction of Apo2L/TRAIL expression.
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Affiliation(s)
- Takehiro Matsuda
- Division of Molecular Virology and Oncology, Graduate School of Medicine, University of the Ryukyus, Uehara 207, Nishihara, Okinawa 903-0215, Japan
- Division of Child Health and Welfare, Faculty of Medicine, University of the Ryukyus, Uehara 207, Nishihara, Okinawa 903-0215, Japan
| | - Alex Almasan
- Department of Cancer Biology, Lerner Research Institute, Cleveland Clinic Foundation, Cleveland, OH 44195, USA
| | - Mariko Tomita
- Division of Molecular Virology and Oncology, Graduate School of Medicine, University of the Ryukyus, Uehara 207, Nishihara, Okinawa 903-0215, Japan
| | - Kazumi Tamaki
- Division of Molecular Virology and Oncology, Graduate School of Medicine, University of the Ryukyus, Uehara 207, Nishihara, Okinawa 903-0215, Japan
| | - Mika Saito
- Division of Molecular Virology and Oncology, Graduate School of Medicine, University of the Ryukyus, Uehara 207, Nishihara, Okinawa 903-0215, Japan
| | - Masayuki Tadano
- Division of Molecular Virology and Oncology, Graduate School of Medicine, University of the Ryukyus, Uehara 207, Nishihara, Okinawa 903-0215, Japan
| | - Hideo Yagita
- Department of Immunology, Juntendo University School of Medicine, Hongo 2-1-1, Bunkyo-ku, Tokyo 113-8421, Japan
| | - Takao Ohta
- Division of Child Health and Welfare, Faculty of Medicine, University of the Ryukyus, Uehara 207, Nishihara, Okinawa 903-0215, Japan
| | - Naoki Mori
- Division of Molecular Virology and Oncology, Graduate School of Medicine, University of the Ryukyus, Uehara 207, Nishihara, Okinawa 903-0215, Japan
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Matsuda T, Almasan A, Tomita M, Uchihara JN, Masuda M, Ohshiro K, Takasu N, Yagita H, Ohta T, Mori N. Resistance to Apo2 ligand (Apo2L)/tumor necrosis factor-related apoptosis-inducing ligand (TRAIL)-mediated apoptosis and constitutive expression of Apo2L/TRAIL in human T-cell leukemia virus type 1-infected T-cell lines. J Virol 2005; 79:1367-78. [PMID: 15650163 PMCID: PMC544134 DOI: 10.1128/jvi.79.3.1367-1378.2005] [Citation(s) in RCA: 19] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/31/2023] Open
Abstract
Adult T-cell leukemia (ATL), a CD4+-T-cell malignancy caused by human T-cell leukemia virus type 1 (HTLV-1), is difficult to cure, and novel treatments are urgently needed. Apo2 ligand (Apo2L; also tumor necrosis factor-related apoptosis-inducing ligand [TRAIL]) has been implicated in antitumor therapy. We found that HTLV-1-infected T-cell lines and primary ATL cells were more resistant to Apo2L-induced apoptosis than uninfected cells. Interestingly, HTLV-1-infected T-cell lines and primary ATL cells constitutively expressed Apo2L mRNA. Inducible expression of the viral oncoprotein Tax in a T-cell line up-regulated Apo2L mRNA. Analysis of the Apo2L promoter revealed that this gene is activated by Tax via the activation of NF-kappaB. The sensitivity to Apo2L was not correlated with expression levels of Apo2L receptors, intracellular regulators of apoptosis (FLICE-inhibitory protein and active Akt). NF-kappaB plays a crucial role in the pathogenesis and survival of ATL cells. The resistance to Apo2L-induced apoptosis was reversed by N-acetyl-L-leucinyl-L-leucinyl-lLnorleucinal (LLnL), an NF-kappaB inhibitor. LLnL significantly induced the Apo2L receptors DR4 and DR5. Our results suggest that the constitutive activation of NF-kappaB is essential for Apo2L gene induction and protection against Apo2L-induced apoptosis and that suppression of NF-kappaB may be a useful adjunct in clinical use of Apo2L against ATL.
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Affiliation(s)
- Takehiro Matsuda
- Division of Molecular Virology and Oncology, Graduate School of Medicine, University of the Ryukyus, 207 Uehara, Nishihara, Okinawa 903-0215, Japan
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Farrow B, Thomas RP, Wang XF, Evers BM. Activation of conventional PKC isoforms increases expression of the pro-apoptotic protein Bad and TRAIL receptors. INTERNATIONAL JOURNAL OF GASTROINTESTINAL CANCER 2003; 32:63-72. [PMID: 12794242 DOI: 10.1385/ijgc:32:2-3:63] [Citation(s) in RCA: 13] [Impact Index Per Article: 0.6] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 12/14/2022]
Abstract
BACKGROUND Pancreatic cancer is a leading cause of cancer death worldwide; current treatment options have been ineffective in prolonging survival. Agents that target specific signaling pathways (e.g., protein kinase C [PKC]) may regulate apoptotic gene expression rendering resistant cancers sensitive to the effects of other chemotherapeutic drugs. The purpose of our study was to assess the effect of PKC stimulation on apoptotic gene expression in pancreatic cancer cells. METHODS The human pancreatic cancer cell line, PANC-1, was treated with PKC-stimulating agents, phorbol 12-myristate 13-acetate (PMA) or bryostatin-1, and analyzed for expression of apoptosis-related genes. RESULTS Both PMA and bryostatin-1 induced expression of the pro-apoptotic gene Bad in a dose dependent fashion. The expression of Bad was blocked by the PKC inhibitors GF109203x, Gö6983, and Ro-31-8220, suggesting a role for the conventional isoforms of PKC. In addition, treatment with the MEK inhibitors PD98059 or UO126 reduced PMA-mediated induction of Bad gene expression. PMA also increased the expression of TRAIL receptors DR4 and DR5; this expression was inhibited by the PKC inhibitors GF109203x, Gö6983, and Ro-31-8220 and the MEK inhibitor UO126, suggesting a role for conventional PKC isoforms and MEK in the regulation of TRAIL receptor expression. CONCLUSIONS PKC stimulation in PANC-1 cells increases expression of the pro-apoptotic gene Bad and the TRAIL receptors, DR4 and DR5, through both conventional PKC- and MEK-dependent pathways. Agents that stimulate PKC may sensitize pancreatic cancer cells to apoptosis and provide a potential adjuvant therapy for the treatment of chemoresistant pancreatic cancers.
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Affiliation(s)
- Buckminster Farrow
- Department of Surgery, The University of Texas Medical Branch, Galveston, TX 77555, USA
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Mi QS, Ly D, Lamhamedi-Cherradi SE, Salojin KV, Zhou L, Grattan M, Meagher C, Zucker P, Chen YH, Nagle J, Taub D, Delovitch TL. Blockade of tumor necrosis factor-related apoptosis-inducing ligand exacerbates type 1 diabetes in NOD mice. Diabetes 2003; 52:1967-75. [PMID: 12882912 DOI: 10.2337/diabetes.52.8.1967] [Citation(s) in RCA: 80] [Impact Index Per Article: 3.6] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 01/02/2023]
Abstract
Tumor necrosis factor (TNF)-related apoptosis-inducing ligand (TRAIL) is expressed in different tissues and cells, including pancreas and lymphocytes, and can induce apoptosis in various tumor cells but not in most normal cells. The specific roles of TRAIL in health and disease remain unclear. Here we show by cDNA array analyses that TRAIL gene expression is upregulated in pancreatic islets during the development of autoimmune type 1 diabetes in nonobese diabetic (NOD) mice and in Min6 islet beta-cells activated by TNF-alpha + interferon-gamma. However, stimulation of freshly isolated pancreatic islets or Min6 cells with TRAIL did not induce their apoptosis. TRAIL blockade exacerbates the onset of type 1 diabetes in NOD.Scid recipients of transferred diabetogenic T-cells and in cyclophosphamide-treated NOD mice. TRAIL inhibits the proliferation of NOD diabetogenic T-cells by suppressing interleukin (IL)-2 production and cell cycle progression, and this inhibition can be rescued in the presence of exogenous IL-2. cDNA array and Western blot analyses indicate that TRAIL upregulates the expression of the cdk inhibitor p27(kip1). Our data suggest that TRAIL is an important immune regulator of the development of type 1 diabetes.
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Affiliation(s)
- Qing-Sheng Mi
- Autoimmunity/Diabetes Group, the John P. Robarts Research Institute, London, Ontario, Canada
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Katz MH, Spivack DE, Takimoto S, Fang B, Burton DW, Moossa AR, Hoffman RM, Bouvet M. Gene therapy of pancreatic cancer with green fluorescent protein and tumor necrosis factor-related apoptosis-inducing ligand fusion gene expression driven by a human telomerase reverse transcriptase promoter. Ann Surg Oncol 2003; 10:762-72. [PMID: 12900367 DOI: 10.1245/aso.2003.01.021] [Citation(s) in RCA: 31] [Impact Index Per Article: 1.4] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/30/2022]
Abstract
BACKGROUND Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) induces apoptosis in malignant cells but not in normal cells. Ad/g-TRAIL, an adenoviral vector in which expression of green fluorescent protein (GFP) and TRAIL is driven by a human telomerase reverse transcriptase promoter, has shown promise as a targeted antitumor agent. METHODS To investigate the effects of TRAIL gene therapy on pancreatic cancer, BxPC-3, MIA-PaCa-2, Panc-1, and ASPC-1 cells were treated with Ad/g-TRAIL. Transfection and protein expression were determined by using immunoblotting and identification of GFP with fluorescent microscopy and flow cytometry. Cell viability was determined by proliferation assay. Cell-cycle analysis and quantification of caspase-3 were used to identify apoptosis. The in vivo efficacy of Ad/g-TRAIL was characterized in a novel red fluorescent protein murine model of MIA-PaCa-2 pancreatic cancer. RESULTS Cells treated with Ad/g-TRAIL expressed GFP and exhibited apoptotic morphology within 2 days of treatment. Treatment with this vector in vitro resulted in less cell viability, increased caspase-3 activity, and a greater apoptotic fraction than treatment with controls. In vivo, treatment with Ad/g-TRAIL significantly suppressed tumor growth. CONCLUSIONS TRAIL gene therapy induces apoptosis of pancreatic tumor cells both in vitro and in vivo and is a promising therapy in the treatment of pancreatic cancer.
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Affiliation(s)
- Matthew H Katz
- Department of Surgery, University of California at San Diego, San Diego, California 92161, USA
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Abstract
AIM: To observe the localization of TRAIL/TRAIR (DR4, DR5, DcR1, DcR2) in the fetal pancreas.
METHODS: Fetal pancreas of 32 wk of pregnancy were obtained from induced abortions, embedded in paraffin, and 4-μm sections were prepared. The localization of TRAIL/TRAILR in fetal pancreas was investigated by fluorescence immunohistochemical method combined with laser scanning confocal microscopy.
RESULTS: TRAIL immunoreactive cells were mainly located on the periphery of the pancreas islets. There were a few DcR1 and DcR2 positive cells whereas there were no immunoreactive cells of DR4 and DR5 in the pancreas islets. In the acini and the ducts of the exocrine pancreas there were no TRAIL/TRAILR immunoreactive cells.
CONCLUSION: This study not only describes the distribution of TRAIL/TRAILR in the fetal pancreas, but also provides a morphological basis for deducing the function of TRAIL/TRAILR in pancreas, suggesting that in normal pancreatic islets, the pancreatic cells are resistant towards apoptosis too.
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Affiliation(s)
- Li-Hua Chen
- Department of Immunology, the Fourth Military Medical University, 17 West Changle Road, Xi'an 710032, Shaanxi Province, China
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Thomas RP, Farrow BJ, Kim S, May MJ, Hellmich MR, Evers BM. Selective targeting of the nuclear factor-kappaB pathway enhances tumor necrosis factor-related apoptosis-inducing ligand-mediated pancreatic cancer cell death. Surgery 2002; 132:127-34. [PMID: 12219002 DOI: 10.1067/msy.2002.124930] [Citation(s) in RCA: 53] [Impact Index Per Article: 2.3] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/03/2023]
Abstract
BACKGROUND Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL), a member of the tumor necrosis factor family, selectively induces apoptosis in various cancer cells; however, certain cancers can evade TRAIL-mediated apoptosis. FLICE-like inhibitory protein (FLIP), an inhibitor of caspase-8, (also known as FLICE) is regulated by the transcription factor nuclear factor-kappaB (NF-kappaB) and can contribute to TRAIL resistance. The purpose of our study was to determine whether inhibition of NF-kappaB can enhance TRAIL-mediated pancreatic cancer cell death and decrease FLIP levels. METHODS The human pancreatic cancer cell lines MIA PaCa-2 and L3.6 were treated with TRAIL, NEMO-binding domain (NBD) peptide (a novel selective NF-kappaB inhibitor), or a combination of both. Cell viability and apoptosis were measured. Gel mobility shift assays were performed to assess NF-kappaB binding activity. Western blots were performed to assess FLIP levels after treatment with NBD or infection with an adenovirus encoding mutated IkappaBalpha. RESULTS The aggressive L3.6 cell line was resistant to TRAIL treatment, whereas MIA PaCa-2 cells were sensitive to TRAIL. The combination of TRAIL and NBD significantly decreased cell viability and increased apoptosis in L3.6 cells. Cellular levels of FLIP were decreased by inhibition of NF-kappaB (either by NBD treatment or mutant IkappaBalpha infection). CONCLUSIONS Our findings demonstrate resistance of the aggressive L3.6 pancreatic cell line to TRAIL treatment alone; inhibition of NF-kappaB by NBD increased TRAIL-mediated cell death and decreased FLIP protein levels. Novel agents that selectively target the NF-kappaB pathway may be useful adjuvant therapies for chemoresistant pancreatic cancers.
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Affiliation(s)
- Robert P Thomas
- Department of Surgery, The University of Texas Medical Branch, Galveston 77555, USA
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Kaneko K, Satoh K, Masamune A, Satoh A, Shimosegawa T. Expression of ROCK-1 in human pancreatic cancer: its down-regulation by morpholino oligo antisense can reduce the migration of pancreatic cancer cells in vitro. Pancreas 2002; 24:251-7. [PMID: 11893932 DOI: 10.1097/00006676-200204000-00007] [Citation(s) in RCA: 36] [Impact Index Per Article: 1.6] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 12/10/2022]
Abstract
INTRODUCTION Invasion and metastasis of cancer cells require cell motility and adhesion. The small GTPase Rho and one of its effector molecules ROCK regulate cytoskeleton and actomyosin contractility, and play a crucial role in cell adhesion and motility. Results of previous studies showed that the elevated activity of ROCK-1, one of the isomers of ROCK kinases, led to an increase in the activity of invasion and metastasis of cancer cell lines. AIM To investigate the importance of ROCK-1 in cancer invasion and metastasis. METHODOLOGY We investigated the expression of ROCK-1 in two cancer cell lines and 31 human pancreatic tissues (21 pancreatic cancers [PC] and 10 histologically normal tissues) by immunoblotting and immunohistochemistry. We also examined by haptotaxis assay whether the migratory activity of PC cells could be suppressed by treatment with the morpholino antisense oligonucleotide in vitro. RESULTS The expression of ROCK-1 was found in 18 of 21 PC tissues (85.7%), but not in normal pancreatic tissues by immunoblotting and immunohistochemistry. Antisense oligo against ROCK-1 significantly inhibited the haptotaxis of Panc-1 in a dose-dependent manner, compared with the control oligo. CONCLUSION These results suggest that ROCK-1 may contribute to pancreatic cancer cell invasion and/or metastasis by facilitating cancer cell migration.
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Affiliation(s)
- Kenzo Kaneko
- Department of Gastroenterology, Division of Internal Medicine, Tohoku University Graduate School of Medicine, Sendai, Miyagi, Japan.
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