ABSTRACT
STUDY QUESTION: Does the flavonoid naringenin inhibit proliferation of human endometriosis cells? SUMMARY ANSWER: Naringenin suppresses proliferation and increases apoptosis via depolarization of mitochondrial membrane potential and generation of reactive oxygen species (ROS) in human endometriosis cells. WHAT IS KNOWN ALREADY: For management of endometriosis, hormonal therapy is commonly used to decrease production of estrogens by the ovaries, but that has limitations including undesirable side effects with long-term therapies. To overcome these limitations, it is important to discover novel compounds which have no adverse effects, but inhibit expression of target molecules involved in the pathogenesis of endometriosis. STUDY DESIGN SIZE, DURATION: Well-established endometriosis cell lines (VK2/E6E7 and End1/E6E7) were purchased from the American Type Culture Collection. Effects of naringenin on VK2/E6E7 and End1/E6E7 cells were assessed in diverse assays in a dose- and time-dependent manner. PARTICIPANTS/MATERIALS, SETTING, METHODS: Effects of naringenin on viability, apoptosis (Annexin V expression, propidium iodide staining, TUNEL and invasion assays), mitochondria-mediated apoptosis, production of ROS and endoplasmic reticulum (ER) stress proteins of VK2/E6E7 and End1/E6E7 cells were determined. Signal transduction pathways in VK2/E6E7 and End1/E6E7 cells in response to naringenin were determined by western blot analyses. MAIN RESULTS AND THE ROLE OF CHANCE: In the present study, we demonstrated that naringenin suppressed proliferation and increased apoptosis through depolarization of mitochondrial membrane potential and inducing pro-apoptotic proteins, Bax and Bak, in both endometriosis cell lines. In addition, naringenin increased ROS, ER stress, through activation of eIF2α and IRE1α, GADD153 and GRP78 proteins in a dose-dependent manner. Furthermore, the induction of apoptosis by naringenin involved activation of MAPK and inactivation of PI3K pathways in VK2/E6E7 and End1/E6E7 cells. LIMITATIONS REASONS FOR CAUTION: Lack of in vivo animal studies is a major limitation of this research. Effectiveness of naringenin to induce apoptosis of human endometriosis cells requires further investigation. WIDER IMPLICATIONS OF THE FINDINGS: Our results suggest that naringenin is a promising therapeutic compound for treatment of endometriosis in women. STUDY FUNDING/COMPETING INTEREST(S): This work was supported by grants from the Korea Health Technology R&D Project through the Korea Health Industry Development Institute (KHIDI), funded by the Ministry of Health & Welfare, Republic of Korea (No. HI15C0810 awarded to G.S. and HI17C0929 awarded to W.L.). The authors declare that there are no conflicts of interest.
Subject(s)
Endoplasmic Reticulum Stress/drug effects , Estrogen Antagonists/pharmacology , Flavanones/pharmacology , Mitochondria/drug effects , Mitogen-Activated Protein Kinases/genetics , Stromal Cells/drug effects , Annexin A5/genetics , Annexin A5/metabolism , Apoptosis/drug effects , Cell Line , Dose-Response Relationship, Drug , Endometriosis/metabolism , Endometriosis/pathology , Endometrium/metabolism , Endometrium/pathology , Endoplasmic Reticulum Chaperone BiP , Endoribonucleases/genetics , Endoribonucleases/metabolism , Eukaryotic Initiation Factor-2/agonists , Eukaryotic Initiation Factor-2/genetics , Eukaryotic Initiation Factor-2/metabolism , Female , Gene Expression Regulation , Heat-Shock Proteins/agonists , Heat-Shock Proteins/genetics , Heat-Shock Proteins/metabolism , Humans , Membrane Potential, Mitochondrial/drug effects , Mitochondria/metabolism , Mitogen-Activated Protein Kinases/metabolism , Phosphatidylinositol 3-Kinases/genetics , Phosphatidylinositol 3-Kinases/metabolism , Phosphoinositide-3 Kinase Inhibitors , Protein Serine-Threonine Kinases/genetics , Protein Serine-Threonine Kinases/metabolism , Reactive Oxygen Species/agonists , Reactive Oxygen Species/metabolism , Signal Transduction , Stromal Cells/metabolism , Stromal Cells/pathology , Transcription Factor CHOP/agonists , Transcription Factor CHOP/genetics , Transcription Factor CHOP/metabolism , bcl-2 Homologous Antagonist-Killer Protein/agonists , bcl-2 Homologous Antagonist-Killer Protein/genetics , bcl-2 Homologous Antagonist-Killer Protein/metabolism , bcl-2-Associated X Protein/agonists , bcl-2-Associated X Protein/genetics , bcl-2-Associated X Protein/metabolismABSTRACT
The integrated stress response comprises multiple signaling pathways for detecting and responding to cellular stress that converge at a single event-the phosphorylation of Ser51 on the α-subunit of eukaryotic translation initiation factorâ 2 (eIF2α). Phosphorylation of eIF2α (eIF2α-P) results in attenuation of global protein synthesis via the inhibitory effects of eIF2α-P on eIF2B, the guanine exchange factor (GEF) for eIF2. Herein we describe structure-activity relationship (SAR) studies of bis-O-arylglycolamides, first-in-class integrated stress response inhibitors (ISRIB). ISRIB analogues make cells insensitive to the effects of eIF2α-P by activating the GEF activity of eIF2B and allowing global protein synthesis to proceed with residual unphosphorylated eIF2α. The SAR studies described herein support the proposed pharmacology of ISRIB analogues as binding across a symmetrical protein-protein interface formed between protein subunits of the dimeric eIF2B heteropentamer.
Subject(s)
Eukaryotic Initiation Factor-2/metabolism , Glycolates/pharmacology , Stress, Physiological/drug effects , Dose-Response Relationship, Drug , Eukaryotic Initiation Factor-2/agonists , Eukaryotic Initiation Factor-2/chemistry , Glycolates/chemical synthesis , Glycolates/chemistry , HEK293 Cells , Humans , Molecular Structure , Phosphorylation/drug effects , Protein Binding/drug effects , Structure-Activity RelationshipABSTRACT
Eukaryotic translation initiation factor 2 alpha (eIF2α), which is a component of the eukaryotic translation initiation complex, functions in cell death and survival under various stress conditions. In this study, we investigated the roles of eIF2α phosphorylation in cell death using the breast cancer cell lines MCF-7 and MCF-7/ADR. MCF-7/ADR cells are MCF-7-driven cells that have acquired resistance to doxorubicin (ADR). Treatment of doxorubicin reduced the viability and induced apoptosis in both cell lines, although susceptibility to the drug was very different. Treatment with doxorubicin induced phosphorylation of eIF2α in MCF-7 cells but not in MCF-7/ADR cells. Basal expression levels of Growth Arrest and DNA Damage 34 (GADD34), a regulator of eIF2α, were higher in MCF-7/ADR cells compared to MCF-7 cells. Indeed, treatment with salubrinal, an inhibitor of GADD34, resulted in the upregulation of eIF2α phosphorylation and enhanced doxorubicin-mediated apoptosis in MCF-7/ADR cells. However, MCF-7 cells did not show such synergic effects. These results suggest that dephosphorylation of eIF2α by GADD34 plays an important role in doxorubicin resistance in MCF-7/ADR cells.
Subject(s)
Antibiotics, Antineoplastic/pharmacology , Cinnamates/pharmacology , Doxorubicin/pharmacology , Drug Resistance, Neoplasm/drug effects , Eukaryotic Initiation Factor-2/genetics , Gene Expression Regulation, Neoplastic , Protein Phosphatase 1/genetics , Thiourea/analogs & derivatives , Apoptosis/drug effects , Cell Line, Tumor , Cell Survival/drug effects , Drug Resistance, Neoplasm/genetics , Drug Synergism , Eukaryotic Initiation Factor-2/agonists , Eukaryotic Initiation Factor-2/metabolism , Female , Humans , MCF-7 Cells , Phosphorylation/drug effects , Protein Phosphatase 1/metabolism , Signal Transduction , Thiourea/pharmacology , Transcriptional ActivationABSTRACT
Avarol is a sesquiterpenoid hydroquinone with potent cytotoxicity. Although resolving endoplasmic reticulum (ER) stress is essential for intracellular homeostasis, erratic or excessive ER stress can lead to apoptosis. Here, we reported that avarol selectively induces cell death in pancreatic ductal adenocarcinomas (PDAC), which are difficult to treat owing to the availability of few chemotherapeutic agents. Analyses of the molecular mechanisms of avarol-induced apoptosis indicated upregulation of ER stress marker BiP and ER stress-dependent apoptosis inducer CHOP in PDAC cells but not in normal cells, suggesting that avarol selectively induces ER stress responses. We also showed that avarol activated the PERK-eIF2α pathway but did not affect the IRE1 and ATF6 pathways. Moreover, CHOP downregulation was significantly suppressed by avarol-induced apoptosis. Thus, the PERK-eIF2α-CHOP signaling pathway may be a novel molecular mechanism of avarol-induced apoptosis. The present data indicate that avarol has potential as a chemotherapeutic agent for PDAC and induces apoptosis by activating the PERK-eIF2α pathway.
Subject(s)
Antineoplastic Agents/pharmacology , Carcinoma, Pancreatic Ductal/drug therapy , Pancreatic Neoplasms/drug therapy , Sesquiterpenes/pharmacology , Signal Transduction/drug effects , Transcription Factor CHOP/agonists , eIF-2 Kinase/metabolism , Animals , Antineoplastic Agents/adverse effects , Apoptosis/drug effects , Biomarkers/metabolism , Carcinoma, Pancreatic Ductal/metabolism , Cell Line , Cell Line, Tumor , Dysidea/chemistry , Endoplasmic Reticulum Chaperone BiP , Endoplasmic Reticulum Stress/drug effects , Eukaryotic Initiation Factor-2/agonists , Eukaryotic Initiation Factor-2/metabolism , Gene Expression Regulation, Neoplastic/drug effects , Heat-Shock Proteins/agonists , Heat-Shock Proteins/genetics , Heat-Shock Proteins/metabolism , Humans , Inhibitory Concentration 50 , Neoplasm Proteins/agonists , Neoplasm Proteins/antagonists & inhibitors , Neoplasm Proteins/genetics , Neoplasm Proteins/metabolism , Pancreatic Neoplasms/metabolism , RNA Interference , Sesquiterpenes/adverse effects , Transcription Factor CHOP/antagonists & inhibitors , Transcription Factor CHOP/genetics , Transcription Factor CHOP/metabolism , Up-Regulation/drug effects , eIF-2 Kinase/chemistryABSTRACT
It has been recently reported that p53 activation can lead to the induction of PKR-eIF2alpha phosphorylation pathway. However, we demonstrate that PKR and eIF2alpha phosphorylation at serine 51 are not downstream of p53 in response to doxorubicin treatment suggesting that PKR is not a universal target of p53 as proposed.
Subject(s)
Colorectal Neoplasms/metabolism , Cyclin-Dependent Kinase Inhibitor p21/metabolism , DNA Damage , Eukaryotic Initiation Factor-2/metabolism , Tumor Suppressor Protein p53/metabolism , eIF-2 Kinase/metabolism , Animals , Colorectal Neoplasms/drug therapy , Cyclin-Dependent Kinase Inhibitor p21/agonists , Doxorubicin/pharmacology , Doxorubicin/therapeutic use , Eukaryotic Initiation Factor-2/agonists , Humans , Mice , Phosphorylation/drug effects , Phosphorylation/physiology , Tumor Suppressor Protein p53/agonists , Tumor Suppressor Protein p53/genetics , Xenograft Model Antitumor Assays , eIF-2 Kinase/drug effectsABSTRACT
The mechanisms that determine whether a tumor cell that has disseminated to a secondary site will resume growth immediately, die or enter a state of dormancy are poorly understood. Although tumor dormancy represents a common clinical finding, studying the mechanisms behind this stage of tumor progression has been challenging. Furthermore, it is thought that dormant tumor cells are refractory to chemotherapy due to their lack of proliferation. However, whether this is the only reason for their chemo-resistance remains to be proven. In this review we summarize recent findings that provide a mechanistic explanation about how stress signaling through the p38(SAPK) pathway and ER-stress signaling may coordinate the induction of growth arrest and drug-resistance in a model of squamous carcinoma dormancy. We further discuss how dormant tumor cells may enter this stage to adapt to strenuous conditions that do not favor immediate growth after dissemination. Finally, we propose that this response may recapitulate an evolutionarily conserved program of life-span extension through adaptation and tolerance to stress.