TFEB Regulates ATP7B Expression to Promote Platinum Chemoresistance in Human Ovarian Cancer Cells

ATP7B is a hepato-specific Golgi-located ATPase, which plays a key role in the regulation of copper (Cu) homeostasis and signaling. In response to elevated Cu levels, ATP7B traffics from the Golgi to endo-lysosomal structures, where it sequesters excess copper and further promotes its excretion to the bile at the apical surface of hepatocytes. In addition to liver, high ATP7B expression has been reported in tumors with elevated resistance to platinum (Pt)-based chemotherapy. Chemoresistance to Pt drugs represents the current major obstacle for the treatment of large cohorts of cancer patients. Although the mechanisms underlying Pt-tolerance are still ambiguous, accumulating evidence suggests that lysosomal sequestration of Pt drugs by ion transporters (including ATP7B) might significantly contribute to drug resistance development. In this context, signaling mechanisms regulating the expression of transporters such as ATP7B are of great importance. Considering this notion, we investigated whether ATP7B expression in Pt-resistant cells might be driven by transcription factor EB (TFEB), a master regulator of lysosomal gene transcription. Using resistant ovarian cancer IGROV-CP20 cells, we found that TFEB directly binds to the predicted coordinated lysosomal expression and regulation (CLEAR) sites in the proximal promoter and first intron region of ATP7B upon Pt exposure. This binding accelerates transcription of luciferase reporters containing ATP7B CLEAR regions, while suppression of TFEB inhibits ATP7B expression and stimulates cisplatin toxicity in resistant cells. Thus, these data have uncovered a Pt-dependent transcriptional mechanism that contributes to cancer chemoresistance and might be further explored for therapeutic purposes.

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HOTTIP-miR-205-ZEB2 Axis Confers Cisplatin Resistance to Ovarian Cancer Cells

Frontiers in Cell and Developmental Biology ◽

10.3389/fcell.2021.707424 ◽

2021 ◽

Vol 9 ◽

Author(s):

Yu-Jie Dong ◽

Wei Feng ◽

Yan Li

Keyword(s):

Ovarian Cancer ◽

Cancer Cells ◽

Cisplatin Resistance ◽

Clinical Problem ◽

Ovarian Cancer Cells ◽

Stem Cell Markers ◽

Gene Target ◽

Gynecological Malignancy ◽

Clonogenic Potential ◽

Resistant Cells

Ovarian cancer is a deadly gynecological malignancy with resistance to cisplatin a major clinical problem. We evaluated a role of long non-coding (lnc) RNA HOTTIP (HOXA transcript at the distal tip) in the cisplatin resistance of ovarian cancer cells, using paired cisplatin sensitive and resistant A2780 cells along with the SK-OV-3 cells. HOTTIP was significantly elevated in cisplatin resistant cells and its silencing reversed the cisplatin resistance of resistant cells. HOTTIP was found to sponge miR-205 and therefore HOTTIP silenced cells had higher levels of miR-205. Downregulation of miR-205 could attenuate HOTTIP-silencing effects whereas miR-205 upregulation in resistant cells was found to re-sensitize cells to cisplatin. HOTTIP silencing also led to reduced NF-κB activation, clonogenic potential and the reduced expression of stem cell markers SOX2, OCT4, and NANOG, an effect that could be attenuated by miR-205. Finally, ZEB2 was identified as the gene target of miR-205, thus completing the elucidation of HOTTIP-miR-205-ZEB2 as the novel axis which is functionally involved in the determination of cisplatin resistance in ovarian cancer cells.

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Short-Term Serum Deprivation Confers Sensitivity to Taxanes in Platinum-Resistant Human Ovarian Cancer Cells

International Journal of Gynecological Cancer ◽

10.1097/igc.0b013e318231b994 ◽

2011 ◽

Vol 21 (9) ◽

pp. 1547-1554 ◽

Cited By ~ 2

Author(s):

Seiji Isonishi ◽

Motoaki Saito ◽

Misato Saito ◽

Tadao Tanaka

Keyword(s):

Ovarian Cancer ◽

Cancer Cells ◽

Flow Cytometric Analysis ◽

Serum Deprivation ◽

Ovarian Cancer Cells ◽

Enhancement Effect ◽

Human Ovarian Cancer ◽

Platinum Resistant ◽

Mitochondrial Functions ◽

Resistant Cells

BackgroundBased on the evidences showing that serum deprivation provokes apoptosis in a variety of cells, we have investigated the effect of serum deprivation on drug sensitivity.MethodsAfter human ovarian cancer cells were preincubated in 0.5 % serum containing medium for 12 hours, cellular drug sensitivities were determined by colony-forming assay.ResultsSerum deprivation treatment resulted in significant increase in paclitaxel sensitivity by factors of mean ± SD, 148.6 ± 28.1 and 10.1 ± 1.0 (n = 3;P< 0.001) fold in platinum-resistant C13 and CP70 cells, respectively. Similarly, serum deprivation induced significant docetaxel sensitivity in these cell lines. However, no enhancement effect of serum deprivation was observed in platinum-sensitive 2008 and A2780 cells. Serum deprivation did not have any effect on the sensitivities to cisplatin, vincristin, and doxorubicin in all of these cells. More than 7-fold increase of apoptotic cells were observed in C13 or CP70 cells when they were treated by serum deprivation followed by paclitaxel compared with the treatment of either serum deprivation or paclitaxel alone. Confocal laser microscopy using rhodamine 123 and flow cytometric analysis with 3,3′-dihexyloxacarbocyanine iodide revealed that serum deprivation decreased mitochondrial membrane potential in C13 or CP70 cells, whereas no change was observed in 2008 and A2780 cells. This indicates that serum deprivation induced depolarization specifically in platinum-resistant cells. Electron microscopy revealed that serum deprivation caused regeneration of mitochondrial matrix structure in C13 or CP70 cells where mitochondria were usually destructed and disappeared.DiscussionsThese results indicate that serum deprivation confers taxane hypersensitivity specifically in platinum-resistant cells by recovering their impaired mitochondrial functions. The evidence might be clinically beneficial for the development of new chemotherapeutic technology, particularly for the patients with platinum-resistant ovarian cancer.

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MiR-200c sensitizes Olaparib-resistant ovarian cancer cells by targeting Neuropilin 1

Journal of Experimental & Clinical Cancer Research ◽

10.1186/s13046-019-1490-7 ◽

2020 ◽

Vol 39 (1) ◽

Cited By ~ 5

Author(s):

Enrica Vescarelli ◽

Giulia Gerini ◽

Francesca Megiorni ◽

Eleni Anastasiadou ◽

Paola Pontecorvi ◽

...

Keyword(s):

Ovarian Cancer ◽

Western Blot ◽

Cell Lines ◽

Target Genes ◽

Model Systems ◽

Ovarian Cancer Cells ◽

Drug Resistant ◽

Neuropilin 1 ◽

Skov3 Cells ◽

Resistant Cells

Abstract Background Ovarian cancer (OC) is the most lethal gynecological malignancy and the second leading cause of cancer-related death in women. Treatment with PARP inhibitors (PARPi), such as Olaparib, has been recently introduced for OC patients, but resistance may occur and underlying mechanisms are still poorly understood. The aim of this study is to identify target genes within the tumor cells that might cause resistance to Olaparib. We focused on Neuropilin 1 (NRP1), a transmembrane receptor expressed in OC and correlated with poor survival, which has been also proposed as a key molecule in OC multidrug resistance. Methods Using three OC cell lines (UWB, UWB-BRCA and SKOV3) as model systems, we evaluated the biological and molecular effects of Olaparib on OC cell growth, cell cycle, DNA damage and apoptosis/autophagy induction, through MTT and colony forming assays, flow cytometry, immunofluorescence and Western blot analyses. We evaluated NRP1 expression in OC specimens and cell lines by Western blot and qRT-PCR, and used RNA interference to selectively inhibit NRP1. To identify miR-200c as a regulator of NRP1, we used miRNA target prediction algorithms and Pearsons’ correlation analysis in biopsies from OC patients. Then, we used a stable transfection approach to overexpress miR-200c in Olaparib-resistant cells. Results We observed that NRP1 is expressed at high levels in resistant cells (SKOV3) and is upmodulated in partially sensitive cells (UWB-BRCA) upon prolonged Olaparib treatment, leading to poor drug response. Our results show that the selective inhibition of NRP1 is able to overcome Olaparib resistance in SKOV3 cells. Moreover, we demonstrated that miR-200c can target NRP1 in OC cells, causing its downmodulation, and that miR-200c overexpression is a valid approach to restore Olaparib sensitivity in OC resistant cells. Conclusions These data demonstrate that miR-200c significantly enhanced the anti-cancer efficacy of Olaparib in drug-resistant OC cells. Thus, the combination of Olaparib with miRNA-based therapy may represent a promising treatment for drug resistant OC, and our data may help in designing novel precision medicine trials for optimizing the clinical use of PARPi.

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Polyurea Dendrimer Folate-Targeted Nanodelivery of l-Buthionine Sulfoximine as a Tool to Tackle Ovarian Cancer Chemoresistance

Antioxidants ◽

10.3390/antiox9020133 ◽

2020 ◽

Vol 9 (2) ◽

pp. 133 ◽

Cited By ~ 1

Author(s):

Adriana Cruz ◽

Pedro Mota ◽

Cristiano Ramos ◽

Rita F. Pires ◽

Cindy Mendes ◽

...

Keyword(s):

Ovarian Cancer ◽

Cancer Cells ◽

Buthionine Sulfoximine ◽

Ovarian Cancer Cells ◽

Metabolic Remodeling ◽

Metabolic Mechanism ◽

In Vitro Inhibition ◽

New Formulation ◽

Cancer Chemoresistance

Ovarian cancer is a highly lethal disease, mainly due to chemoresistance. Our previous studies on metabolic remodeling in ovarian cancer have supported that the reliance on glutathione (GSH) bioavailability is a main adaptive metabolic mechanism, also accounting for chemoresistance to conventional therapy based on platinum salts. In this study, we tested the effects of the in vitro inhibition of GSH synthesis on the restoration of ovarian cancer cells sensitivity to carboplatin. GSH synthesis was inhibited by exposing cells to l-buthionine sulfoximine (l-BSO), an inhibitor of γ-glutamylcysteine ligase (GCL). Given the systemic toxicity of l-BSO, we developed a new formulation using polyurea (PURE) dendrimers nanoparticles (l-BSO@PUREG4-FA2), targeting l-BSO delivery in a folate functionalized nanoparticle.

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Metformin inhibits the growth of ovarian cancer cells by promoting the Parkin-induced p53 ubiquitination

Bioscience Reports ◽

10.1042/bsr20200679 ◽

2020 ◽

Cited By ~ 1

Author(s):

Xiaojia Min ◽

Tingting Zhang ◽

Ying Lin ◽

Bo Wang ◽

Kean Zhu

Keyword(s):

Ovarian Cancer ◽

Cancer Cells ◽

Molecular Mechanisms ◽

Ovarian Cancer Cells ◽

Related Factors ◽

P53 Signaling ◽

P53 Signaling Pathway ◽

Resistant Cells

Ovarian cancer is the most lethal diseases among women. The chemo-resistance has been a big challenge for the cancer treatment. It has been reported that metformin may inhibit ovarian cancer and is able to impede the development of drug resistance, but the molecular mechanisms remain elusive. In this study, we explored the molecular roles of metformin in Parkin expression and p53 ubiquitination in chemo-resistant ovarian cancer cells. Firstly, ovarian cancer and chemo-resistant ovarian cancer cells were selected for determining the expression of Parkin, p53, and p53 signaling pathway-related factors. Then the cell proliferation and viability after loss- and gain-of-function assays were measured. Besides, immunoprecipitation (IP) was used to determine the interactions between Parkin and p53, and the ubiquitination level of p53 was measured using in vitro ubiquitination assay. Finally, the degradation of p53 proteasome regulated by Parkin was monitored using the MG132 proteasome inhibitor. We found that metformin significantly inhibited the growth of ovarian cancer parental cells and chemo-resistant cells, and metformin promoted Parkin expression in chemo-resistant cells. Further, up-regulated Parkin expression promoted the ubiquitination and degradation of p53, and metformin inhibited the expression of p53 to suppress the proliferation of chemo-resistant ovarian cancer cells. Mechanistically, metformin could inhibit the growth of ovarian cancer cells by promoting the Parkin-induced p53 ubiquitination. Altogether, our study demonstrated an inhibitory role of metformin in the growth of chemo-resistant cancer cells through promoting the Parkin-induced p53 ubiquitination, which provides a novel mechanism of metformin for treating ovarian cancer.

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Activation of the reverse transsulfuration pathway through NRF2/CBS confers erastin-induced ferroptosis resistance

British Journal of Cancer ◽

10.1038/s41416-019-0660-x ◽

2019 ◽

Vol 122 (2) ◽

pp. 279-292 ◽

Cited By ~ 18

Author(s):

Nan Liu ◽

Xiaoli Lin ◽

Chengying Huang

Keyword(s):

Ovarian Cancer ◽

Cell Death ◽

Cancer Cells ◽

Therapeutic Response ◽

Lipid Peroxide ◽

Ovarian Cancer Cells ◽

Transsulfuration Pathway ◽

Selective Elimination ◽

Sustained Activation ◽

Resistant Cells

Abstract Background Ferroptosis is an iron-dependent, lipid peroxide-mediated cell death that may be exploited to selective elimination of damaged and malignant cells. Recent studies have identified that small-molecule erastin specifically inhibits transmembrane cystine–glutamate antiporter system xc−, prevents extracellular cystine import and ultimately causes ferroptosis in certain cancer cells. In this study, we aimed to investigate the molecular mechanism underlying erastin-induced ferroptosis resistance in ovarian cancer cells. Methods We treated ovarian cancer cells with erastin and examined cell viability, cellular ROS and metabolites of the transsulfuration pathway. We also depleted cystathionine β-synthase (CBS) and NRF2 to investigate the CBS and NRF2 dependency in erastin-resistant cells. Results We found that prolonged erastin treatment induced ferroptosis resistance. Upon exposure to erastin, cells gradually adapted to cystine deprivation via sustained activation of the reverse transsulfuration pathway, allowing the cells to bypass erastin insult. CBS, the biosynthetic enzyme for cysteine, was constantly upregulated and was critical for the resistance. Knockdown of CBS by RNAi in erastin-resistant cells caused ferroptotic cell death, while CBS overexpression conferred ferroptosis resistance. We determined that the antioxidant transcriptional factor, NRF2 was constitutively activated in erastin-resistant cells and NRF2 transcriptionally upregulated CBS. Genetically repression of NRF2 enhanced ferroptosis susceptibility. Conclusions Based on these results, we concluded that constitutive activation of NRF2/CBS signalling confers erastin-induced ferroptosis resistance. This study demonstrates a new mechanism underlying ferroptosis resistance, and has implications for the therapeutic response to erastin-induced ferroptosis.

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Corilagin Reduces Resistance to PARP Inhibitors by Inhibiting the ERK Signaling Pathway in Ovarian Cancers

10.21203/rs.3.rs-318225/v1 ◽

2021 ◽

Author(s):

Baoxin Luan ◽

Hongbo Zhao ◽

Robert C. Bast ◽

Zhen Lu ◽

Yinhua Yu

Keyword(s):

Ovarian Cancer ◽

Cancer Cells ◽

Cell Lines ◽

Excision Repair ◽

Resistant Cell ◽

Erk Signaling ◽

Ovarian Cancer Cells ◽

Dependent Manner ◽

Ovarian Cancers ◽

Resistant Cells

Abstract Background: Corilagin is a compound with hepatoprotective and antiviral activity extracted from Phyllanthus niruri L. Our previous work demonstrated that corilagin inhibits the growth of ovarian cancer cells by regulating the TGF-β/AKT/ERK signaling. Corilagin was also found to sensitize ovarian cancer cells to paclitaxel and carboplatin by inhibiting the Snail-glycolysis pathway. We have now studied whether corilagin could overcome resistance of ovarian cancer cells to poly ADP ribose polymerase inhibitors (PARPi). PARPi block DNA base excision repair and have been approved for treatment of ovarian cancers. Drug resistance has limited efficacy of PARPi. Methods: We have assessed the effect of corilagin alone and in combination with PARPi in two pairs of ovarian cancer cell lines-A2780CP/A2780CP_R and UWB1.289/UWB1.289_R-that are sensitive or resistant to PARPi. CulcuSyn software (BIOSOFT-Software for Science, Cambridge, U.K.) was used to calculate synergy between two drug combinations. Results: Corilagin was active against all four cell lines and enhanced BMN673 activity synergistically in both PARPi resistant cell lines. PARPi-BMN673 down-regulated the expression levels of PARP and up-regulated pH2AX, it decreased pERK activity in sensitive cell lines, but not in resistant cell lines. While corilagin affected DNA repair function to some extent, it inhibited pERK activity in both PARPi sensitive and resistant cells in a dose dependent manner. Corilagin, but not the BMN673, inhibited ZEB1 in resistant cells. Conclusions: Corilagin deserves further evaluation as a drug that could enhance the activity of PARPi in PARPi-resistant ovarian cancer cells.

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Overexpression of miR-138-5p Sensitizes Taxol-Resistant Epithelial Ovarian Cancer Cells through Targeting Cyclin-Dependent Kinase 6

Gynecologic and Obstetric Investigation ◽

10.1159/000518510 ◽

2021 ◽

pp. 1-9

Author(s):

Man Liang ◽

Qin Li ◽

Shuai Shi ◽

Ya-ning Tian ◽

Yanhong Feng ◽

...

Keyword(s):

Ovarian Cancer ◽

Epithelial Ovarian Cancer ◽

Cancer Cells ◽

Cell Line ◽

Cancer Cell ◽

Luciferase Assay ◽

Ovarian Cancer Cells ◽

Cyclin Dependent Kinase ◽

Cancer Tissues ◽

Resistant Cells

Background: Ovarian cancer, one of the most malignant diseases in female, is associated with poor diagnosis and low 5-year survival rate. Taxol is a widely used chemotherapeutic drug for the treatment of ovarian cancer by targeting the microtubules of the mitotic spindle to induce cancer cell death. However, with the widespread clinical applications of Taxol, a large fraction of ovarian cancer patients developed drug resistance. Results: Here, we report miR-138-5p is significantly downregulated in epithelial ovarian cancer tissues compared with their matched normal ovarian tissues. Overexpression of miR-138-5p effectively sensitized ovarian cancer cells to Taxol. By establishing Taxol-resistant cell line from the epithelial ovarian cancer cell line, HO-8910, we found miR-138-5p was significantly downregulated in Taxol-resistant cells. Furthermore, overexpression of miR-138-5p dramatically overcame the chemoresistance of Taxol-resistant cells. Intriguingly, bioinformatic analysis indicated miR-138-5p had putative binding sites for cyclin-dependent kinase 6 (CDK6). This negative regulation was further verified from epithelial ovarian cancer tissues. Luciferase assay demonstrated miR-138-5p could directly bind to 3′UTR of CDK6. Importantly, silencing CDK6 expression by siRNA successfully increased the sensitivity of both parental and Taxol-resistant ovarian cancer cells. Finally, rescue experiments clearly elucidated restoration of CDK6 in miR-138-5p-overexpressing ovarian cancer cells successfully recovered the Taxol resistance. Conclusion: In summary, these findings suggest important molecular mechanisms for the miR-138-5p-mediated Taxol sensitivity of ovarian cancer via directly targeting CDK6, suggesting miR-138-5p is an effective therapeutic target for the noncoding RNA-based anti-chemoresistance treatment.

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Esomeprazole improves the acidic microenvironment of epithelial ovarian cancer by inhibiting the expression of VATPase

Tropical Journal of Pharmaceutical Research ◽

10.4314/tjpr.v19i8.3 ◽

2020 ◽

Vol 19 (8) ◽

pp. 1585-1590

Author(s):

Yasong Chi ◽

Ruiqin Yue ◽

Yanru Lv ◽

Wei Liao ◽

Ruchang Yin

Keyword(s):

Ovarian Cancer ◽

Epithelial Ovarian Cancer ◽

Cancer Cells ◽

Ph Value ◽

Ovarian Cancer Cells ◽

Control Group ◽

Acidic Microenvironment ◽

Paclitaxel Group ◽

Resistant Cells

Purpose: To determine the effect of esomeprazole on apoptosis of ovarian cancer cells and their sensitivity to paclitaxel, and the underlying mechanism.Methods: Human ovarian paclitaxel-resistant cancer cells were cultured in vitro, and treated with esomeprazole at doses of 50, 100 and 250 mol/L. Cell proliferation was determined using MTT assay. Paclitaxel-resistant cells were divided into control group, esomeprazole group, paclitaxel group, and esomeprazole + taxol group. Western blot was employed for the assay of protein levels of bcl-2, Bcl-xl, P-gp and V-ATPase, while BCECF-AM method was employed to determine changes in intracellular pH.Results: Esomeprazole significantly inhibited the proliferation of paclitaxel-resistant cells in a dosedependent manner. The half-maximal inhibitory concentration (IC50) value of esomeprazole + paclitaxel was significantly low, when compared with those of the other treatments (p < 0.05). Apoptosis was significantly higher in esomeprazole + paclitaxel group than in any other treatment group (p < 0.05). The expressions of Bcl-2 and P-gp in esomeprazole + paclitaxel group decreased significantly, relative to the corresponding values for other groups, while protein expression of bcl-xl was markedly increased. The intracellular pH value of esomeprazole + paclitaxel group was significantly lower than those for other treatment groups (p < 0.05).Conclusion: Esomeprazole improves the acidic microenvironment of epithelial ovarian cancer by inhibiting the expression of V-ATPase, and restores the sensitivity of ovarian cancer cells to paclitaxel by inhibiting their proliferation and apoptosis. This revelation may explain patients’ resistance topaclitaxel. Keywords: Esomeprazole, V-ATPase, Apoptosis, Ovarian cancer, Taxol, Sensitivity

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Protein Expression Patterns in ovarian cancer cells Associated with Monofunctional Platinums Treatment

10.1101/628958 ◽

2019 ◽

Cited By ~ 1

Author(s):

Laila Arzuman ◽

Mohammad Ali Moni ◽

Philip Beale ◽

Jun Q. Yu ◽

Mark Molloy ◽

...

Keyword(s):

Ovarian Cancer ◽

Patient Survival ◽

Expression Patterns ◽

Differentially Expressed ◽

Ovarian Cancer Cells ◽

Altered Expression ◽

Expression Levels ◽

Risk Of Death ◽

Platinum Based Chemotherapy ◽

Resistant Cells

ABSTRACTPlatinum drugs cisplatin and carboplatin, given in combination with paclitaxel, constitute the standard chemotherapy against ovarian cancer (OC). Oc chemoresistance is a major obstacle to effective treatment, but knowledge of the mechanisms that underlie it remains incomplete. We thus sought to discover key proteins associated with platinum resistance by comparing A2780 OC cells with A2780cisR cells (resistant cells derived from the A2780 line) to identify proteins with markedly altered expression levels in the resistant cells. We also determined which proteins in these cells had altered expression in response to treatment with either designed monofunctional platinum alone or a combination with cisplatin with selected phytochemical therapeutic agents.We thus performed proteomic analysis using 2D-gel electrophoresis A2780 and A2780cisR to identify proteins with differential expression; these were eluted and analysed by mass spectrometry to identify them. A total of 122 proteins were found to be differentially expressed between A2780 and A2780cisR cell lines in the absence of any drug treatment. Among them, levels of 27 proteins in A2780cisR cell line were further altered (up-or down-regulated) in response to one or more of the drug treatments. We then investigated primary OC tissue RNA expression levels (compared to l ovarian tissue) of genes coding for these candidate 27 proteins using publically available datasets (The Cancer Genome Atlas). We assessed how expression of these genes in OC tissue associates with patient survival using Cox Proportional Hazard (PH) regression models to determine relative risk of death associated with each factor. Our Cox PH regression-based machine learning method confirmed a significant relationship of mortality with altered expression of ARHGDIA, CCT6A and HISTIH4F genes. This indicated that these genes affect OC patient survival, i.e., provided mechanistic evidence, in addition to that of the clinical traits, that these genes may be critical mediators of the processes that underlie OC progression and mortality.Thus, we identified differentially expressed proteins that are implicated in platinum-based chemotherapy resistance mechanisms which may serve as resistance biomarkers. These drug resistance associated proteins may also serve as potential OC therapeutic targets whose blockade may enhance the effectiveness of platinum based drugs.

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