Metabolic shift toward oxidative phosphorylation in docetaxel resistant prostate cancer cells.
ABSTRACT: Drug resistance of cancer cells is recognized as the primary cause of failure of chemotherapeutic treatment in most human cancers. Growing evidences support the idea that deregulated cellular metabolism is linked to such resistance. Indeed, both components of the glycolytic and mitochondrial pathways are involved in altered metabolism linked to chemoresistance of several cancers. Here we investigated the drug-induced metabolic adaptations able to confer advantages to docetaxel resistant prostate cancer (PCa) cells. We found that docetaxel-resistant PC3 cells (PC3-DR) acquire a pro-invasive behavior undergoing epithelial-to-mesenchymal-transition (EMT) and a decrease of both intracellular ROS and cell growth. Metabolic analyses revealed that PC3-DR cells have a more efficient respiratory phenotype than sensitive cells, involving utilization of glucose, glutamine and lactate by the mitochondrial oxidative phosphorylation (OXPHOS). Consequently, targeting mitochondrial complex I by metformin administration, impairs proliferation and invasiveness of PC3-DR cells without effects on parental cells. Furthermore, stromal fibroblasts, which cause a "reverse Warburg" phenotype in PCa cells, reduce docetaxel toxicity in both sensitive and resistant PCa cells. However, re-expression of miR-205, a microRNA strongly down-regulated in EMT and associated to docetaxel resistance, is able to shift OXPHOS to a Warburg metabolism, thereby resulting in an elevated docetaxel toxicity in PCa cells. Taken together, these findings suggest that resistance to docetaxel induces a shift from Warburg to OXPHOS, mandatory for conferring a survival advantage to resistant cells, suggesting that impairing such metabolic reprogramming could be a successful therapeutic approach.
Project description:BACKGROUND:Prostate cancer (PCa) is one of the most frequently diagnosed tumors in men. In general, therapies for localized PCa are curative. However, treatment of advanced PCa is considered palliative since development of therapy resistance occurs rapidly. It has been shown that tumor-initiating cells are likely involved in therapy resistance. They are not eliminated by conventional therapies and thereby lead to tumor progression and relapse. The aim of this study was to evaluate the effects of the known stem cell inhibitor salinomycin on this critical subpopulation of cells. METHODS:Expression of the cell surface markers CD24 and CD44 was assessed by immunofluorescence and fluorescence-activated cell sorting. Colony formation efficiency and classification of colony types with varying tumor-initiating potential (holoclones, meroclones, and paraclones) were analyzed in an automated way by the newly developed CATCH-colonies software in the absence or presence of salinomycin. RESULTS:Automated high-resolution colony formation analysis consistently identified the various colony types in a broad range of PCa cell lines. Serial clonogenic assays confirmed that holoclones show the highest colony formation potential and maintain their tumor-initiating capacity over multiple rounds. Furthermore, holoclones showed high expression of CD44, while CD24 was not expressed in these clones, thus representing the well-described tumor-initiating CD24- /CD44high population. Salinomycin decreased the CD24- /CD44high population in both docetaxel-sensitive PC3 and docetaxel-resistant (DR) PC3-DR. Moreover, treatment of PC3, DU145, PC3-DR, and DU145-DR with salinomycin led to a significant reduction in the colony formation potential by targeting the colonies with high tumor-initiating potential. CONCLUSIONS:Taken together, we demonstrated that salinomycin specifically targets the tumor-initiating cell population in docetaxel-sensitive and docetaxel-resistant PCa cells and may represent a potential therapeutic approach for the treatment of advanced PCa.
Project description:This study focuses on the effect of miR-129-5p on docetaxel-resistant (DR) prostate cancer (PCa) cells invasion, migration and apoptosis. In our study, the expression of CAMK2N1 was assessed by qRT-PCR in PCa patient tissues and cell lines including PC-3 and PC-3-DR. Cells transfected with miR-129-5p mimics, inhibitor, CAMK2N1 or negative controls (NC) were used to interrogate their effects on DR cell invasions, migrations and apoptosis during docetaxel (DTX) treatments. The apoptosis rate of the PCa cells was validated by flow cytometry. Relationships between miR-129-5p and CAMK2N1 levels were identified by qRT-PCR and dual-luciferase reporter assay. CAMK2N1 was found to be down-expressed in DR PCa tissue sample, and low levels of CAMK2N1 were correlated with high docetaxel resistance and clinical prediction of poor survival. CAMK2N1 levels were decreased in DR PCa cells treated with DXT. We further explored that up-regulation of miR-129-5p could promote DR PCa cells viability, invasion and migration but demote apoptosis. Involved molecular mechanism studies revealed that miR-129-5p reduced downstream CAMK2N1 expression to further impact on chemoresistance to docetaxel of PCa cells, indicating its vital role in PCa docetaxel resistance. Our findings revealed that miR-129-5p contributed to the resistance of PC-3-DR cells to docetaxel through suppressing CAMK2N1 expression, and thus targeting miR-129-5p may provide a novel therapeutic approach in sensitizing PCa to future docetaxel treatment.
Project description:Acquired docetaxel-resistance of prostate cancer (PCa) remains a clinical obstacle due to the lack of effective therapies. Acetyl-11-keto-?-boswellic acid (AKBA) is a pentacyclic triterpenic acid isolated from the fragrant gum resin of the Boswellia serrata tree, which has shown intriguing antitumor activity against human cell lines established from PCa, colon cancer, malignant glioma, and leukemia. In this study, we examined the effects of AKBA against docetaxel-resistant PCa in vitro and in vivo as well as its anticancer mechanisms. We showed that AKBA dose-dependently inhibited cell proliferation and induced cell apoptosis in docetaxel-resistant PC3/Doc cells; its IC50 value in anti-proliferation was ?17??M. Furthermore, AKBA dose-dependently suppressed the chemoresistant stem cell-like properties of PC3/Doc cells, evidenced by significant decrease in the ability of mammosphere formation and down-regulated expression of a number of stemness-associated genes. The activation of Akt and Stat3 signaling pathways was remarkably enhanced in PC3/Doc cells, which contributed to their chemoresistant stem-like phenotype. AKBA (10-30??M) dose-dependently suppressed the activation of Akt and Stat3 signaling pathways in PC3/Doc cells. In contrast, overexpression of Akt and Stat3 significantly attenuated the inhibition of AKBA on PC3/Doc cell proliferation. In docetaxel-resistant PCa homograft mice, treatment with AKBA significantly suppresses the growth of homograft RM-1/Doc, equivalent to its human PC3/Doc, but did not decrease their body weight. In summary, we demonstrate that AKBA inhibits the growth inhibition of docetaxel-resistant PCa cells in vitro and in vivo via blocking Akt and Stat3 signaling, thus suppressing their cancer stem cell-like properties.
Project description:Resistance to docetaxel is a major clinical problem in advanced prostate cancer. The overexpression of AXL receptor tyrosine kinase (AXL) has been correlated with chemotherapeutic drug resistance. However, the role of AXL expression in docetaxel resistance in prostate cancer is yet unclear. In this study, we demonstrate that AXL is overexpressed and activated independent of Gas6 in docetaxel-resistant prostate cancer cells (PC3-DR and DU145-DR). Moreover, we show that forced overexpression of AXL in PC3 and DU145 cells is sufficient to induce resistance to docetaxel in these cell lines. Notably, genetic or pharmacologic inhibition of AXL in the resistant models suppressed cell proliferation, migration, invasion, and tumor growth, and these effects were significantly augmented when AXL inhibition was combined with docetaxel treatment. Mechanistically, we found that AXL inhibition led to reversion of the epithelial-mesenchymal transition (EMT) phenotype and decreased the expression of ATP-binding cassette B1 (ABCB1). Overall, our results identify AXL as an important mediator of docetaxel resistance in prostate cancer. We propose that AXL-targeted therapy, in combination with docetaxel, has the potential to improve the response to docetaxel therapy and reduce resistance induced by prolonged docetaxel therapy in prostate cancer.
Project description:Long noncoding RNAs (lncRNAs) affect docetaxel chemosensitivity. However, the biological role and regulatory mechanisms of lncRNAs in docetaxel-resistant prostate cancer remain unclear. Differences in lncRNAs were evaluated by lncRNA sequencing and evaluated using quantitative real-time polymerase chain reaction, and TrkB expression was measured through Western blot analysis. Proliferation was measured using the MTS, while apoptosis and cell cycle were measured using flow cytometry. Additionally, migration and invasion were measured using transwell assays. Forty-eight female BALB/c nude mice were used for subcutaneous tumorigenicity and lung metastasis assays. We found that LINC01963 was overexpressed in the PC3-DR cells. LINC01963 silencing enhanced the chemosensitivity of PC3-DR to docetaxel and inhibited tumorigenicity and lung metastasis, while LINC01963 overexpression enhanced the chemoresistance of PC3 cells to docetaxel. It was found that LINC01963 sponges miR-216b-5p. The miR-216b-5p inhibitor reversed the suppressive effect of sh-LINC01963 on PC3-DR cell proliferation, migration, and invasion. Furthermore, miR-216b-5p can bind to the 3’-UTR of NTRK2 and inhibit TrkB protein levels. TrkB enhances docetaxel resistance in prostate cancer and reverses the effects of LINC01963 silencing and miR-216b-5p overexpression. In conclusion, silencing LINC01963 enhanced the chemosensitivity of PC3-DR to docetaxel by sponging miR-216b-5p to inhibit TrkB protein levels. Overall design: Examination of 2 different PC3 cells, of which was PC3 and docetaxel-resistance PC3.
Project description:In the current study, we investigated a combination of docetaxel and thalidomide (DT therapy) in castration-resistant prostate cancer (CRPC) patients. We identified marker genes that predict the effect of DT therapy. Using an androgen-insensitive PC3 cell line, we established a docetaxel-resistant PC-3 cell line (DR-PC3). In DR-PC3 cells, DT therapy stronger inhibited proliferation/viability than docetaxel alone. Based on gene ontology analysis, we found versican as a selective gene. This result with the findings of cDNA microarray and validated by quantitative RT-PCR. In addition, the effect of DT therapy on cell viability was the same as the effect of docetaxel plus versican siRNA. In other words, silencing of versican can substitute for thalidomide. In the clinical setting, versican expression in prostate biopsy samples (before DT therapy) correlated with PSA reduction after DT therapy (p<0.05). Thus targeting versican is a potential therapeutic strategy in docetaxel-resistant prostate cancer.
Project description:BACKGROUND:Although docetaxel-based chemohormonal therapy (CHT) is one of the standard treatments for castration-resistant prostate cancer (CRPC), pertinent biomarkers and precise mechanisms involved in the resistance for CHT for CRPC remain unknown. We investigated the relationship between chemohormonal resistance and the expression of steroid receptors and Hippo pathway proteins using a docetaxel-resistant prostate cancer (PCa) cell line and human PCa tissues in patients who underwent surgery with and without neoadjuvant therapy. METHODS:A docetaxel-resistant subline (22Rv1-DR) was generated to assess Hippo pathway protein expression and the effect of YAP1 inhibition on cellular characteristics. A tissue microarray with 203 cores from 70 high-risk localized PCa tissues was performed to assess steroid receptor and Hippo pathway protein expressions. RESULTS:Nuclear YAP (nYAP) expression was higher in 22RV-1-DR than in parental 22Rv-1 and YAP1 knockdown suppressed cell proliferation of 22Rv1-DR. Steroid receptor and Hippo pathway protein expressions varied among three different neoadjuvant groups, and nYAP1 expression was the highest in the CHT group. The patients with high nYAP in residual cancer after neoadjuvant CHT had a significantly higher biochemical recurrence (BCR) rate than those with low nYAP1. On multivariate analysis, the high nYAP1 was an independent prognostic factor for BCR. CONCLUSIONS:nYAP expression is a potential biomarker in high-risk patients treated with docetaxel-based CHT. Steroid receptors and Hippo pathway proteins may play a role in the chemohormonal resistance in advanced PCa.
Project description:The establishment of docetaxel-based chemotherapeutic treatments has improved the survival of castration-resistant prostate cancer (CRPC) patients. However, most patients develop resistance supporting the development of therapy. The current study was undertaken to establish the therapeutic benefit to target hedgehog signaling cascade using GDC-0449 to improve the efficacy of chemotherapeutic drug, docetaxel. Here, we show that the combination of GDC-0449 plus docetaxel inhibited the proliferation of WPE1-NB26 cells and PC3 cells via a blockade of G1 and G2M phases. The combined treatment significantly inhibited PC cell migration in vitro. Moreover, the apoptotic effect induced by GDC-0449 plus docetaxel on PC3 cells was mediated, at least partly, via the mitochondrial membrane depolarization, H2O2 production and caspase cascade activation. Interestingly, GDC-0449 was effective at inhibiting the prostasphere formation, inducing the prostasphere disintegration and apoptotic death of side population (SP) from PC3 cells and reversing the resistance of SP cells to docetaxel. In addition, GDC-0449 plus docetaxel also have shown a greater anti-tumoral growth inhibitory effect on PC3 cell xenografts. These findings support the use of the hedgehog inhibitor GDC-0449, which is currently in clinical trials, for improving the anticarcinogenic efficacy of docetaxel-based chemotherapeutic treatments against locally advanced, AI and metastatic PC.
Project description:Rationale: Docetaxel-mediated chemotherapy is the first-line standard approach and has been determined to show a survival advantage for metastatic castration-resistant prostate cancer (mCRPC) patients. However, a substantial proportion of patients eventually becomes refractory due to drug resistance. The detailed mechanisms remain unclear. We have previously reported that Prostate Leucine Zipper (PrLZ), a specific oncogene of prostate cancer (PCa), promotes PCa cell growth at the castration-resistant stage, thus suggesting a vital role of PrLZ in the progression of CRPC. In this study, we aimed to investigate the role of PrLZ in docetaxel resistance in PCa, focusing on PrLZ-regulating autophagy pathway. Methods: Human PCa PC3, LNCaP and C4-2 cell lines were used as the model system in vitro and PCa xenografts and PrLZ-knockout mice were used as the model system in vivo. Docetaxel-induced cell death and apoptosis in PCa were determined by MTT and flow cytometry assay. The role of PrLZ on the regulation of autophagy and liver kinase B1/AMP-activated protein kinase (LKB1/AMPK) signaling pathway was analyzed using immunoblotting, immunoprecipitation, siRNA silencing and plasmid overexpression. Results: PrLZ increased docetaxel-mediated drug resistance both in vitro and in vivo. Mechanistic dissection revealed that PrLZ interacted with LKB1 and further inhibited the activation of LKB1/AMPK signals, which negatively contributed to the induction of autophagy. Moreover, PrLZ/LKB1-mediated autophagy conferred resistance to docetaxel-induced cell death and apoptosis both in vitro and in vivo. Conclusion: These findings identify a novel role of PrLZ in autophagy manipulation and provide new insight into docetaxel chemoresistance in PCa, suggesting a new strategy for treating mCRPC by targeting this newly identified signaling pathway.
Project description:<h4>Aim</h4>To investigate whether microRNA-21 was involved in mediating the chemoresistance of prostate cancer cells to docetaxel.<h4>Methods</h4>A microarray technique was used to determine the miRNA profile in docetaxel-resistant PC3 cells. Real-time PCR was used to confirm the array results. miR-21 mimics and inhibitors were synthesized and introduced to cells using Lipofectamine 2000. Cell proliferation was examined with the CCK-8 assay. Luciferase reporter containing PDCD 3'UTR was constructed and the activity was detected by a dual luciferase assay. PDCD4 protein expression was evaluated using Western blot.<h4>Results</h4>A docetaxel-resistant prostate cancer PC3 cell line (PC3R) was established . Using microarrays, miR-21 was found to be up-regulated in PC3R cells. Ectopic expression of miR-21 increased the resistance to docetaxel in PC3 wild type cells. In contrast, silencing of miR-21 in PC3R cells sensitized the cells to docetaxel. The IC(50) values for miR-21-silencing cells and control cells were 28.31 and 35.89 nmol/L, respectively. PDCD4, a direct target gene of miR-21, could mediate chemoresistance to docetaxel in PC3 cells.<h4>Conclusion</h4>Our findings suggest that miR-21 contributed to the resistance of PC3 cells to docetaxel, and that targeting miR-21 may offer a promising therapeutic approach in sensitizing prostate cancer to docetaxel treatment.