Toll-like receptor 3 (TLR3) is an endosomal receptor expressed in several immune and epithelial cells. Recent studies have highlighted its expression also in solid tumors, including prostate cancer (PCa), and have described its role primarily in the proinflammatory response and induction of apoptosis. It is up-regulated in some castration-resistant prostate cancers. However, the role of TLR3 in prostate cancer progression remains largely unknown. The current study experimentally demonstrated that exogenous TLR3 activation in PCa cell lines leads to a significant induction of secretion of the cytokines IL-6, IL-8, and interferon-β, depending on the model and chemoresistance status. Transcriptomic analysis of TLR3-overexpressing cells revealed a functional program that is enriched for genes involved in the regulation of cell motility, migration, and tumor invasiveness. Increased motility, migration, and invasion in TLR3-overexpressing cell line were confirmed by several in vitro assays and using an orthotopic prostate xenograft model in vivo. Furthermore, TLR3-ligand induced apoptosis via cleavage of caspase-3/7 and poly (ADP-ribose) polymerase, predominantly in TLR3-overexpressing cells. These results indicate that TLR3 may be involved in prostate cancer progression and metastasis; however, it might also represent an Achilles heel of PCa, which can be exploited for targeted therapy.

• MeSH
• apoptóza MeSH
• lidé MeSH
• nádorové buněčné linie MeSH
• nádory prostaty * patologie   MeSH
• poly I-C farmakologie   MeSH
• prostata patologie   MeSH
• toll-like receptor 3 * genetika   metabolismus   MeSH
• zvířata MeSH
• Check Tag
• lidé MeSH
• mužské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

The transcription factor c-Myb is an oncoprotein promoting cell proliferation and survival when aberrantly activated/expressed, thus contributing to malignant transformation. Overexpression of c-Myb has been found in leukemias, breast, colon and adenoid cystic carcinoma. Recent studies revealed its expression also in osteosarcoma cell lines and suggested its functional importance during bone development. However, the relevance of c-Myb in control of osteosarcoma progression remains unknown. A retrospective clinical study was carried out to assess a relationship between c-Myb expression in archival osteosarcoma tissues and prognosis in a cohort of high-grade osteosarcoma patients. In addition, MYB was depleted in metastatic osteosarcoma cell lines SAOS-2 LM5 and 143B and their growth, chemosensitivity, migration and metastatic activity were determined. Immunohistochemical analysis revealed that high c-Myb expression was significantly associated with poor overall survival in the cohort and metastatic progression in young patients. Increased level of c-Myb was detected in metastatic osteosarcoma cell lines and its depletion suppressed their growth, colony-forming capacity, migration and chemoresistance in vitro in a cell line-dependent manner. MYB knock-out resulted in reduced metastatic activity of both SAOS-2 LM5 and 143B cell lines in immunodeficient mice. Transcriptomic analysis revealed the c-Myb-driven functional programs enriched for genes involved in the regulation of cell growth, stress response, cell adhesion and cell differentiation/morphogenesis. Wnt signaling pathway was identified as c-Myb target in osteosarcoma cells. Taken together, we identified c-Myb as a negative prognostic factor in osteosarcoma and showed its involvement in the regulation of osteosarcoma cell growth, chemosensitivity, migration and metastatic activity.

• MeSH
• lidé MeSH
• myši MeSH
• nádorové buněčné linie MeSH
• nádory kostí * patologie   MeSH
• osteosarkom * patologie   MeSH
• pohyb buněk genetika   MeSH
• prognóza MeSH
• proliferace buněk MeSH
• regulace genové exprese u nádorů MeSH
• retrospektivní studie MeSH
• signální dráha Wnt MeSH
• zvířata MeSH
• Check Tag
• lidé MeSH
• myši MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

As treatment options for patients with incurable metastatic castration-resistant prostate cancer (mCRPC) are considerably limited, novel effective therapeutic options are needed. Checkpoint kinase 1 (CHK1) is a highly conserved protein kinase implicated in the DNA damage response (DDR) pathway that prevents the accumulation of DNA damage and controls regular genome duplication. CHK1 has been associated with prostate cancer (PCa) induction, progression, and lethality; hence, CHK1 inhibitors SCH900776 (also known as MK-8776) and the more effective SCH900776 analog MU380 may have clinical applications in the therapy of PCa. Synergistic induction of DNA damage with CHK1 inhibition represents a promising therapeutic approach that has been tested in many types of malignancies, but not in chemoresistant mCRPC. Here, we report that such therapeutic approach may be exploited using the synergistic action of the antimetabolite gemcitabine (GEM) and CHK1 inhibitors SCH900776 and MU380 in docetaxel-resistant (DR) mCRPC. Given the results, both CHK1 inhibitors significantly potentiated the sensitivity to GEM in a panel of chemo-naïve and matched DR PCa cell lines under 2D conditions. MU380 exhibited a stronger synergistic effect with GEM than clinical candidate SCH900776. MU380 alone or in combination with GEM significantly reduced spheroid size and increased apoptosis in all patient-derived xenograft 3D cultures, with a higher impact in DR models. Combined treatment induced premature mitosis from G1 phase resulting in the mitotic catastrophe as a prestage of apoptosis. Finally, treatment by MU380 alone, or in combination with GEM, significantly inhibited tumor growth of both PC339-DOC and PC346C-DOC xenograft models in mice. Taken together, our data suggest that metabolically robust and selective CHK1 inhibitor MU380 can bypass docetaxel resistance and improve the effectiveness of GEM in DR mCRPC models. This approach might allow for dose reduction of GEM and thereby minimize undesired toxicity and may represent a therapeutic option for patients with incurable DR mCRPC.

• MeSH
• buněčná smrt účinky léků   MeSH
• checkpoint kinasa 1 antagonisté a inhibitory   metabolismus   MeSH
• chemorezistence účinky léků   MeSH
• deoxycytidin analogy a deriváty   farmakologie   MeSH
• docetaxel farmakologie   MeSH
• lidé MeSH
• mitóza * účinky léků   MeSH
• myši SCID MeSH
• nádorové buněčné linie MeSH
• nádory prostaty patologie   MeSH
• piperidiny chemie   farmakologie   MeSH
• proliferace buněk účinky léků   MeSH
• pyrazoly chemie   farmakologie   MeSH
• pyrimidiny chemie   farmakologie   MeSH
• S fáze účinky léků   MeSH
• xenogenní modely - testy protinádorové aktivity MeSH
• zvířata MeSH
• Check Tag
• lidé MeSH
• mužské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

Skp2 is a crucial component of SCFSkp2 E3 ubiquitin ligase and is often overexpressed in various types of cancer, including prostate cancer (PCa). The epithelial-to-mesenchymal transition (EMT) is involved in PCa progression. The acquisition of a mesenchymal phenotype that results in a cancer stem cell (CSC) phenotype in PCa was described. Therefore, we aimed to investigate the expression and localization of Skp2 in clinical samples from patients with PCa, the association of Skp2 with EMT status, and the role of Skp2 in prostate CSC. We found that nuclear expression of Skp2 was increased in patients with PCa compared to those with benign hyperplasia, and correlated with high Gleason score in PCa patients. Increased Skp2 expression was observed in PCa cell lines with mesenchymal and CSC-like phenotype compared to their epithelial counterparts. Conversely, the CSC-like phenotype was diminished in cells in which SKP2 expression was silenced. Furthermore, we observed that Skp2 downregulation led to the decrease in subpopulation of CD44+CD24- cancer stem-like cells. Finally, we showed that high expression levels of both CD24 and CD44 were associated with favorable recurrence-free survival for PCa patients. This study uncovered the Skp2-mediated CSC-like phenotype with oncogenic functions in PCa.

• MeSH
• antigen CD24 genetika   MeSH
• antigeny CD44 genetika   MeSH
• buňky PC-3 MeSH
• epitelo-mezenchymální tranzice * MeSH
• lidé MeSH
• myši nahé MeSH
• myši MeSH
• nádorové buněčné linie MeSH
• nádorové kmenové buňky metabolismus   fyziologie   MeSH
• nádory prostaty genetika   metabolismus   patofyziologie   MeSH
• proteiny asociované s kinázou S-fáze genetika   MeSH
• regulace genové exprese u nádorů * MeSH
• stupeň nádoru MeSH
• xenogenní modely - testy protinádorové aktivity MeSH
• zvířata MeSH
• Check Tag
• lidé MeSH
• mužské pohlaví MeSH
• myši MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

PURPOSE: The purpose of the study was to determine whether the GDF-15 is present in follicular fluid; to evaluate if there is a relation between follicular and serum levels of GDF-15 and fertility status of study subjects; and to test whether granulosa cells, oocytes, or both produce GDF-15. METHODS: This study used follicular fluid (FF, serum, and oocytes obtained under informed consent from women undergoing oocyte retrieval for in vitro fertilization. It also used ovaries from deceased preterm newborns. Collection of FF and blood at the time of oocyte retrieval, ELISA and western blot were performed to determine levels and forms of GDF-15. Concentrations of GDF-15 in FF and serum, its expression in ovarian tissue, and secretion from granulosa cells were analyzed. RESULTS: GDF-15 concentration in FF ranged from 35 to 572 ng/ml, as determined by ELISA. Western blot analysis revealed the GDF-15 pro-dimer only in FF. Both normal healthy and cancerous granulosa cells secreted GDF-15 into culture media. Primary oocytes displayed cytoplasmic GDF-15 positivity in immunostained newborn ovaries, and its expression was also observed in fully grown human oocytes. CONCLUSIONS: To the best of our knowledge, this is the first documentation of cytokine GDF-15 presence in follicular fluid. Its concentration was not associated with donor/patient fertility status. Our data also show that GDF-15 is expressed and inducible in both normal healthy and cancerous granulosa cells, as well as in oocytes.

• MeSH
• buněčná diferenciace genetika   MeSH
• dospělí MeSH
• fertilizace in vitro MeSH
• folikulární buňky metabolismus   MeSH
• folikulární tekutina metabolismus   MeSH
• lidé MeSH
• odběr oocytu MeSH
• oocyty metabolismus   MeSH
• růstový diferenciační faktor 15 genetika   izolace a purifikace   MeSH
• vývojová regulace genové exprese MeSH
• Check Tag
• dospělí MeSH
• lidé MeSH
• ženské pohlaví MeSH
• Publikační typ
• časopisecké články MeSH

Checkpoint-mediated dependency of tumor cells can be deployed to selectively kill them without substantial toxicity to normal cells. Specifically, loss of CHK1, a serine threonine kinase involved in the surveillance of the G2-M checkpoint in the presence of replication stress inflicted by DNA-damaging drugs, has been reported to dramatically influence the viability of tumor cells. CHK1's pivotal role in maintaining genomic stability offers attractive opportunity for increasing the selectivity, effectivity, and reduced toxicity of chemotherapy. Some recently identified CHK1 inhibitors entered clinical trials in combination with DNA antimetabolites. Herein, we report synthesis and profiling of MU380, a nontrivial analogue of clinically profiled compound SCH900776 possessing the highly unusual N-trifluoromethylpyrazole motif, which was envisioned not to undergo metabolic oxidative dealkylation and thereby provide greater robustness to the compound. MU380 is a selective and potent inhibitor of CHK1 which sensitizes a variety of tumor cell lines to hydroxyurea or gemcitabine up to 10 times. MU380 shows extended inhibitory effects in cells, and unlike SCH900776, does not undergo in vivo N-dealkylation to the significantly less selective metabolite. Compared with SCH900776, MU380 in combination with GEM causes higher accumulation of DNA damage in tumor cells and subsequent enhanced cell death, and is more efficacious in the A2780 xenograft mouse model. Overall, MU380 represents a novel state-of-the-art CHK1 inhibitor with high potency, selectivity, and improved metabolic robustness to oxidative N-dealkylation. Mol Cancer Ther; 16(9); 1831-42. ©2017 AACR.

• MeSH
• apoptóza účinky léků   MeSH
• biologické markery MeSH
• buněčný cyklus účinky léků   MeSH
• checkpoint kinasa 1 antagonisté a inhibitory   MeSH
• chemorezistence účinky léků   MeSH
• dealkylace účinky léků   MeSH
• inhibitory proteinkinas chemická syntéza   farmakologie   MeSH
• kontrolní body buněčného cyklu účinky léků   MeSH
• lidé MeSH
• metylace MeSH
• modely nemocí na zvířatech MeSH
• molekulární struktura MeSH
• myši MeSH
• nádorové buněčné linie MeSH
• protinádorové látky chemická syntéza   farmakologie   MeSH
• pyrazoly farmakologie   MeSH
• pyrimidiny farmakologie   MeSH
• vztah mezi dávkou a účinkem léčiva MeSH
• xenogenní modely - testy protinádorové aktivity MeSH
• zvířata MeSH
• Check Tag
• lidé MeSH
• myši MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH

Lung cancer is the leading cause of cancer deaths, and effective treatments are urgently needed. Loss-of-function mutations in the DNA damage response kinase ATM are common in lung adenocarcinoma but directly targeting these with drugs remains challenging. Here we report that ATM loss-of-function is synthetic lethal with drugs inhibiting the central growth factor kinases MEK1/2, including the FDA-approved drug trametinib. Lung cancer cells resistant to MEK inhibition become highly sensitive upon loss of ATM both in vitro and in vivo. Mechanistically, ATM mediates crosstalk between the prosurvival MEK/ERK and AKT/mTOR pathways. ATM loss also enhances the sensitivity of KRAS- or BRAF-mutant lung cancer cells to MEK inhibition. Thus, ATM mutational status in lung cancer is a mechanistic biomarker for MEK inhibitor response, which may improve patient stratification and extend the applicability of these drugs beyond RAS and BRAF mutant tumours.

• MeSH
• ATM protein genetika   metabolismus   MeSH
• benzamidy farmakologie   MeSH
• difenylamin analogy a deriváty   farmakologie   MeSH
• inhibitory proteinkinas farmakologie   MeSH
• lidé MeSH
• močovina analogy a deriváty   farmakologie   MeSH
• mutace * MeSH
• myši nahé MeSH
• nádorové buněčné linie MeSH
• nádory plic genetika   metabolismus   prevence a kontrola   MeSH
• proliferace buněk účinky léků   genetika   MeSH
• protoonkogenní proteiny B-raf genetika   metabolismus   MeSH
• pyridony farmakologie   MeSH
• pyrimidinony farmakologie   MeSH
• ras proteiny genetika   metabolismus   MeSH
• RNA interference MeSH
• thiofeny farmakologie   MeSH
• xenogenní modely - testy protinádorové aktivity MeSH
• zvířata MeSH
• Check Tag
• lidé MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

Plasticity of cancer cells, manifested by transitions between epithelial and mesenchymal phenotypes, represents a challenging issue in the treatment of neoplasias. Both epithelial-mesenchymal transition (EMT) and mesenchymal-epithelial transition (MET) are implicated in the processes of metastasis formation and acquisition of stem cell-like properties. Mouse double minute (MDM) 2 and MDMX are important players in cancer progression, as they act as regulators of p53, but their function in EMT and metastasis may be contradictory. Here, we show that the EMT phenotype in multiple cellular models and in clinical prostate and breast cancer samples is associated with a decrease in MDM2 and increase in MDMX expression. Modulation of EMT-accompanying changes in MDM2 expression in benign and transformed prostate epithelial cells influences their migration capacity and sensitivity to docetaxel. Analysis of putative mechanisms of MDM2 expression control demonstrates that in the context of defective p53 function, MDM2 expression is regulated by EMT-inducing transcription factors Slug and Twist. These results provide an alternative context-specific role of MDM2 in EMT, cell migration, metastasis, and therapy resistance.

• MeSH
• epitelo-mezenchymální tranzice fyziologie   MeSH
• fenotyp MeSH
• heterografty MeSH
• jaderné proteiny biosyntéza   MeSH
• lidé MeSH
• myši nahé MeSH
• myši MeSH
• nádorové buněčné linie MeSH
• nádory prostaty genetika   metabolismus   patologie   MeSH
• nádory prsu genetika   metabolismus   patologie   MeSH
• protoonkogenní proteiny c-mdm2 biosyntéza   MeSH
• protoonkogenní proteiny biosyntéza   MeSH
• transfekce MeSH
• zvířata MeSH
• Check Tag
• lidé MeSH
• mužské pohlaví MeSH
• myši MeSH
• ženské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

BACKGROUND: Tumor heterogeneity and the plasticity of cancer cells present challenges for effective clinical diagnosis and therapy. Such challenges are epitomized by neuroendocrine transdifferentiation (NED) and the emergence of neuroendocrine-like cancer cells in prostate tumors. This phenomenon frequently arises from androgen-depleted prostate adenocarcinoma and is associated with the development of castration-resistant prostate cancer and poor prognosis. RESULTS: In this study, we showed that NED was evoked in both androgen receptor (AR)-positive and AR-negative prostate epithelial cell lines by growing the cells to a high density. Androgen depletion and high-density cultivation were both associated with cell cycle arrest and deregulated expression of several cell cycle regulators, such as p27Kip1, members of the cyclin D protein family, and Cdk2. Dual inhibition of Cdk1 and Cdk2 using pharmacological inhibitor or RNAi led to modulation of the cell cycle and promotion of NED. We further demonstrated that the cyclic adenosine 3', 5'-monophosphate (cAMP)-mediated pathway is activated in the high-density conditions. Importantly, inhibition of cAMP signaling using a specific inhibitor of adenylate cyclase, MDL-12330A, abolished the promotion of NED by high cell density. CONCLUSIONS: Taken together, our results imply a new relationship between cell cycle attenuation and promotion of NED and suggest high cell density as a trigger for cAMP signaling that can mediate reversible NED in prostate cancer cells.

• MeSH
• AMP cyklický metabolismus   MeSH
• androgenní receptory metabolismus   MeSH
• androgeny farmakologie   MeSH
• cyklin-dependentní kinasa 2 metabolismus   MeSH
• cyklin-dependentní kinasy metabolismus   MeSH
• epitelové buňky účinky léků   enzymologie   patologie   MeSH
• imunohistochemie MeSH
• inhibitory proteinkinas farmakologie   MeSH
• kontrolní body buněčného cyklu účinky léků   MeSH
• lidé MeSH
• nádorové buněčné linie MeSH
• nádory prostaty patologie   MeSH
• neuroendokrinní buňky účinky léků   patologie   MeSH
• počet buněk MeSH
• signální transdukce účinky léků   MeSH
• transdiferenciace buněk * účinky léků   MeSH
• Check Tag
• lidé MeSH
• mužské pohlaví MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

The principles of large screening strategies, which are developed by industrial companies, have been recently adopted by researchers in the fields of molecular biology and oncology as invaluable tools for translational medicine. The declining costs of laboratory robotic machines have allowed high-throughput screening to become more available to academic centres with limited resources. Here, we describe how a robotic conventional liquid handling system could be used on a daily basis in laboratories to obtain consistent and reproducible results. Our approach allowed us to quickly screen a panel of more than 20 tumorigenic and non-tumorigenic cell lines for their responses to hydroxyurea, which is a DNA-damaging anticancer therapeutic drug. The format of 384-well microplates was used for manual cell seeding, and the effect of hydroxyurea was screened at multiple concentrations. The fluorescence-based CyQuant assay was employed as the readout method to analyse the cellular DNA content. The effectiveness of our approach was demonstrated in the experimental results.

• MeSH
• automatizace MeSH
• buněčné kultury metody   MeSH
• hydroxymočovina farmakologie   MeSH
• karcinogeneze účinky léků   patologie   MeSH
• lidé MeSH
• nádorové buněčné linie MeSH
• poškození DNA * MeSH
• protinádorové látky farmakologie   MeSH
• regresní analýza MeSH
• screeningové testy protinádorových léčiv * MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH