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siRNA Oligonucleotides (sirna + oligonucleotide)
Selected AbstractsADAM8 expression is associated with increased invasiveness and reduced patient survival in pancreatic cancerJOURNAL OF CELLULAR AND MOLECULAR MEDICINE, Issue 5 2007N. Valkovskaya Abstract ADAM8 belongs to a family of transmembrane proteins implicated in cell,cell interactions, proteolysis of membrane proteins, and various aspects of carcinogenesis. In the present study, we aimed to evaluate the expression and function of ADAM8 in pancreatic cancer. ADAM8 mRNA levels were analysed by quantitative RT-PCR and correlated to patient survival. Immunohistochemistry was performed to localize ADAM8 in pancreatic tis-sues. Silencing of ADAM8 expression was carried out by transfection with specific siRNA oligonucleotides. Cell growth and invasion assays were used to assess the functional consequences of ADAM8 silencing. SELDI-TOF-MS was performed to detect the proteolytic activity of ADAM8 in pancreatic cancer cells. ADAM8 mRNA was significantly overexpressed in pancreatic ductal adenocarcinoma (PDAC) compared with normal pancreatic tissues (5.3-fold increase; P= 0.0008), and high ADAM8 mRNA and protein expression levels correlated with reduced survival time of PDAC patients (P= 0.048 and P= 0.065, respectively). Silencing of ADAM8 expression did not significantly influence pancreatic cancer cell growth but suppressed invasiveness. In addition, decreased proteolytic activity was measured in cell culture supernatants following silencing of ADAM8. In conclusion, ADAM8 is overexpressed in PDAC, influences cancer cell invasiveness and correlates with reduced survival, suggesting that ADAM8 might be a potential target in pancreatic cancer therapy. [source] Expression of heregulin by mouse mammary tumor cells: Role in activation of ErbB receptors,MOLECULAR CARCINOGENESIS, Issue 7 2006M. Schmitt Abstract The inappropriate activation of one or more members of the ErbB family of receptor tyrosine kinases [ErbB-1 (EGFR), ErbB-2, ErbB-3, ErbB-4] has been linked with oncogenesis. ErbB-2 is frequently coexpressed with ErbB-3 in breast cancer cells and in the presence of the ligand heregulin (HRG) the ErbB-2/ErbB-3 receptors form a signaling heterodimer that can affect cell proliferation and apoptosis. The major goal of the present study was to determine whether endogenous HRG causes autocrine/paracrine activation of ErbB-2/ErbB-3 and contributes to the proliferation of mammary epithelial tumor cells. Tyrosine-phosphorylated (activated) ErbB-2 and ErbB-3 receptors were detected in the majority of extracts from tumors that had formed spontaneously or as a result of oncogene expression. HRG-1 transcripts and protein were found in the epithelial cells of most of these mouse mammary tumors. Various mouse mammary cell lines also contained activated ErbB-2/ErbB-3 and HRG transcripts. A ,50 kDa C-terminal fragment of pro-HRG was detected, which indicates that the HRG-1 precursor is readily processed by these cells. It is likely that the secreted mature HRG activated the ErbB-2/3 receptors. Addition of an antiserum against HRG to the mammary epithelial tumor cell line TM-6 reduced ErbB-3 Tyr-phosphorylation. Treatment with HRG-1 siRNA oligonucleotides or infection with a retroviral construct to stably express HRG siRNA effectively reduced HRG protein levels, ErbB-2/ErbB-3 activation, and the rate of proliferation, which could be reversed by the addition of HRG. The cumulative findings from these experiments show that coexpression of the HRG ligand contributes to activation of ErbB-2/Erb-3 in mouse mammary tumor cells in an autocrine or paracrine fashion. Published 2006 Wiley-Liss, Inc. [source] Inhibition of Aurora Kinase A enhances chemosensitivity of medulloblastoma cell lines,PEDIATRIC BLOOD & CANCER, Issue 1 2010Ayman El-Sheikh MD Abstract Background Medulloblastoma comprises approximately 20% of all primary pediatric brain tumors. Despite recent advances, the survival rate for high-risk patients and the morbidity associated with these treatments remains suboptimal. To improve outcomes and decrease morbidity, more targeted therapy is required. One possible target is the Aurora Kinase family. The objective of this study was to evaluate the impact of Aurora Kinase A inhibition in medulloblastoma cell lines. Procedure Cell proliferation was measured using an MTS assay after adding an Aurora Kinase inhibitor (C1368) at different concentrations. Cell cycle analysis was carried out by Flow Cytometry using propidium iodide (PI). RNAi experiments were performed using siRNA oligonucleotides. Luciferase experiments were carried out using the Cignal Finder 10 Pathway Reporter Arrays. Results Inhibition of Aurora Kinase A induces cell death in medulloblastoma cells and lowers the IC50 of other chemotherapeutic agents (etoposide and cisplatin) used in medulloblastoma treatment. Cell arrest at G2/M phase was significantly increased in medulloblastoma cell lines treated with C1368 Sigma at IC30 or transfected siRNA. Inhibition of Aurora Kinase A resulted in decreased activity of pro-proliferative signaling pathways including Wnt, Myc, and RB as measured by luciferase reporter assays. Conclusions These data indicate that inhibition of Aurora Kinase A inhibits cell growth in medulloblastoma through inhibition of pro-proliferative signaling pathways Wnt, Myc, and RB. Additionally, combining Aurora Kinase A inhibition with other chemotherapeutic agents significantly lowers their IC50, which make it a promising small molecule target for medulloblastoma therapy. Pediatr Blood Cancer 2010;55:35,41. © 2010 Wiley-Liss, Inc. [source] Downmodulation of Bcl-2 sensitizes metastatic LNCaP-LN3 cells to undergo apoptosis via the intrinsic pathwayTHE PROSTATE, Issue 6 2010Renduo Song Abstract BACKGROUND We explored the mechanisms of apoptosis after Bcl-2 protein downmodulation in metastatic LNCaP-LN3 cells (LN3). METHODS LNCaP, LNCaP-Pro5 (Pro5) and LN3 cells were cultured in 5% charcoal-stripped serum (CSS) or in R1881 (synthetic androgen) and bicalutamide (synthetic anti-androgen) and growth inhibition was assessed. Expression levels of androgen receptor (AR) and Bcl-2 were determined. LN3 cells were transfected with small interfering RNA Bcl-2 (siRNA Bcl-2) or control siRNA oligonucleotides. Rates of apoptosis and proliferation were obtained. Cytochrome c localization in treated and control cells was assessed,±,cyclosporine A (CsA). Caspases 9, 3, and poly (ADP-ribose) polymerase cleavage (PARP) were measured upon downmodulation of Bcl-2; and cell growth inhibition in vitro after Bcl-2 modulation combined with docetaxel chemotherapy was determined. RESULTS LN3 cells maintained growth under castrate conditions in vitro. AR protein amplification did not explain castrate-resistant LN3 cell growth. Bcl-2 protein levels in LN3 cells were significantly higher than in Pro5 cells, and were effectively downmodulated by siRNA Bcl-2. Subsequently increased apoptosis and decreased proliferation mediated by cytochrome c was noted and this was reversed by CsA. siRNA Bcl-2-transfected LN3 cells exhibited elevated levels of caspases 9, 3, and PARP cleavage. Exposure of LN3 cells to docetaxel led to increased apoptosis, and simultaneous downmodulation of Bcl-2 substantially enhanced this effect. CONCLUSIONS Downmodulation of Bcl-2 in metastatic castrate-resistant LNCaP-LN3 cells led to apoptosis via a cytochrome c -dependent pathway that was enhanced with docetaxel treatment. Prostate 70: 571,583, 2010. © 2009 Wiley-Liss, Inc. [source] | ||||||||||