cx-4945 and Lung-Neoplasms

Tumor microenvironment (TME) plays a vital role in determining the outcomes of radiotherapy. As an important component of TME, vascular endothelial cells are involved in the perivascular resistance niche (PVRN), which is formed by inflammation or cytokine production induced by ionizing radiation (IR). Protein kinase CK2 is a constitutively active serine/threonine kinase which plays a vital role in cell proliferation and inflammation. In this study, we investigated the potential role of CK2 in PVRN after IR exposure.. Specific CK2 inhibitors, Quinalizarin and CX-4945, were employed to effectively suppressed the kinase activity of CK2 in human umbilical vein endothelial cells (HUVECs) without affecting their viability. Results showing that conditioned medium from IR-exposed HUVECs increased cell viability of A549 and H460 cells, and the pretreatment of CK2 inhibitors slowed down such increment. The secretion of IL-8 and IL-6 in HUVECs was induced after exposure with IR, but significantly inhibited by the addition of CK2 inhibitors. Furthermore, IR exposure elevated the nuclear phosphorylated factor-κB (NF-κB) p65 expression in HUVECs, which was a master factor regulating cytokine production. But when pretreated with CK2 inhibitors, such elevation was significantly suppressed.. This study indicated that protein kinase CK2 is involved in the key process of the IR induced perivascular resistant niche, namely cytokine production, by endothelial cells, which finally led to radioresistance of non-small cell lung cancer cells. Thus, the inhibition of CK2 may be a promising way to improve the outcomes of radiation in non-small cell lung cancer cells.

Topics: Anthraquinones; Blotting, Western; Carcinoma, Non-Small-Cell Lung; Casein Kinase II; Cytokines; Endothelial Cells; Endothelium, Vascular; Humans; Lung Neoplasms; Naphthyridines; Phenazines; Protein Kinase Inhibitors

Protein kinase CK2 is a ubiquitously expressed kinase in eukaryotes, which is known to phosphorylate many protein substrates. Because CK2 is involved in the regulation of various signaling pathways, we wondered whether CK2 participated in the regulation of ionizing radiation (IR) induced biological process. In this study, we investigated the effect of IR on the subcellular localization and kinase activity in human non-small cell lung cancer (NSCLC) cells. Immunofluorescent results showed that CK2 subunits shuttle into the nucleus mostly beginning 1 h after IR and lasting more than 6 h. We also conducted in vitro kinase assay and observed an increase in CK2 kinase activity at 6 h after IR. Furthermore, an increase in S phase was observed at 6 h after IR. Colony formation assay results demonstrated that CK2 inhibitor CX-4945 significantly enhanced the effect of irradiation in NSCLC cells. These results indicated that CK2 may be implicated in the regulation of IR-induced biological process.

Topics: Carcinoma, Non-Small-Cell Lung; Casein Kinase II; Cell Cycle; Cell Line, Tumor; Cell Nucleus; Cell Survival; Cytoplasm; Cytosol; Gene Expression Regulation, Enzymologic; Gene Expression Regulation, Neoplastic; Genotype; Humans; Lung Neoplasms; Naphthyridines; Phenazines; Phosphorylation; Radiation, Ionizing; Signal Transduction

Kinase inhibitors are important cancer therapeutics. Polypharmacology is commonly observed, requiring thorough target deconvolution to understand drug mechanism of action. Using chemical proteomics, we analyzed the target spectrum of 243 clinically evaluated kinase drugs. The data revealed previously unknown targets for established drugs, offered a perspective on the "druggable" kinome, highlighted (non)kinase off-targets, and suggested potential therapeutic applications. Integration of phosphoproteomic data refined drug-affected pathways, identified response markers, and strengthened rationale for combination treatments. We exemplify translational value by discovering SIK2 (salt-inducible kinase 2) inhibitors that modulate cytokine production in primary cells, by identifying drugs against the lung cancer survival marker MELK (maternal embryonic leucine zipper kinase), and by repurposing cabozantinib to treat FLT3-ITD-positive acute myeloid leukemia. This resource, available via the ProteomicsDB database, should facilitate basic, clinical, and drug discovery research and aid clinical decision-making.

Topics: Animals; Antineoplastic Agents; Cell Line, Tumor; Cytokines; Drug Discovery; fms-Like Tyrosine Kinase 3; Humans; Leukemia, Myeloid, Acute; Lung Neoplasms; Mice; Molecular Targeted Therapy; Protein Kinase Inhibitors; Protein Serine-Threonine Kinases; Proteomics; Xenograft Model Antitumor Assays

The response to chemotherapeutic drugs in non-small cell lung cancer (NSCLC) is unsatisfactory, leading to poor outcomes. This study the aimed to investigates anticancer effects of CX-4945, a potent casein kinase II (CK2) inhibitor, in chemorefractory NSCLC cells.. Cell proliferation and apoptosis assay were carried-out by annexin V-FITC and FACScan after drug treatment with paclitaxel, cisplatin and CX-4945. AKT/mTOR and CK2α signals were measured by western blotting. Treatment was carried-out using siRNA to inhibit CK2α.. Paclitaxel, and cisplatin effectively inhibited cell proliferation and induced apoptosis in A549 cells, while not in H1299, Calu-1 and H358 cells. In these chemorefractory cell lines, AKT signalling was maintained despite drug treatment. However, CX-4945 suppressed cell growth, with cell-cycle arrest at G2/M phase and induced apoptosis with an increase of cleaved caspase-3 and PARP1 in a dose-dependent manner. Accordingly, AKT and its downstream signals such as mTOR and p70S6K were down-regulated by CX-4945. Transfection of CK2α siRNA had similar effects to CX-4945 treatment on cell proliferation and apoptosis.. CX-4945 shows a promising anticancer action through down-regulation of AKT/mTOR signals, suggesting its possible application for treatment of chemorefractory lung cancer.

Topics: Apoptosis; Carcinoma, Non-Small-Cell Lung; Casein Kinase II; Cell Cycle Checkpoints; Cell Line, Tumor; Cisplatin; Down-Regulation; Humans; Lung Neoplasms; Naphthyridines; Paclitaxel; Phenazines; Proto-Oncogene Proteins c-akt; TOR Serine-Threonine Kinases

Here, we demonstrate the possible applications of micropillar arrays in screening anti-metastasis drugs. Human lung adenocarcinoma A549 cells incubated in multiwell plates containing micropillars exhibited markedly different physical/biochemical behavior depending on pillar dimensions. In particular, A549 cells grown in plates containing 2-μm diameter, 16-μm pitched pillar arrays showed epithelial-to-mesenchymal transition (EMT)-like behavior; cell body elongation, and highly increased activation of the focal adhesion kinase (FAK)-Src-paxillin signaling cascade. FAK is the most prominent kinase involved in dynamic regulation of the actin cytoskeleton and cell adhesion, migration, and invasion. Activation of FAK, a hallmark of cancer cell adhesion and migration, is normally induced by various growth factors, such as transforming growth factor-β (TGF-β). Here, we found that pillar-mediated activation of signaling molecules mimicked that induced by TGF-β. Notably, micropillar arrays with specific dimensions accelerated the elongation of cells, an effect linked to the activation of signaling molecules related to EMT. Micropillar-induced FAK activation could be arrested by the casein kinase-2 (CK2) inhibitor CX-4945, a drug candidate with activity against TGF-β-induced cancer cell metastasis, demonstrating the possibility of using inorganic microstructures for cell-based drug screening.. In this work, we have fabricated flexible substrates with regular arrays of micrometersized pillars, and used them to grow A549 human lung adenocarcinoma cells. Cells exhibit dramatically different behavior depending on the intervals of pillars. Especially, cells grown in certain pillar structures show epithelial-to mesenchmal transition (EMT)-like morphology and related molecules, which is similar to the activation obtained using expensive cytokine TGF-β. Based on the fact that pillar arrays may activate EMT like transition, screening of anti-cancer drug using pillar arrays have demonstrated as well in our work. Our study confirms that mechanical stimulation may exert similar effects with chemical stimulation, and such mechanical structures could be used as a large-scale drug screening platforms. Cell morphogenesis on engineered substrate is not new, but the present work could be distinguished with its unique fabrication process that can mass produce the structures and it could be applied for high-throughput drug screening. Also, we suggest the formation of focal adhesions on pillar structures and consequent strain as the possible mechanism behind the observed EMT-like transition. Currently, we are working on full-scale profiling of metabolomics and proteomics of cells grown in large-scale pillar arrays as well.

Topics: Apoptosis; Biological Assay; Casein Kinase II; Cell Line, Tumor; Cell Survival; Dose-Response Relationship, Drug; Drug Screening Assays, Antitumor; Enzyme Activation; Equipment Design; Equipment Failure Analysis; Focal Adhesion Kinase 1; Humans; Lung Neoplasms; Mechanotransduction, Cellular; Naphthyridines; Paxillin; Phenazines; src-Family Kinases; Tissue Array Analysis; Treatment Outcome

Protein kinase CK2 has diverse functions promoting and maintaining cancer phenotypes. We investigated the effect of CK2 inhibition in lung cancer cells with T790M-mediated resistance to the EGFR-TK inhibitor. Resistant sublines of PC-9 to gefitinib (PC-9/GR) and erlotinib (PC-9/ER) were established by previous study, and T790M secondary mutation was found in both resistant sublines. A decrease of EGFR by siRNA treatment effectively controlled the growth of resistant cells, thus suggesting that they still have EGFR-dependency. CX-4945, a potent and selective CK2 inhibitor, induced autophagy in PC-9/GR and PC-9/ER, and which was supported by the induction of autophagic vacuoles and microtubule-associated protein 1 light chain 3 (LC3) expression, and the increase of punctate fluorescent signals in resistant cells pre-transfected with green fluorescent protein (GFP)-tagged LC3. However, the withdrawal of CX-4945 led to the recovery of cancer cells with autophagy. We found that the induction of autophagy by CX-4945 in both resistant cells was CK2 dependent by using small interfering RNA against CK2. The treatment with CX-4945 alone induced a minimal growth inhibition in resistant cells. However, combined treatment of CX-4945 and EGFR-TKI effectively inhibited cancer-cell proliferation and induced apoptosis. CX-4945 increased the translocation of EGFR from the cell surface into the autophagosome, subsequently leading to the decrease of EGFR while inhibition of autophagy by 3MA or Atg7-targeted siRNA pretreatment reduced the decrease of EGFR by CX-4945. Accordingly, apoptosis by a combination of CX-4945 and EGFR-TKI was suppressed by 3MA or Atg7-targeted siRNA pretreatment, thus suggesting that autophagosome-mediated EGFR down-regulation would have an important role regarding apoptotic cell death by EGFR-TKI. Combined treatment of the CK2 inhibitor and EGFR-TKI may be a promising strategy for overcoming T790M-mediated resistance.

Topics: Apoptosis; Autophagy; Casein Kinase II; Cell Line, Tumor; Drug Resistance, Neoplasm; ErbB Receptors; Erlotinib Hydrochloride; Gefitinib; Gene Expression Regulation, Neoplastic; Humans; Lung Neoplasms; Mutation; Naphthyridines; Phagosomes; Phenazines; Protein Kinase Inhibitors; Quinazolines

Casein kinase 2 (CK2) is known to be involved in various cellular processes such as cell cycle, apoptosis and proliferation. It has been reported that the inhibition of CK2 induced by recently developed small molecule CX4945 shows anti-cancer effects including anti-proliferation and anti-angiogenesis in several different cancers including prostate cancer. Here we report that migration and invasion of A549 human lung cancer cells are suppressed by the inhibition of CK2 induced by CX4945. We found that CX4945 sequentially attenuates the proteins in PI3K/Akt and MAPK pathways, two signaling pathways related with cell migration. This sequential control of signal pathways inhibits the expression of membrane type 1-matrix metalloproteinase and this leads to the selective attenuation of one of the gelatinases, MMP-2, which can degrade components of extracellular matrix, and metastasis of A549 human lung cancer cell.

Topics: Antineoplastic Agents; Casein Kinase II; Cell Line, Tumor; Cell Movement; Cell Survival; Gelatinases; Humans; Lung Neoplasms; Matrix Metalloproteinase 1; Matrix Metalloproteinase 2; Mitogen-Activated Protein Kinase Kinases; Naphthyridines; Phenazines; Phosphatidylinositol 3-Kinases; Proto-Oncogene Proteins c-akt; Signal Transduction

Protein kinase CK2 is frequently elevated in a variety of human cancers. The Notch1 signalling pathway has been implicated in stem cell maintenance and its aberrant activation has been shown in several types of cancer including lung cancer. Here, we show, for the first time, that CK2α is a positive regulator of Notch1 signalling in lung cancer cell lines A549 and H1299. We found that Notch1 protein level was reduced after CK2α silencing. Down-regulation of Notch1 transcriptional activity was demonstrated after the silencing of CK2α in lung cancer cells. Furthermore, small-molecule CK2α inhibitor CX-4945 led to a dose-dependent inhibition of Notch1 transcriptional activity. Conversely, forced overexpression of CK2α resulted in an increase in Notch1 transcriptional activity. Finally, the inhibition of CK2α led to a reduced proportion of stem-like CD44 + /CD24- cell population. Thus, we report that the inhibition of CK2α down-regulates Notch1 signalling and subsequently reduces a cancer stem-like cell population in human lung cancer cells. Our data suggest that CK2α inhibitors may be beneficial to the lung cancer patients with activated Notch1 signalling.

Topics: Casein Kinase II; CD24 Antigen; Cell Line, Tumor; DNA; Dose-Response Relationship, Drug; Down-Regulation; Flow Cytometry; Gene Expression Regulation, Neoplastic; Humans; Hyaluronan Receptors; Immunohistochemistry; Lung Neoplasms; Naphthyridines; Neoplastic Stem Cells; Phenazines; Phenotype; Receptor, Notch1; RNA, Small Interfering; Signal Transduction; Transcription, Genetic

The epithelial-to-mesenchymal transition (EMT) is a major phenotype of cancer metastasis and invasion. As a druggable cancer target, the inhibition of protein kinase CK2 (formally named to casein kinase 2) has been suggested as a promising therapeutic strategy to treat EMT-controlled cancer metastasis. This study aimed to evaluate the effect of the CK2 inhibitor CX-4945 on the processes of cancer migration and invasion during the EMT in A549 human lung adenocarcinoma cells.. The effect of CX-4945 on TGF-β1-induced EMT was evaluated in A549 cells treated with TGF-β1 (5 ng/ml) and CX-4945. The effect of CX-4945 on TGF-β1-induced cadherin switch and activation of key signaling molecules involved in Smad, non-Smad, Wnt and focal adhesion signaling pathways were investigated by Western blot analysis, immunocytochemistry and reporter assay. Additionally, the effect of CX-4945 on TGF-β1-induced migration and invasion was investigated by wound healing assay, Boyden chamber assay, gelatin zymography, and the quantitative real-time PCR.. CX-4945 inhibits the TGF-β1-induced cadherin switch and the activation of key signaling molecules involved in Smad (Smad2/3, Twist and Snail), non-Smad (Akt and Erk), Wnt (β-catenin) and focal adhesion signaling pathways (FAK, Src and paxillin) that cooperatively regulate the overall process of EMT. As a result, CX-4945 inhibits the migration and invasion of A549 cells accompanied with the downregulation of MMP-2 and 9.. Clinical evaluation of CX-4945 in humans as a single agent in solid tumors and multiple myeloma has established its promising pharmacokinetic, pharmacodynamic, and safety profiles. Beyond regression of tumor mass, CX-4945 may be advanced as a new therapy for cancer metastasis and EMT-related disorders.

Topics: Adenocarcinoma; Adenocarcinoma of Lung; Cadherins; Casein Kinase II; Cell Line, Tumor; Cell Movement; Epithelial-Mesenchymal Transition; Focal Adhesion Kinase 1; Humans; Lung Neoplasms; Naphthyridines; Neoplasm Invasiveness; Paxillin; Phenazines; Protein Kinase Inhibitors; Signal Transduction; Smad Proteins; Transforming Growth Factor beta1; Wnt Proteins