ag-490 and Lung-Neoplasms

TRIM32 is a member of the tripartite motif (TRIM) family, which has been associated with tumorigenesis. However, its expression and potential functional role(s) in lung cancer progression have not been fully understood. To evaluate the relationship between the expression of TRIM32 and the prognosis of patients with lung cancer, an independent data set (The Human Protein Atlas website) was introduced. The expression and function analysis of TRIM32 in lung cancer cell lines were also performed by using cell counting kit-8, flow cytometry, transwell, real-time polymerase chain reaction and Western blot analysis. Our data showed that TRIM32 was overexpressed in lung cancer tissues and cell lines and was associated with a poor prognosis. TRIM32 silencing inhibited cell proliferation, migration, invasion, adhesion, and the activation of janus kinase 2 (JAK2)/signal transducer and activator of transcription 3 (STAT3) signaling. The results showed knockdown of TRIM32 in NCI-H446 cells also inhibited cell growth in nude mice in the xenograft model. Additionally, TRIM32 overexpression promoted lung cancer cell proliferation and motility and mediated the expression of Bax, Bcl-2, cleaved caspase-3, matrix metalloproteinase-2 (MMP-2) and MMP-9 were inhibited by JAK2/STAT3 signaling inhibitor (AG490). Taken together, our findings suggest that TRIM32 may regulate lung cancer cell proliferation, apoptosis, and motility through activating the JAK2/STAT3-signaling pathway and may be a novel and promising target for lung cancer.

Topics: Animals; Apoptosis; Carcinogenesis; Cell Adhesion; Cell Line, Tumor; Cell Movement; Cell Proliferation; Gene Expression Regulation, Neoplastic; Gene Silencing; Humans; Janus Kinase 2; Lung Neoplasms; Mice, Nude; Neoplasm Invasiveness; Neoplasm Proteins; Prognosis; Signal Transduction; STAT3 Transcription Factor; Transcription Factors; Tripartite Motif Proteins; Tyrphostins; Ubiquitin-Protein Ligases

To explore AG490 (the inhibitor of Janus kinase (JAK) 2/signal transducer and activator of transcription (STAT) 3 pathway) in cisplatin (DDP)-induced acute kidney injury (AKI) in mice with lung cancer.. Mice were randomly divided into normal, model, AG490, DDP and DDP + AG490 groups. The lung cancer models were established except for Normal group. The levels of blood urea nitrogen (BUN) and creatinine and the status of oxidative stress were detected. Then, histological changes were assessed by HE and PAS staining and apoptosis by TUNEL experiment. The molecule expressions were detected by qRT-PCR and western blot, and immunohistochemistry, respectively.. DDP inhibited the tumor growth in mice with lung cancer, which was further promoted by the combination with AG490. Mice in the DDP group had elevated levels of BUN and creatinine than those in the Normal group with the increased inflammatory cytokines (TNF-α, IL-6, MCP-1 and CXCL-1) and malondialdehyde (MDA) level and the decreased glutathione (GSH), superoxide dismutase (SOD) and catalase (CAT). In addition, DDP could activate the JAK2/STAT3 pathway to promote the apoptosis by upregulating Bax, cleaved caspase-9 and cleaved caspase-3 while downregulating the Bcl-2 in the kidney tissues. DDP + AG490 group showed the alleviated AKI and the improvements in oxidative stress, inflammatory responses and apoptosis in the kidney tissues, as compared to DDP group.. AG490 alleviated DDP-induced AKI in lung cancer mice with improved oxidative stress and inflammation, and the suppression of JAK2/STAT3 pathway.

Topics: Acute Kidney Injury; Animals; Apoptosis; Blood Urea Nitrogen; Carcinoma, Lewis Lung; Cisplatin; Creatinine; Female; Inflammation; Janus Kinase 2; Lung Neoplasms; Male; Mice; Mice, Inbred C57BL; Oxidative Stress; Signal Transduction; STAT3 Transcription Factor; Tyrphostins

Lung cancer, which is typically diagnosed at later stages, is a leading cause of cancer death among both males and females given its highly invasive and rapidly metastasizing nature. Rho GTPase activating protein 15 (ARHGAP15) is a member of the RhoGAP family and functions in multiple biological processes, such as cell proliferation and migration. However, the effect of ARHGAP15 in lung cancer and its underlying molecular mechanisms remain unclear.. In this study, immunohistochemistry and Real Time PCR were performed to detect ARHGAP15 expression in lung cancer tissues and cells. Proliferation, transwell, and Western blot assays were further performed to explore the role and underlying mechanism of ARHGAP15 in lung cancer.. Reduced ARHGAP15 expression was observed in lung cancer tissues and cells. In vitro upregulation of ARHGAP15 in lung cancer cells strongly suppressed cell proliferation, migration, and invasion and was accompanied by reduced matrix metalloproteinase-2 (MMP2), MMP9, vascular endothelial growth factor (VEGF) expression, and the phosphorylation of the signal transducer and activator of transcription-3 (p-STAT3). In contrast, interleukin-6 (IL-6) had the opposite effect and the induction of IL-6 was counteracted by ARHGAP15 upregulation. In addition, the proliferation, migration, and cell invasion induced by ARHGAP15 silencing were potentially inhibited by the STAT3 inhibitor AG490 (100 µM), MMP2, MMP9, VEGF, and p-STAT3 levels decreased.. These results suggest that ARGFAP15 suppressed the proliferation and metastasis of lung cancer cells, which may occur through the inhibition of MMP2, MMP9, and VEGF expression via the STAT3 pathway inactivation.

Topics: Cell Proliferation; Cells, Cultured; GTPase-Activating Proteins; Humans; Lung Neoplasms; Signal Transduction; STAT3 Transcription Factor; Tyrphostins

Interferon-γ (IFN-γ) is conventionally regarded as an inflammatory cytokine that has a pivotal role in anti-infection and tumor immune surveillance. It has been used clinically to treat a variety of malignancies. However, increased evidence has suggested IFN-γ can act to induce tumor progression. The role of IFN-γ in regulating antitumor immunity appears to be complex and paradoxical. The mechanism underlying the dual aspects of IFN-γ function in antitumor immunity is not clear.. (1) Lung cancer cells (A549 cells) were cultured with pleural effusion or supernatant of tumor-associated macrophages (TAMs supernatant), and the expression levels of PD-L1 were detected by flow cytometer. The invasion capacity was measured in vitro using trans-well migration assays. (2) Pleural effusion mononuclear cells (PEMC) were separated by Ficoll Hypaque gradient. The expression of interleukin (IL)-6, IL-10, tumor necrosis factor (TNF)-α, and INF-γ in the tumor-associated macrophages was analyzed by flow cytometry. (3) A549 cells were stimulated with IL-6, IL-10, TNF-α, or IFN-γ and then the expression levels were detected by flow cytometry. (4) The expression levels of phospho-ERK (p-ERK), phospho-AKT (p-AKT), and phospho-Sat3 (p-Stat3) were analyzed with Western blot after stimulation with IFN-γ. (5) Cotreatment of the A549 cells with MAPK/ERK-specific inhibitor PD98059, PI3K/AKT-specific inhibitor LY294002, or JAK/STAT3-specific inhibitor AG490, respectively, blocked IFN-γ-induced PD-L1 expression, and then PD-L1 expression was detected by flow cytometry.. We demonstrated that TAMs could induce the expression of PD-L1 by the secretion of IFN-γ through the Janus kinase/signal transducer and activator of transcription 3 (JAK/STAT3) signaling pathway and the phosphatidylinositol 3-kinase (PI3K)/AKT signaling pathway in A549 cells. Furthermore, the signal pathway blockers LY294002 or AG490 could block the induced expression of PD-L1 by IFN-γ.. IFN-γ was not always successful as an antitumor agent. It also can promote tumor cells to evade immune surveillance. Researchers should be cautious in using IFN-γ as a therapeutic agent for cancer treatment.

Topics: B7-H1 Antigen; Cell Line, Tumor; Flavonoids; Humans; Interferon-gamma; Interleukin-10; Interleukin-6; Janus Kinases; Lung Neoplasms; Macrophages; Phosphatidylinositol 3-Kinases; Protein Kinase Inhibitors; Proto-Oncogene Proteins c-akt; Signal Transduction; STAT3 Transcription Factor; Tumor Necrosis Factor-alpha; Tyrphostins

Epithelial-mesenchymal transition (EMT), a key step in the early stages of cancer metastasis, is orchestrated by several signaling pathways, including IL-6/JAK/STAT3 and TGF-β/Smad signaling. However, an association between the two signaling pathways during the EMT process is largely unknown. Here, we focused on lung cancer and demonstrated that TGF-β1 induced the phosphorylation of Smad3 (p-Smad3), upregulation of Snail, a fibroblast-like morphology, and downregulation of E-cadherin as well as upregulation of vimentin in lung cancer cell lines. SIS3 (an inhibitor of Smad3) suppressed TGF-β1-induced activation of Smad3, upregulation of Snail and the EMT process. Importantly, the JAK2/STAT3-specific inhibitor AG490 blocked Stat3 phosphorylation, resulting in attenuated levels of TGF-β1-induced p-Smad3, Snail, MMP2, and Smad-mediated PAI-1 promoter reporter gene activity in A549 and H1650 cells. Subsequently, AG490 inhibited TGF-β-induced cell migration and invasion. Moreover, exogenous IL-6 treatment stimulated Stat3 activation, enhanced TGF-β-induced expression of p-Smad3 and Snail, aggravated the EMT process, and increased lung cancer cell migration and invasion induced by TGF-β1. Our findings show that the JAK/STAT3 pathway is required for TGF-β-induced EMT and cancer cell migration and invasion via upregulation of the expression of p-Smad3 and Snail, and the IL-6/JAK/STAT3 and TGF-β/Smad signaling synergistically enhance EMT in lung carcinomas. The present study suggests a novel rationale for inhibiting cancer metastasis using anti-IL-6/JAK/STAT3 and anti-TGF-β/Smad therapeutic strategies.

Topics: Antineoplastic Agents; Cell Line, Tumor; Cell Movement; Epithelial-Mesenchymal Transition; Gene Expression Regulation, Neoplastic; Humans; Interleukin-6; Janus Kinases; Lung Neoplasms; Neoplasm Invasiveness; Phosphorylation; Signal Transduction; Smad3 Protein; Snail Family Transcription Factors; STAT3 Transcription Factor; Transcription Factors; Transforming Growth Factor beta; Tyrphostins

Combination chemotherapy with multiple drugs has been widely applied to cancer treatment owing to enhanced efficacy and reduced drug resistance. For drug combination experiment analysis, response surface modeling has been commonly adopted. In this paper, we introduce a Hill-based global response surface model and provide an application of the model to a 512-run drug combination experiment with three chemicals, namely AG490, U0126, and indirubin-3  ' -monoxime (I-3-M), on lung cancer cells. The results demonstrate generally improved goodness of fit of our model from the traditional polynomial model, as well as the original Hill model on the basis of fixed-ratio drug combinations. We identify different dose-effect patterns between normal and cancer cells on the basis of our model, which indicates the potential effectiveness of the drug combination in cancer treatment. Meanwhile, drug interactions are analyzed both qualitatively and quantitatively. The distinct interaction patterns between U0126 and I-3-M on two types of cells uncovered by the model could be a further indicator of the efficacy of the drug combination.

Topics: Adenosine Triphosphate; Antineoplastic Combined Chemotherapy Protocols; Butadienes; Cell Line, Tumor; Data Interpretation, Statistical; Dose-Response Relationship, Drug; Humans; Indoles; Lung Neoplasms; Models, Statistical; Nitriles; Oximes; Tyrphostins

In the present study, the effect of radiation on the invasion of the pulmonary adenocarcinoma cell line, A549, was investigated. Invasion of A549 cells irradiated with 2 and 4 Gy doses of γ‑ray was detected using the transwell Matrigel invasion assay. Levels of matrix metalloproteinase 2 (MMP‑2) and phosphorylated signal transducer and activator of transcription 3 (STAT3) were detected by reverse transcription PCR (RT-PCR) and/or immunoblotting. The enzyme activity of MMP‑2 was examined by gelatin zymography. Results demonstrated that the invasion of A549 cells was significantly enhanced by γ‑ray radiation at doses of 2 or 4 Gy. In addition, exposure to radiation was found to promote transcriptional expression of MMP‑2 and increase MMP‑2 enzyme activity. Irradiation activated the phosphorylation of STAT3 and promoted the nuclear localization of STAT3. The blockage of STAT3 phosphorylation using a specific inhibitor (AG490) suppressed the irradiation‑induced elevation of MMP‑2 expression, enzyme activity and invasion of A549 cells. Finally, the expression of vascular endothelial growth factor (VEGF) was found to be upregulated by radiation, which was associated with the activation of STAT3. Results of the current study indicate that irradiation leads to activation of STAT3 and translocation to the nucleus, leading to activation of VEGF and MMP‑2 transcription, resulting in the increased invasion of A549 cells.

Topics: Adenocarcinoma; Cell Line, Tumor; Cell Movement; Gamma Rays; Humans; Lung Neoplasms; Matrix Metalloproteinase 2; Phosphorylation; STAT3 Transcription Factor; Tyrphostins; Up-Regulation; Vascular Endothelial Growth Factor A

Cytokines in tumor microenvironment play an important role in the success or failure of molecular targeted therapies. We have chosen tumor necrosis factor α (TNF-α), TNF related apoptosis inducing ligand (TRAIL), insulin-like growth factor 1 (IGF-1) and transforming growth factor β (TGF-β) as representative pro-inflammatory, pro-apoptotic, anti-apoptotic and anti-inflammatory tumor derived cytokines. Analysis of Oncomine database revealed the differential expression of these cytokines in a subset of cancer patients. The effects of these cytokines on cytotoxicity of FDA approved drugs - cisplatin and taxol and inhibitors of epidermal growth factor receptor - AG658, Janus kinase - AG490 and SIRT1 - sirtinol were assessed in A549 lung cancer cells. TRAIL augmented cytotoxicity of sirtinol and IGF-1 had a sparing effect. Since TRAIL and IGF-1 differentially modulated sirtinol cytotoxicity, further studies were carried out to identify the mechanisms. Sirtinol or knockdown of SIRT1 increased the expression of death receptors DR4 and DR5 and sensitized A549 cells to TRAIL. Increased cell death in presence of TRAIL and sirtinol was caspase independent and demonstrated classical features of necroptosis. Inhibition of iNOS increased caspase activity and switched the mode of cell death to caspase mediated apoptosis. Interestingly, sirtinol or SIRT1 knockdown did not increase IGF-1R expression. Instead, it abrogated ligand induced downregulation of IGF-1R and increased cell survival through PI3K-AKT pathway. In conclusion, these findings reveal that the tumor microenvironment contributes to modulation of cytotoxicity of drugs and that combination therapy, with agents that increase TRAIL signaling and suppress IGF-1 pathway may potentiate anticancer effect.

Topics: Apoptosis; Benzamides; Cell Line, Tumor; Cell Survival; Cisplatin; Cytokines; Drug Resistance, Neoplasm; ErbB Receptors; Humans; Insulin-Like Growth Factor I; Janus Kinases; Lung Neoplasms; Molecular Targeted Therapy; Naphthols; Paclitaxel; Phosphatidylinositol 3-Kinases; Receptor, IGF Type 1; Receptors, TNF-Related Apoptosis-Inducing Ligand; Receptors, Tumor Necrosis Factor; Sirtuin 1; TNF-Related Apoptosis-Inducing Ligand; Transforming Growth Factor beta; Tumor Microenvironment; Tumor Necrosis Factor-alpha; Tyrphostins

Metastasis, a multistep process, is a major cause of mortality in cancer patients. Thus, it is hoped that inhibition of metastasis at any step, such as proliferation, migration, or invasion, using small-molecule inhibitors will reduce this mortality. Recent study suggests that the Janus kinase/signal transducer and activator of transcription 3 signal transduction pathway is a central pathway that regulates tumor progression and metastasis and can be blocked using tyrosine kinase inhibitors. In this study we used a synthetic tyrosine kinase inhibitor, AG490, to block the constitutive activation of the Janus kinase/signal transducer and activator of transcription 3 pathway in A549 lung carcinoma and A375 melanoma cell lines. Our results show that AG490 at subtoxic doses can effectively suppress tumor cell proliferation by limiting the expression of cyclin D1. Furthermore, AG490 is seen to induce apoptosis, inhibit cellular migration by disrupting actin organization, and suppress matrix metalloproteinase 2 activity. Taken together, these data demonstrate that AG490 can exert antimetastatic activity by inhibiting cellular proliferation, invasion, and migration.

Topics: Actins; Apoptosis; Cell Line, Tumor; Cell Movement; Cell Proliferation; Cyclin D1; Dose-Response Relationship, Drug; Enzyme Inhibitors; Humans; In Vitro Techniques; Janus Kinase 2; Lung Neoplasms; Matrix Metalloproteinase 2; Melanoma; Protein-Tyrosine Kinases; STAT3 Transcription Factor; Tyrphostins

Small cell lung cancer (SCLC) is an aggressive, rapidly metastasising tumour. Previously, we demonstrated the influence of CXCL12-CXCR4 interaction on processes involved in metastasis and chemoresistance in SCLC. We show here that STAT3 is expressed in both primary SCLC tumour tissues and SCLC cell lines. We investigated the function of STAT3 upon CXCL12 stimulation in SCLC cell lines. Small cell lung cancer cell lines present constitutive phosphorylation of STAT3, and in the reference cell lines NCI-H69 and NCI-H82 constitutive phosphorylation was further increased by CXCL12 stimulation. Further investigating this signalling cascade, we showed that it involves interactions between CXCR4 and JAK2 in both cell lines. However CXCL12-induced adhesion to VCAM-1 could be completely inhibited by the JAK2 inhibitor AG490 only in NCI-H82. Furthermore, CXCR4 antagonist but not AG490 inhibited cell adhesion whereas both antagonisms were shown to inhibit growth of the cells in soft agar, indicating the central involvement of this signalling in anchorage-independent growth of SCLC cells. Most interestingly, while using primary tumour material, we observed that in contrast to non-small-cell lung cancer samples from primary tumour tissues, all analysed samples from SCLC were strongly positive for tyrosine-phosphorylated STAT3. Taken together, these data indicate that STAT3 is constitutively phosphorylated in SCLC and is important in SCLC growth and spreading thus presenting an interesting target for therapy.

Topics: Antineoplastic Agents; Blotting, Western; Cell Adhesion; Humans; Immunoenzyme Techniques; Immunoprecipitation; Janus Kinase 2; Lung Neoplasms; Phosphorylation; Receptors, CXCR4; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; Small Cell Lung Carcinoma; STAT3 Transcription Factor; Stromal Cells; Tumor Cells, Cultured; Tyrphostins; Vascular Cell Adhesion Molecule-1

We previously reported that matrix metalloproteinase (MMP)-10 mRNA levels were significantly lower in tumor tissues than in adjacent normal tissues in human non-small cell lung cancer (NSCLC), whereas protein levels of MMP-10 were higher in the tumor tissues than the adjacent tissues. The mechanism of this divergence is still unknown. In the present study the role of Janus kinase 2/signal transducers and activators of transcription 3 (JAK2/STAT3) on interleukin (IL)-6 mediated regulation of MMP-10 expression was investigated in a human lung adenocarcinoma cell line (A549 cells) and the molecular regulatory mechanism of MMP-10 expression was explored. A549 cells were stimulated by different concentrations of IL-6 with or without AG490, a specific JAK2 inhibitor. It was demonstrated that IL-6 moderately reduced the MMP-10 mRNA levels, whereas it significantly enhanced the MMP-10 protein mass in the A549 cells. This phenomenon mimicked the divergence of mRNA level and protein mass of MMP-10 in human NSCLC. Moreover, the present study indicated that IL-6 regulation of MMP-10 expression was via the JAK2/STAT3 pathway. STAT3 mRNA levels were significantly increased when the cells were treated with IL-6, whereas when AG490 (50 muM) was added to the cell cultures, IL-6-induced increase of STAT3 mRNA levels was abolished. Meanwhile, AG490 blocked the IL-6-induced inhibition of MMP-10 mRNA as well as blocking the IL-6-induced increase of MMP-10 protein mass in the A549 cells. Neither IL-6 nor AG490 influenced JAK2 mRNA levels in the A549 cell cultures. It is concluded that the JAK2/STAT3 pathway is involved in the IL-6-mediated regulation of MMP-10, and IL-6 can moderately reduce MMP-10 mRNA levels and strongly increase MMP-10 protein mass in human lung adenocarcinoma A549 cells. Contrasting effects of IL-6 on MMP-10 mRNA level and protein concentration in A549 cells may partially explain the divergence of MMP-10 mRNA level and protein mass in human NSCLC.

Topics: Adenocarcinoma; Cell Line, Tumor; Gene Expression Regulation, Enzymologic; Gene Expression Regulation, Neoplastic; Humans; Interleukin-6; Janus Kinase 2; Lung Neoplasms; Matrix Metalloproteinase 10; RNA, Messenger; Signal Transduction; STAT3 Transcription Factor; Tyrphostins

Up-regulated signal transducers and activators of transcription (STAT)-mediated signaling is believed to contribute to the pathogenesis of a variety of solid and hematologic cancers. Consequently, inhibition of STAT-mediated signaling has recently been proposed as a potential new therapeutic approach to the treatment of cancers. Having shown previously that the pan-cyclin-dependent kinase inhibitor flavopiridol binds to DNA and seems to kill cancer cells via that process in some circumstances, we evaluated the hypothesis that flavopiridol might consequently disrupt STAT3/DNA interactions, attenuate STAT3-directed transcription, and down-regulate STAT3 downstream polypeptides, including the antiapoptotic polypeptide Mcl-1. SDS-PAGE/immunoblotting and reverse transcription-PCR were used to assess RNA and polypeptide levels, respectively. DNA cellulose affinity chromatography and a nuclear elution assay were used to evaluate the ability of flavopiridol to disrupt STAT3/DNA interactions. A STAT3 luciferase reporter assay was used to examine the ability of flavopiridol to attenuate STAT3-directed transcription. Colony-forming assays were used to assess cytotoxic synergy between flavopiridol and AG490. Flavopiridol was found to (a) disrupt STAT3/DNA interactions (DNA cellulose affinity chromatography and nuclear elution assay), (b) attenuate STAT3-directed transcription (STAT3 luciferase reporter assay), and (c) down-regulate the STAT3 downstream antiapoptotic polypeptide Mcl-1 at the transcriptional level (reverse transcription-PCR and SDS-PAGE/immunoblotting). Furthermore, flavopiridol, but not the microtubule inhibitor paclitaxel, could be combined with the STAT3 pathway inhibitor AG490 to achieve cytotoxic synergy in A549 human non-small cell lung cancer cells. Collectively, these data suggest that flavopiridol can attenuate STAT3-directed transcription in a targeted fashion and may therefore be exploitable clinically in the development of chemotherapy regimens combining flavopiridol and other inhibitors of STAT3 signaling pathways.

Topics: Antineoplastic Combined Chemotherapy Protocols; Carcinoma, Non-Small-Cell Lung; DNA; Down-Regulation; Drug Screening Assays, Antitumor; Enzyme Inhibitors; Flavonoids; Humans; Janus Kinase 1; Lung Neoplasms; Myeloid Cell Leukemia Sequence 1 Protein; Neoplasm Proteins; Phosphoproteins; Piperidines; Protein-Tyrosine Kinases; Proto-Oncogene Proteins c-bcl-2; RNA Polymerase II; STAT3 Transcription Factor; Transcription, Genetic; Tumor Cells, Cultured; Tyrphostins

Neuregulin-1 (NRG-1) is part of a family of proteins whose members are structurally related to epidermal growth factor. NRG-1 induces cell proliferation through a high-affinity receptor complex composed of a heterodimer of human epidermal growth factor-like receptor (HER) 2 and 3. In this study, we show that NRG-1 activates the Janus kinases (JAK) and signal transducer and activator of transcription proteins (STAT). NRG-1 induced a rapid and transient increase in tyrosine phosphorylation of TYK2 and JAK3, but not JAK1 or JAK2, and induced STAT3 and STAT5 tyrosine phosphorylation. Upon phosphorylation, STAT3 translocated to the nucleus within 1 h. Activation of the JAK-STAT pathway was dependent on HER2/HER3 heterodimerization and was necessary for NRG-1-induced proliferation. Inhibition of HER2's ability to dimerize using the HER2-specific antibody 2C4 completely blocked NRG-1-induced JAK3, TYK2, STAT3, and STAT5 tyrosine phosphorylation. Blocking the JAK-STAT pathway with a specific JAK-STAT pathway inhibitor, AG490, inhibited NRG-1-induced JAK and STAT phosphorylation and cell proliferation. These data suggest that NRG-1 activates the JAK-STAT signal transduction pathway through its high-affinity receptor, the HER2/HER3 heterodimer. This pathway plays an important role in NRG-1-stimulated proliferation of pulmonary epithelial cells.

Topics: Cell Division; Cell Nucleus; DNA-Binding Proteins; Enzyme Inhibitors; Epithelial Cells; Humans; Janus Kinase 1; Janus Kinase 3; Lung; Lung Neoplasms; Milk Proteins; Neuregulin-1; Phosphorylation; Protein Transport; Protein-Tyrosine Kinases; Receptor, ErbB-2; Receptor, ErbB-3; STAT3 Transcription Factor; STAT5 Transcription Factor; STAT6 Transcription Factor; Time Factors; Trans-Activators; Tumor Cells, Cultured; Tyrosine; Tyrphostins

Disruption of apoptosis may allow metastatic cell survival and confer resistance to chemotherapeutic drugs. We have analysed the molecular pathways that activate these survival genes in specific sites of metastasis. Estrogen receptor-negative breast cancer cell line MDA-MB435 and two metastatic sublines derived from lung (435L) and brain (435B) were analysed for the expression of members of the Bcl-2 family of apoptosis regulators. The levels of Bcl-2 were higher in the metastatic sublines than in parental cells, which correlated with the activation of Stat3, but not with the expression and/or activation of known bcl-2 transcription factors (CREB and WT1). In the brain subline, both expression of Bcl-2 and Stat3 activation were induced by epidermal growth factor and abrogated after treatment with kinase inhibitors specific for epidermal growth factor receptor or Jak2. Furthermore, transfection of 435B with a dominant-negative Stat3 markedly reduced the expression of Bcl-2 protein, whereas transient expression of a constitutively active Stat3 increased Bcl-2 in parental 435 cells. In addition, blockade of Stat3 activation by treatment with epidermal growth factor receptor and Jak2 kinase inhibitors or transfection with a dominant negative Stat3, sensitizes 435B cells to chemotherapy-induced apoptosis. Our data suggest that an increased activation of the Stat3-Bcl-2 pathway in estrogen receptor-negative metastatic breast cancer cell lines confer a survival advantage to these cells and contribute to their chemoresistance.

Topics: Antineoplastic Agents; Apoptosis; bcl-X Protein; Brain Neoplasms; Breast Neoplasms; Carcinoma; Cyclic AMP Response Element-Binding Protein; DNA-Binding Proteins; Enzyme Inhibitors; ErbB Receptors; Female; Humans; Janus Kinase 2; Lung Neoplasms; Neoplasm Metastasis; Neoplasms, Hormone-Dependent; Protein-Tyrosine Kinases; Proto-Oncogene Proteins; Proto-Oncogene Proteins c-bcl-2; Receptors, Estrogen; STAT3 Transcription Factor; Trans-Activators; Transcription, Genetic; Tumor Cells, Cultured; Tyrphostins