High STAT3 Expression Research Articles

123P - Signal transducer and activator of transcription–3 (STAT3) expression concordance in paired primary and metastatic colorectal cancers (mCRC)

STAT3 is a constitutively activated transcription factor in several cancers. In patients with mCRC overexpression of STAT3 is associated with worse survival. To determine if STAT3 is a potential target for therapy and to evaluate expression through metastatic progression in mCRC the concordance of expression in primary and metastatic tumors was assessed. Patients treated at the Ottawa Hospital from 2001-2012 were retrospectively identified and included if tumor tissue was available from both the primary and a metastasis. Tissue microarrays were constructed using 2 x 2mm cores for each tumor. Nuclear phosphorylated STAT3 expression intensity by immunohistochemistry was evaluated by 2 independent pathologists as 0 (absent), 1-2 (low), or 3 (high). The primary outcome was concordance of STAT3 expression between primary and metastatic sites. Secondary outcomes included correlation of STAT3 expression with demographic and disease characteristics as well as clinical outcomes. Among 38 patients identified 52% were male, median age at diagnosis was 61, and 36% of metastases were synchronous. Expression of STAT3 in primary tumors was 5% high, 42% low and absent in 53% compared to 16% high, 47% low, and 37% absent in metastatic samples. Expression between paired primary and metastatic samples was concordant in 33% of patients whereas it increased in 21% and decreased in 45%. A weak correlation was observed between primary and metastatic STAT3 expression (Pearson's correlation 0.1). After a median follow-up of 13.1 years, 30 of 35 patients included in the survival analysis died. Median survival was 4.6 years. Higher STAT3 expression in primary tumors showed a trend towards worse survival (HR 1.7, 95% CI 0.81-3.64, p = 0.1), while there was no prognostic correlation of STAT3 expression in metastases. In patients with mCRC there was low concordance of STAT3 expression in primary and metastatic tumors. STAT3 expression in the primary, but not the metastatic site, was related to survival, indicating that the prognostic value of STAT3 depended on tumor sampling. These results have important implications for further research in the use of STAT3 as a biomarker in patients with mCRC.

Abstract 4545: The E3 ubiquitin ligase COP1 controls STAT3 turnover and its loss leads to increased STAT3 stabilization and activation in prostate cancer

Abstract The E3 ubiquitin ligase COP1 acts as a tumor suppressor and is deleted in a small percentage of prostate cancers. We reported that COP1 was repressed in prostate tumors through miRNA-mediated silencing, which represents an alternative and more frequent event than genetic deletion. COP1 controls ubiquitination and turnover of c-Jun and ETV1 and its loss has been associated with over-expression of these transcription factors. In this study, we identify STAT3 as a novel substrate of COP1 and report that concomitant deregulation of COP1 and STAT3 leads to prostate cancer progression. In a panel of prostate cancer cell lines, low expression of COP1 was associated with increased level of STAT3 protein and a more aggressive phenotype. Knockdown of COP1 in normal prostate epithelial RWPE-1 cells increased total (tSTAT3) and phosphorylated STAT3 (pSTAT3) and promoted tumorigenic properties. Conversely, over-expression of COP1 in DU145 cells, expressing low level of COP1, reversed the transformed phenotype and reduced tSTAT3 and pSTAT3 level and activation. COP1/STAT3 anti-correlation suggested that STAT3 was a substrate of COP1 for ubiquitination and degradation by the ubiquitin-proteasome system. Consistently, the proteasome inhibitor PS-341 prevented down-regulation of STAT3 in response to COP1 in DU145 cells. Furthermore, COP1 knockdown delayed significantly STAT3 protein turnover in RWPE-1 cells, indicating that COP1 regulated STAT3 degradation in these cells. Co-IP showed that COP1 and STAT3 directly interacted in RWPE-1 and DU145 cells. Interestingly, co-IP with the wild type and phosphorylation defective Y705F mutant STAT3 showed that the interaction with COP1 did not depend on STAT3 phosphorylation. Furthermore, the level of ubiquitinated STAT3 in RWPE1 cells was reduced after COP1 knockdown, whereas increased in DU145 cells after COP1 over-expression. These data provided evidence of COP1-dependent ubiquitination and degradation of STAT3 in normal prostate epithelial cells and, conversely, STAT3 accumulation and activation in prostate cancer cells with loss of COP1. To assess the clinical relevance of these findings, we examined the level of COP1, tSTAT3 and pSTAT3 by IHC in primary prostate tumors (n = 136) from patients with long-term clinical follow up. Low COP1 levels were significantly associated with high tSTAT3 expression (Fisher test p<0.001). Furthermore, pSTAT3 was prevalently observed in the group of COP1 negative/STAT3 high tumors indicative of STAT3 activation. The combination of low COP1 and high STAT3 expression was associated with significantly higher risk of disease recurrence after prostatectomy (p≤0.01) marking patients at risk of poor clinical outcome. Collectively, this study identifies STAT3 as a substrate of COP1 and provides evidence of a novel pathway leading to STAT3 activation in human tumors. Citation Format: Cecilia Dallavalle, George Thalmann, Carlo V. Catapano, Giuseppina M. R. Carbone. The E3 ubiquitin ligase COP1 controls STAT3 turnover and its loss leads to increased STAT3 stabilization and activation in prostate cancer. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 4545.

Molecular heterogeneity in a patient-derived glioblastoma xenoline is regulated by different cancer stem cell populations.

Malignant glioblastoma (GBM) is a highly aggressive brain tumor with a dismal prognosis and limited therapeutic options. Genomic profiling of GBM samples has identified four molecular subtypes (Proneural, Neural, Classical and Mesenchymal), which may arise from different glioblastoma stem-like cell (GSC) populations. We previously showed that adherent cultures of GSCs grown on laminin-coated plates (Ad-GSCs) and spheroid cultures of GSCs (Sp-GSCs) had high expression of stem cell markers (CD133, Sox2 and Nestin), but low expression of differentiation markers (βIII-tubulin and glial fibrillary acid protein). In the present study, we characterized GBM tumors produced by subcutaneous and intracranial injection of Ad-GSCs and Sp-GSCs isolated from a patient-derived xenoline. Although they formed tumors with identical histological features, gene expression analysis revealed that xenografts of Sp-GSCs had a Classical molecular subtype similar to that of bulk tumor cells. In contrast xenografts of Ad-GSCs expressed a Mesenchymal gene signature. Adherent GSC-derived xenografts had high STAT3 and ANGPTL4 expression, and enrichment for stem cell markers, transcriptional networks and pro-angiogenic markers characteristic of the Mesenchymal subtype. Examination of clinical samples from GBM patients showed that STAT3 expression was directly correlated with ANGPTL4 expression, and that increased expression of these genes correlated with poor patient survival and performance. A pharmacological STAT3 inhibitor abrogated STAT3 binding to the ANGPTL4 promoter and exhibited anticancer activity in vivo. Therefore, Ad-GSCs and Sp-GSCs produced histologically identical tumors with different gene expression patterns, and a STAT3/ANGPTL4 pathway is identified in glioblastoma that may serve as a target for therapeutic intervention.

Targeting of MDS and AML Stem Cells Via Inhibition of STAT3 By Pyrimethamine

Acute Myeloid Leukemia (AML) and Myelodysplastic syndrome (MDS) arise from accumulation of multiple stepwise genetic and epigenetic changes in hematopoietic stem cells (HSC) and/or committed progenitors. A series of transforming events can initially give rise to pre-leukemia stem cells (pre-LSC) as well as fully transformed leukemia stem cells (LSC), both of which need to be targeted in strategies aimed at curing these diseases. We conducted parallel transcriptional and epigenetic analysis of highly fractionated stem and progenitor populations in individual patients of MDS and identified STAT3 upregulation in MDS HSCs. qRTPCR in an independent set of sorted MDS/AML HSCs (Lineage-negative, CD34+, CD38-) confirmed the significant increase in STAT3 expression in 60% of cases when compared to healthy controls. We further analyzed gene expression profiles of CD34+ cells from 183 MDS patients and found significant increased expression of STAT3 in MDS when compared to healthy controls (FDR<0.1) and importantly, found that increased STAT3 expression was predictive of significantly adverse prognosis (log rank P value < 0.01, median survival of 2.6 years compared to 5.8 years for group with lower STAT3) in patients. This further points to a critical role of STAT3 signaling in AML/MDS pathogenesis and progression.Next, we studied the functional role of STAT3 by using Pyrimethamine as a clinically relevant inhibitor. Pyrimethamine is an FDA approved antifolate compound that was found to be a specific inhibitor of STAT3 activity in an initial screen of 1120 compounds. 3D modeling studies reveal that Pyrimethamine can occupy the SH2 domain of STAT3 protein with high avidity. It shows half-maximal activity (EC50) for STAT3 inhibition at approximately 1.5 μM, well within the plasma concentrations found with clinical use.We assessed the effect of Pyrimethamine on cellular proliferation of multiple leukemic cell lines (KG-1, KT-1 and CMK), and observed that these were significantly inhibited at 120 hours of exposure in a dose dependent fashion (t test, p value <0.004, Mean + SD of 3 experiments). Pyrimethamine was also able to induce significant apoptosis in AML cell lines at 72 hours at a 10 μM concentration. (P Value <0.05)In summary, we have found significant demethylation and increased expression of STAT3 in sorted HSCs from AML and MDS patients. High STAT3 expression is a marker of adverse prognosis in a large cohort of MDS patients. In vitro studies show that Pyrimethamine can inhibit STAT3 activation and is able to inhibit proliferation by inducing apoptosis in leukemic cells. A phase II study evaluating Pyrimethamine for the treatment of intermediate/high-risk MDS patients that have relapsed or are refractory to azanucleoside therapy has been initiated. DisclosuresBrown:Sanofi, Onyx, Vertex, Novartis, Boehringer, GSK, Roche/Genentech, Emergent, Morphosys, Celgene, Janssen, Pharmacyclics, Gilead: Consultancy.

The tumor suppressor miR-124 inhibits cell proliferation by targeting STAT3 and functions as a prognostic marker for postoperative NSCLC patients.

The aim of the present study was to investigate the role of miR-124 in lung cancer and identify the potential predictive value of miR-124 in postoperative non-small cell lung cancer (NSCLC) patients. We detected miR-124 expression in A549, NCL-H460 and normal lung epithelial BEAS-2E cells and showed a significantly lower expression level of miR-124 in NSCLC cells than in BEAS-2E cells. Upregulation of miR-124 expression levels in both A549 and NCL-H460 cells by transfection with miR-124 mimics suppressed cell proliferation and induced apoptosis. Further investigation revealed that miR-124 bound directly to the 3' UTR region of STAT3, thereby inhibiting STAT3 expression. In addition, miR-124 levels detected in NSCLC tissues were lower than those in adjacent normal lung tissues, while the opposite was observed for STAT3. In NSCLC, the expression levels of miR-124 and STAT3 correlated significantly with the tumor node metastases (TNM) stage, differentiation grade and lymph node metastasis, while the levels of these molecules did not differ significantly by gender, age, location, smoking index, pleural invasion or pathological type. The expression level of miR-124 was significantly associated with disease-free survival (DFS) in both positive and negative lymph node groups. Furthermore, patients with low miR-124 or high STAT3 expression generally received a worse prognosis in terms of both overall survival (OS) and DFS. In conclusion, our findings suggest that miR-124 functions as a tumor suppressor by targeting STAT3, and that miR-124 may potentially serve as a useful biomarker for the prognosis of NSCLC patients.

Correlation and malignant transformation of MSCs with the overexpression of SIL-R/GP130 under the tumor microenvironment.

Mesenchymal stem cells (MSCs) can differentiate into chondrogenesis, osteogenesis, myocardium and nerve in specific conditions, but it may undergo malignant transformation when cultivated with tumor cells. This research was to provide preliminary experimental basis for the safe application of MSCs and seek for drugs to avoid the malignant transformation of MSCs in the treatment of tumor. Accordingly, four groups including experimental group, positive control group, negative contrast group and blank group were set. Experimental group was the co-culture group of MSCs and C6 glioma cells. Positive control group was the regular culture group of C6 glioma cells. Negative control group was the co-culture group of MSCs and astrocyte. And blank group was the regular culture group of MSCs. The results showed the expression of IL-6, and IL-6R significantly increased in the group of co-culture with C6; the proliferation situation of MSCs was obviously strengthened; MSCs performed a high expression of GP130, STAT-3, CyclinD1 and BCL-xl, which had statistical significance compared with the contrast group. It can be concluded that the malignant expression of MSCs was related to the overexpression and activation of SIL-R/GP130; and the excessive expression and activation of SIL-6R and GP130 might be one of the important reasons for malignant transformation of MSCs under the microenvironment.

Effects of matrine on JAK-STAT signaling transduction pathways in bleomycin-induced pulmonary fibrosis

The current study aims to investigate the effects of matrine on the JAK-STAT signaling transduction pathways in bleomycin (BLM)-induced pulmonary fibrosis (PF) and to explore its action mechanism. A total of 72 male C57BL/6 mice were randomized into the control, model, and treatment groups. PF models were established by instilling BLM intratracheally. The treatment group was given daily matrine through gastric lavage. Six mice were sacrificed in each group at 3, 7, 14, and 28 days. The lung tissues were observed using hematoxylin-eosin staining. The expression of JAK, STAT1, and STAT3 was observed using immunohistochemistry and then determined using real-time polymerase chain reaction. Alveolitis and PF significantly improved in the treatment group compared with the model group (P < 0.05). The expression of JAK, STAT1, and STAT3 in the model group increased at day 7, peaked at day 14 and then decreased, but the expression was still higher than that in the control group at day 28 (P < 0.05). The three indices in the treatment group were significantly lower than those in the model group at any detection time point (P < 0.05). PF causes high expression of JAK, STAT1, and STAT3. Matrine exerts an anti-PF effect by inhibiting the JAK-STAT signaling transduction pathways.

Abstract 304: Reactive oxygen species induce prostate cancer cells via activation of the IL6/STAT3 pathway.

Abstract Background: Previously we have established a stepwise prostate carcinogenesis model including human primary prostate epithelial EP156T cells, non-malignant mesenchymal EPT1 cells and pre-malignant EPT2-D5 cells. Aims and methods: To further achieve malignant cells, EPT2-D5 cells were grown in protein-free medium and the tumorigenesis was tested in vivo. Results: 1) Lots of spheres were generated in monolayer culture of D5 cells in protein free medium. The sphere forming cells were named D5HS. 2) Different from EPT2-D5 cells, D5HS formed large subcutaneous tumors (EPT3) and metastasis (EPT3M) in SCID mice. 3) Gene expression profiling showed inflammation signatures in D5HS. 4) Cellular Reactive Oxygen Species Detection Assay (DCFDA) showed high reactive oxygen species (ROS) in D5HS, and ELISA and Western blotting showed high secreted IL6 and high STAT3 expression in D5HS. 5) Blocking of ROS or STAT3 signaling reduced the sphere formation. 6) Immunohistochemistry staining showed high pSTAT3 level in the EPT3 tumor. Conclusions: Reactive oxygen species induce prostate cancer cells via activation of the IL6/STAT3 pathway. Citation Format: Yi Qu, Runhui Liu, Jigang Zhang, Mihaela Popa, Yaping Hua, Biaoyang Lin, Emmet McCormack, Weidong Zhang, Anne Margrete Oyan, Karl-Henning Kalland, Xisong Ke. Reactive oxygen species induce prostate cancer cells via activation of the IL6/STAT3 pathway. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 304. doi:10.1158/1538-7445.AM2013-304

Abstract 5452: High-fat diet induces inflammation by increasing estrogen levels through Stat3, estrogen receptor alpha and aromatase in the mouse prostate.

Abstract High-fat diet (HFD), an unfortunate lifestyle choice common in the Western world, is associated with inflammation and thereby is regarded as a risk factor for various diseases including obesity, diabetes, and cancer. The molecular mechanism(s) responsible for HFD-induced inflammation in the prostate gland are not well understood. Estrogen receptor alpha (ER-α), signal transducer and activator of transcription (Stat)-3 and aromatase are important signaling molecules constitutively activated during inflammation and have been shown to be upregulated in prostate cancer. We have previously reported that HFD activates a pro-inflammatory response in the prostate through elevated expression of Stat-3 (Prostate 72:233-43, 2012). In the current study, we sought to investigate a possible link between intraprostatic inflammation, HFD feeding, signaling molecules involved in inflammation viz. ER-α, Stat-3, p-Stat3 (Ser727) and aromatase. C57BL/6 mice were fed either a regular diet (RD) or a HFD for 4, 8 and 12 weeks. Serum, visceral fat and prostate tissues were obtained for analysis. HFD intake caused an increase in fat and serum testosterone levels. No significant changes were observed in the testosterone levels in the prostate. A marked increase in estrogen levels was noted in the prostate and visceral fat at 8 weeks of HFD intake. HFD feeding resulted in a significant increase in the expression of aromatase, ER-α, Stat-3 and p-Stat3 (Ser727) at the message and protein levels. Immunoprecipitation and ChIP analysis demonstrated an increased association between aromatase, p-Stat-3 and ER-α in the HFD group. IHC analysis revealed higher expression of Stat-3, ER-α and aromatase in the prostate, accompanied by the morphologic evidence of increased intraprostatic inflammation in the HFD group. Taken together, our findings suggest that HFD increases estrogen levels by increased binding of ER-α and p-Stat-3 to the promoter of aromatase and their interaction is associated with increased intraprostatic inflammation which might contribute to the initiation and development of prostate cancer. Citation Format: Natarajan Bhaskaran, Sanjeev Shukla, Vijay S. Thakur, Melissa A. Babcook, Gregory T. MacLennan, Guiming Liu, Firouz Daneshgari, Sanjay Gupta. High-fat diet induces inflammation by increasing estrogen levels through Stat3, estrogen receptor alpha and aromatase in the mouse prostate. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 5452. doi:10.1158/1538-7445.AM2013-5452

High nuclear expression of STAT3 is associated with unfavorable prognosis in diffuse large B-cell lymphoma

BackgroundThe purpose of the study was to investigate the expression and prognostic value of STAT3 in diffuse large B-cell lymphoma (DLBCL).MethodsSeventy-four DLBCL patients from 2001 to 2007 were reviewed in the study. The STAT3 expression in their tumor tissues was examined using the immunohistochemistry (IHC) method, and evaluated for its association with clinicopathological parameters.ResultsStrong nuclear staining of STAT3 and phosphorylated-STAT3tyr705 (P-STAT3) were observed in 19 cases (25.7%) and 24 cases (32.4%), respectively, and the expression levels were highly consistent between them (P = 0.001). The high nuclear expression of STAT3 was more frequent in the non-germinal center B cell-like (non-GCB) DLBCL than that in the GCB subtype, but not reaching significance (P < 0.061). The high nuclear expression of STAT3 was found to be correlated with poor overall survival (OS) (P = 0.005). Multivariate Cox regression analysis showed that the STAT3 expression was an independent prognostic factor for DLBCL patients regardless of CHOP or R-CHOP regimen used as the first-line therapy.ConclusionSTAT3 is more frequently expressed in non-GCB DLBCL than that in GCB subtype, and its strong nuclear expression is correlated with poor OS in DLBCL.

Expression of Stat3 and Notch1 is associated with cisplatin resistance in head and neck squamous cell carcinoma

Cisplatin is the most important chemotherapeutic agent involved in treatment of head and neck squamous cell carcinoma (HNSCC), but cisplatin resistance in HNSCC is still a serious problem in clinic. The reasons why patients fail chemotherapy are unclear. We examined 25 HNSCC patients who were all tested for cisplatin sensitivity by CD-DST (collagen gel droplet embedded culture-drug sensitivity) method and expression of Stat3 and Notch1. We found that high expression levels of Stat3 and Notch1 were closely associated with cisplatin resistance respectively (P=0.014, P=0.000). In addition, cisplatin resistance of HNSCC was decreased after inhibition of Stat3 or Notch signaling in vitro. Our results provide first evidence that both high Stat3 and Notch1 expression are associated with cisplatin resistance in HNSCC patients, supporting the hypothesis that co-activation of Stat3 and Notch1 by their crosstalk induces the reprogrammed survival pathways in HNSCC responding to chemotherapy.

An Automated and Quantitative Protein Expression-Based Classification System to Identify High Risk Multiple Myeloma Patients

Background: Autologous stem cell transplantation (ASCT) has dramatically improved the survival of myeloma patients; however, this approach has significant toxicities and nearly 25% of MM patients progress within one year from their transplant. While gene expression profiling-based (GEP) molecular classification has permitted the identification of unresponsive high-risk patients, these approaches have proven too costly and complex to translate into clinical practice. Less expensive and more readily available methods are needed clinically to identify, at the time of diagnosis, MM patients who may benefit from more aggressive or experimental therapies. While protein-based tissue arrays offer such alternative, biases introduced by the “observer-dependent” scoring methods have limited their wide applicability.Methods: We have designed a simplified, fully automated and quantitative protein expression based-classification system that will allow us to accurately predict survival post ASCT in a cost effective and “observer-independent” manner. We constructed tissue microarrays using diagnostic bone marrow biopsies of 82 newly diagnosed MM patients uniformly treated with a dexamethasone based induction regimen and frontline ASCT. Using the HistoRx PM-2000 quantitative immunohistochemistry platform, coupled with the AQUA analysis software, we have examined the expression of the following proteins: FGFR3 which is associated with t(4;14), cyclin B2 and Ki-67 which are associated with cellular proliferation, TACI which is associated with maf deregulation, and phospho-Y705 STAT3 and p65NF-κB, which are associated with myeloma cell growth and survival. For FGFR3, patients were divided into FGFR3 positive and negative groups based on hierarchical clustering of their AQUA score. For all other proteins examined, based on AQUA scores, the top quartiles or quintiles of patients were classified as high expression groups. Based on the univariate analysis, patients were further classified as “High Risk” MM if they had been identified as high expressers of either TACI, p65NF-κB or FGFR3. The Kaplan-Meier method was used to estimate time to progression and overall survival. Multivariate analysis was performed using the Cox regression method.Results: 82 patients were included in this study. In univariate analysis, FGFR3 and p65NF-κB expression were associated with significantly shorter TTP (p=0.018 and p=0.009) but not OS (p=0.365 and p=0.104). TACI expression levels predicted for worse OS (p=0.039) but not TTP (p=0.384). High expression of Ki67 or phospho-Y705 STAT3 did not affect survival. Of the 82 cases, 67 were included in the multivariate analysis since they had AQUA scores available for all markers: 26 (38.8%) were considered as High Risk by their AQUA scores and had significantly shorter TTP (p=0.014) and OS (p=0.006) compared to the Low Risk group. The median TTP for the Low and High Risk groups was 2.9 years and 1.9 years, respectively. The 5-years estimates for OS were 60.6% for the High Risk group versus 83.5% for the Low Risk group. Multivariate analysis was performed using del13q and our risk group classification as variables. Both our risk group classification and del13q were independent predictors for TTP, having 2.4 and 2.3 greater risk of relapse, respectively. Our risk group classification was the only independent predictor of OS with the High Risk group having a 5.9 fold greater risk of death.Conclusions: We have found that the expression of FGFR3, TACI, and p65NF-κB, in an automated and fully quantitative tissue-based array, is a powerful predictor of survival post-ASCT in MM and eliminates the “observer-dependent” bias of scoring TMAs. A validation of this “High Risk” TMA based signature is currently underway in larger and independent cohorts. [Display omitted]

Cooperative signaling through the signal transducer and activator of transcription 3 and nuclear factor-κB pathways in subtypes of diffuse large B-cell lymphoma

AbstractThe activated B cell–like (ABC) subgroup of diffuse large B-cell lymphoma (DLBCL) is characterized by constitutive activation of the nuclear factor-κB (NF-κB) pathway. In this study, we showed that the NF-κB pathway induced the expression of the cytokines interleukin (IL)-6 and IL-10 in ABC DLBCL cell lines, which also have high levels of total and phosphorylated signal transducer and activator of transcription (STAT) 3 protein, suggesting autocrine signaling. Using RNA interference for STAT3, we defined a gene expression signature of IL-6 and IL-10 signaling through STAT3. Based on this signature, we constructed a molecular predictor of STAT3 signaling that defined a subset of ABC DLBCL tumors with high expression of STAT3, IL-6, and/or IL-10 and their downstream targets. Although the STAT3-high and STAT3-low subsets had equivalent expression of genes that distinguish ABC DLBCL from germinal center B cell–like DLBCL, STAT3-high ABC DLBCLs had higher expression of signatures that reflected NF-κB activity, proliferation, and glycolysis. A small-molecule inhibitor of Janus kinase signaling, which blocked STAT3 signature expression, was toxic only for ABC DLBCL lines and synergized with an inhibitor of NF-κB signaling. These findings suggest that the biological interplay between the STAT3 and NF-κB pathways may be exploited for the treatments of a subset of ABC DLBCLs.

Constitutively Activated STAT3 Promotes Cell Proliferation and Survival in the Activated B Cell Subtype of Diffuse Large B Cell Lymphomas.

During B cell development, cell proliferation and survival are regulated by stage-specific transcription factors. Accordingly, distinct oncogenic pathways are employed by B cell lymphomas representing different stages of B cell development. Diffuse large B cell lymphoma (DLBCL) contains at least two main phenotypic subtypes, i.e. the germinal center B cell-like (GCB-DLBCL) and the activated B cell-like (ABC-DLBCL) groups. It has been shown that GCB-DLBCL responds favorably to chemotherapy and expresses high levels of BCL6, a transcription repressor known to play a causative role in lymphomagenesis. In comparison, ABC-DLBCL has lower levels of BCL6, constitutively activated NF-kappaB and tends to be refractory to chemotherapy. In this study, we investigated the relationship between BCL6 and STAT3 expression/activation in DLBCL and normal GC B cells. Our results demonstrate that BCL6 directly inhibits transcription of the STAT3 gene by binding to two BCL6 sites in its 5′ regulatory region. As a result, high level STAT3 expression and activation are preferentially detected in ABC-DLBCL and BCL6-negative normal germinal center B cells. Specifically, in tonsillar GCs, STAT3 expression and activation is restricted to a previously uncharacterized subset of BCL6−Blimp-1− B cells in the apical light zone. The location and phenotype of these cells suggest that they are in the process of exiting the BCL6-directed GC program and transitioning to a plasma cell differentiation process governed by Blimp-1. The reciprocal relationship between BCL6 and STAT3 is also conserved in DLBCL such that STAT3 expression and activation is preferentially associated with the BCL6-low, ABC subtype. Most importantly, inactivating STAT3 by either AG490 or small interference RNA in ABC-DLBCL cells inhibits cell proliferation and triggers apoptosis. These phenotypes are accompanied by decreased expression of several known STAT3 target genes, including c-Myc, JunB and Mcl-1, and increased expression of the cell cycle inhibitor p27. In addition to identifying STAT3 as a novel BCL6 target gene, our results define STAT3 activation as a second oncogenic pathway operating in ABC-DLBCL and suggest that blocking STAT3 may be potentially therapeutic in treatment of these aggressive lymphomas.

Differential expression of STAT5 and Bcl-x L, and high expression of Neu and STAT3 in non-small-cell lung carcinoma

Experimental evidence suggests that in lung cancer, development, progression and an increased proliferation rate can be linked to apoptosis-related factors. The objective of this study is to evaluate the status of Neu, signal transducer and activator of transcription (STAT)-3, STAT5 and Bcl-xL expression in non-small-cell lung cancer. We investigated the immunohistochemical expression of these proteins in 92 non-small-cell lung cancer specimens to establish their role in lung cancer pathogenesis. Neu was overexpressed in 65% of cases, and although STAT3 was overexpressed in 52.1% in cytoplasm, it was expressed in nucleus (activated) in 60.8%. Meanwhile, STAT5 was found overexpressed in 41.3% in cytoplasm and 32.6% in nucleus. Thus, Bcl-xL was overexpressed in cytoplasm in 81.5%. Interestingly, we found nuclear expression of Bcl-xL in 30.4% of cases. Finally, we found correlation among histological types of lung cancer and nuclear expression of both STAT5 (P=0.005) and nuclear Bcl-xL (P=0.003). Besides, nuclear expression of Bcl-xL was correlated with TNM stage IV (distant metastasis) (P=0.02). These results suggest for the first time, a relevant role for STAT5 and Bcl-xL as apoptosis-regulatory proteins in the pathogenesis of lung cancer, and overexpression of both Neu and activated STAT3, could be related with the proliferation rate in lung carcinoma cells.