T helper 17 (Th17), a distinct subset of CD4(+) T cells with IL-17 as their major cytokine, orchestrate the pathogenesis of inflammatory and autoimmune diseases. Dysregulated Th17 cells contribute to inflammatory and autoimmune diseases. Candidate biologics are in development for targeting IL-17, IL-17 receptors or IL-17 pathways. Several drugs that impact the IL-17 pathway are already in clinical trials for the treatment of autoimmune diseases. In this review we provide evidence for the role of Th17 cells in immune-mediated diseases. An understanding of the role of Th17 in these conditions will provide important insights and unravel novel targets for therapeutic intervention.
Doxorubicin-based chemotherapy induces cardiotoxicity, which limits its clinical application. We previously reported the protective effects of quercetin against doxorubicin-induced hepatotoxicity. In this study, we tested the effects of quercetin on the expression of Bmi-1, a protein regulating mitochondrial function and ROS generation, as a mechanism underlying quercetin-mediated protection against doxorubicin-induced cardiotoxicity.
Cancer, a major health problem, affects 12 million people worldwide every year. With surgery and chemo-radiation the long term survival rate for the majority of cancer patients is dismal. Thus novel treatments are urgently needed. Immunotherapy, the harnessing of the immune system to destroy cancer cells is an attractive option with potential for long term anti-tumor benefit. Cytokines are biological response modifiers that stimulate anti-tumor immune responses. In this review, we discuss the anti-tumor efficacy of the chemotactic cytokine CCL21 and its pre-clinical and clinical application in cancer.
Non-small cell lung cancers (NSCLC) are highly heterogeneous at the molecular level and comprise 75% of all lung tumors. We have previously shown that the receptor tyrosine kinase (RTK) MET frequently suffers gain-of-function mutations that significantly promote lung tumorigenesis. Subsequent studies from our lab also revealed that PAX5 transcription factor is preferentially expressed in small cell lung cancer (SCLC) and promotes MET transcription. PAX8, however, is also expressed in NSCLC cell lines. We therefore investigated the role of PAX8 in NSCLC.
Aberrant expression of microRNAs (miRNA) with oncogenic capacities (oncomiRs) has been described for several different malignancies. The first identified oncomiR, miR-17-92, is frequently overexpressed in a variety of cancers and its targets include the tumor suppressor PTEN. The transcription factor c-Myc (MYC) plays a central role in proliferative control and is rapidly upregulated upon mitogenic stimulation. Expression of c-Myc is frequently deregulated in tumors, facilitating proliferation and inhibiting terminal differentiation. The c-Myc-regulated network comprises a large number of transcripts, including those encoding miRNAs. Here, prostaglandin E2 (PGE2) exposure rapidly upregulates the expression of the MYC gene followed by the elevation of miR-17-92 levels, which in turn suppresses PTEN expression, thus enhancing apoptosis resistance in non-small cell lung cancer (NSCLC) cells. Knockdown of MYC expression or the miR-17-92 cluster effectively reverses this outcome. Similarly, miR-17-92 levels are significantly elevated in NSCLC cells ectopically expressing COX-2. Importantly, circulating miR-17-92 was elevated in the blood of patients with lung cancer as compared with subjects at risk for developing lung cancer. Furthermore, in patients treated with celecoxib, miR-17-92 levels were significantly reduced. These data demonstrate that PGE2, abundantly produced by NSCLC and inflammatory cells in the tumor microenvironment, is able to stimulate cell proliferation and promote resistance to pharmacologically induced apoptosis in a c-Myc and miR-17-92-dependent manner.
Tumors treated with fractionated doses of ionizing radiation (IR) often acquire radioresistance. Although histone deacetylase inhibitors (HDIs) have been demonstrated to sensitize intrinsic radioresistant cancer cell lines to IR, little is known on the impact of HDIs on the effects of IR in acquired radioresistant cancer cells. This study evaluates the mechanisms by which HDIs sensitize acquired radioresistant esophageal squamous cell carcinoma cells to IR. The HDIs trichostatin A and sodium butyrate were tested for the irability to sensitize acquired radioresistant KYSE-150R and radiosensitive KYSE-150 parental cells to IR. Although the HDIs induced similar levels of cytotoxicity in the KYSE-150 and the KYSE-150R cells, HDIs increased the: (i) radiosensitivity, (ii) IR-induced ROS generation, and (iii) IR-induced G2/M arrest and apoptosis of KYSE-150R cells compared with those of KYSE-150 cells. These changes were accompanied by increased p21 expression and decreased mitochondrial membrane potential. When combined with IR, HDIs inhibited Bmi-1 expression in KYSE-150R cells and their ability to repair DNA damage. The results demonstrate the potential utility of HDIs in augmenting the efficacy of fractionated radiotherapy.
Malignant pleural mesothelioma (MPM) is a rare malignancy of the pleura that is frequently resistant to conventional therapies. Immunotherapy is a promising investigational approach for MPM that has shown some evidence of clinical benefit in select patients. However, tumor-induced immunosuppression is likely a major impediment to achieving optimal clinical responses to immunotherapeutic intervention. MPM contains a variable degree of infiltrating T-regulatory cells and M2 macrophages, which are believed to facilitate tumor evasion from the host immune system. Additional immunosuppressive factors identified in other human tumor types, such as tumor-associated programmed death ligand-1 expression, may be relevant for investigation in MPM. Conventional cytoreductive therapies, such as radiation, chemotherapy, and surgery, may play a critical role in successful immunotherapeutic strategies by ablating intratumoral and/or systemic immunosuppressive factors, thus creating a host environment more amenable to immunotherapy. This article reviews the immunotherapeutic approaches being evaluated in patients with MPM and discusses how immunotherapy might be rationally combined with conventional tumor cytoreductive therapies for this disease.
Interleukin-27 signaling is mediated by the JAK-STAT pathway via activation of STAT1 and STAT3, which have tumor suppressive and oncogenic activities, respectively. Epithelial-mesenchymal transition (EMT) and angiogenesis are key processes in carcinogenesis. Although IL-27 has been shown to have potent anti-tumor activity in various cancer models, the role of IL-27 in EMT and angiogenesis is poorly understood. In this study, we investigated the role of IL-27 in regulating EMT and angiogenesis through modulation of the STAT pathways in human non-small cell lung carcinoma (NSCLC) cells.
miRNAs are small non-coding RNA molecules that modulate the expression of multiple protein-encoding genes at the post-transcriptional level. They have recently been recognized as powerful regulators of numerous genes and pathways in the pathogenesis of inflammatory and autoimmune diseases. The targets of most miRNAs remain unknown and their roles in biological processes such as cell differentiation, proliferation, and death (apoptosis) are not clearly understood. In this review we will discuss how certain candidate miRNAs affect inflammatory and immune mediated diseases by regulating their cellular and molecular targets. We focused the influence of gender and sex hormones on miRNA. We believe that understanding the role of miRNAs could shed light on the cause and progression of many inflammatory and autoimmune diseases and eventually lay the groundwork for therapeutic options.
We have identified an alternatively spliced, non-functional aberrant E-cadherin transcript that lacks exon 11 and is over expressed in malignant cells as compared to the normal non-malignant cells. This increase in the aberrant transcript is a mechanism of loss of E-cadherin gene expression as it is rapidly degraded by the nonsense mediated decay pathway. To study the mechanism of this gene missplicing we analyzed the role of histone epigenetic modifications in lung cancer cell lines. The treatment of low E-cadherin lung cancer cell lines with histone deacetylase inhibitor (HDACi, MS-275) resulted in the preferential expression of the correctly spliced transcripts in the low E-cadherin expressing cell lines only. Chromatin immunoprecipitation (ChIP) assays revealed that the histone hypoacetylation levels correlate with aberrant exon 11 splicing as there is more aberrant splicing in cell lines with E-cadherin promoter hypoacetylation. Inactivation of histone deacetylases (HDAC) 1, 2 and 3 resulted in an increase in E-cadherin expression and an increase in the ratio of the correctly spliced E-cadherin transcript. As transcription of the gene is closely linked to splicing, we considered the possibility that change in E-cadherin transcription correlates with splicing. The Zeb1 epithelial-mesenchymal transformation (EMT) inducer silences E-cadherin expression and could also alter the splicing of this exon. Inhibition of the E-cadherin promoter transcription with Zeb1 expression increases aberrant splicing and the reverse is observed when Zeb1 is knocked down. The role of HDAC inhibitors was also studied in vivo in a immunodeficient mouse xenograft model. Exposure of mice to HDACi resulted in growth inhibition, increase in E-cadherin expression, alteration of aberrant splicing and the reversal of EMT in mouse tumors. The findings support the modulation of E-cadherin exon 11 inclusion or exclusion by histone epigenetic modifications as they change the overall chromatin structure. The results provide an interesting link between epigenetic alterations in cancer cells and gene splicing in addition to their effect on gene silencing.
The EGFR tyrosine kinase inhibitors (TKIs) demonstrate efficacy in NSCLC patients whose tumors harbor activating EGFR mutations. However, patients who initially respond to EGFR TKI treatment invariably develop resistance to the drugs. Known mechanisms account for approximately 70% of native and acquired EGFR TKI resistance. In the current study we investigated a novel mechanism of NSCLC resistance to erlotinib. Experimental Design: The mechanisms of acquired erlotinib resistance were evaluated by microarray analysis in thirteen NSCLC cell lines and in vivo in mice. Correlations between plasma neutrophil gelatinase associated lipocalin (NGAL) levels, erlotinib response and the EGFR mutational status were assessed in advanced stage NSCLC patients treated with erlotinib.
Lung cancer is the most common cause of cancer mortality worldwide for both men and women, causing approximately 1.2 million deaths per year. With the existing therapeutic efforts, the long-term survival for lung cancer patients remains low with only 15% surviving for 5 years following diagnosis. Therefore, new therapeutic strategies are needed. One such approach is the development of immune therapy for lung cancer. Immune approaches for lung cancer remain attractive because although surgery, chemotherapy and radiotherapy alone or in combination have response rates in all histological types of lung cancer, relapse is frequent. Immunologic targeting of lung cancer has the potential for nontoxic and specific therapy. Strategies that harness the immune system to react against tumors can be integrated with existing forms of therapy for optimal responses toward this devastating disease.
Based on our preclinical findings, we are assessing the efficacy of intratumoral injection of dendritic cells (DC) transduced with an adenoviral vector expressing the secondary lymphoid chemokine (CCL21) gene (Ad-CCL21-DC) in a phase I trial in advanced non-small cell lung cancer (NSCLC). While this approach shows immune enhancement, the preparation of autologous DC for CCL21 genetic modification is cumbersome, expensive and time consuming. We are evaluating a non-DC based approach which utilizes vault nanoparticles for intratumoral CCL21 delivery to mediate antitumor activity in lung cancer.
Chemokines (ie, chemoattractant cytokines) are a family of small secreted molecules that mediate leukocyte migration. It is becoming increasingly more evident that chemokines play an integral role in the initiation of a specific immune response. With respect to cancer, chemokines are being studied for both their role in tumor biology and as promising immunotherapy candidates. We review several areas of chemokine importance in tumor immunity and discuss the experimental evidence that is leading to the clinical use of this cytokine family in new treatment approaches for patients with cancer.
CD4(+)CD25(bright) regulatory T cells (T(reg)) play an important role in cancer-mediated immunosuppression. We and others have previously shown that prostaglandin E2 (PGE(2)) and transforming growth factor beta (TGF-beta) induce CD4(+)CD25(bright)FOXP3(+)T(reg). Based on these studies, we investigated the requirement for PGE(2) in Treg induction by TGF-beta. TGF-beta stimulation of human CD4(+) T cells induced COX-2-dependent production of PGE(2). PGE(2)-neutralizing antibody treatment significantly reduced the suppressive function of TGF-beta-induced T(reg) (TGF-beta-T(reg)) in vitro. TGF-beta concentration measured in the plasma of non-small cell lung cancer (NSCLC) patients directly correlated with the frequency of circulating CD4(+)CD25(bright)FOXP3(+)T cells. Flow cytometry analysis showed increased FOXP3 expression in circulating CD4(+)CD25(+)HLA-DR- cells of lung cancer patients compared to control subjects. Immunohistochemical analysis revealed co-expression of TGF-beta, COX-2, and FOXP3 in serial sections from resected lung tumor tissues. All together these observations suggest interplay between TGF-beta and COX-2 in the induction of T(reg) activities. Interrupting TGF-beta and PGE(2) signaling may be important in therapeutic interventions that aim to limit T(reg)function in lung cancer.
With their resistance to genotoxic and anti-proliferative drugs and potential to grow tumors and metastases from very few cells, cancer stem or tumor-initiating cells (TICs) are a severe limitation for the treatment of cancer by conventional therapies. Here, we explored whether human T cells that are redirected via an EpCAM/CD3-bispecific antibody called MT110 can lyse colorectal TICs and prevent tumor growth from TICs. MT110 recognizes EpCAM, a cell adhesion molecule expressed on TICs from diverse human carcinoma, which was recently shown to promote tumor growth through engagement of elements of the wnt pathway. MT110 was highly potent in mediating complete redirected lysis of KRAS-, PI3 kinase- and BRAF-mutated colorectal TICs, as demonstrated in a soft agar assay. In immunodeficient mice, MT110 prevented growth of tumors from a 5,000-fold excess of a minimally tumorigenic TIC dose. T cells engaged by MT110 may provide a potent therapeutic means to eradicate TICs and bulk tumor cells derived thereof.
As a transcriptional repressor of E-cadherin, Snail has predominantly been associated with epithelial-mesenchymal transition, invasion, and metastasis. However, other important Snail-dependent malignant phenotypes have not been fully explored. Here, we investigate the contributions of Snail to the progression of non-small cell lung cancer (NSCLC).
Inflammatory cytokines have been implicated in the progression of head and neck squamous cell carcinoma (HNSCC). Herein we investigate the mechanisms by which interleukin-1beta (IL-1beta) might contribute to Epithelial-Mesenchymal Transition (EMT) in HNSCC.
The coxsackie-adenovirus receptor (CAR) is a developmentally regulated intercellular adhesion molecule that was previously observed to be required for efficient tumor formation. To confirm that observation, we compared the tumorigenicity of clonally derived test and control cell subsets that were genetically modified for CAR. Silencing CAR in lung cancer cells with high constitutive expression reduced engraftment efficiency. Conversely, overexpressing CAR in lung cancer cells with low constitutive expression did not affect tumor formation or growth kinetics. A blocking antibody to the extracellular domain of CAR inhibited tumor engraftment, implicating that domain as being important to this process. However, differences in adhesion properties attributable to this domain (barrier function and aggregation) could not be distinguished in the test groups in vitro, and the mechanisms underlying CARs contribution to tumor engraftment remain elusive. Because high CAR cells displayed a spindle-shaped morphology at baseline, we considered whether this expression was an accompaniment of other mesenchymal features in these lung cancer cells. Molecular correlates of CAR were compared in model epithelial and mesenchymal type lung cancer cells. CAR expression is associated with an absence of E-cadherin, diminished expression of alpha- and gamma-catenin, and increased Zeb1, Snail, and vimentin expression in lung cancer cells. In contrast, epithelial type (NCI-H292, Calu3) lung cancer cells show comparatively low CAR expression. These data suggest that if the mesenchymal cell phenotype is an accurate measure of an undifferentiated and invasive state, then CAR expression may be more closely aligned with this phenotype of lung cancer cells.
We are evaluating the immune enhancing activities of cytokines for their optimum utility in augmenting cellular immune responses against lung cancer. In this study, we evaluated the mechanism of antitumor responses following IL-7 administration to mice bearing established Lewis lung cancer. IL-7 decreased tumor burden with concomitant increases in the frequency of CD4 and CD8 T lymphocyte subsets, T cell activation markers CXCR3, CD69, and CD127(low), effector memory T cells, and T cell cytolytic activity against parental tumor cells. Accompanying the antitumor responses were increases in IFN-gamma, CXCL9, CXCL10, and IL-12. Individual neutralization of CD4, CD8 T lymphocytes, or the CXCR3 ligands CXCL9 and CXCL10 reversed the antitumor benefit of IL-7, indicating their importance for optimal responses in vivo. Furthermore, IL-7 decreased the tumor-induced apoptosis of T cells with subsequent decrease of the proapoptotic marker Bim. We assessed the impact of IL-7 treatment on regulatory T cells that negatively impact antitumor immune responses. IL-7 decreased regulatory T Foxp3 as well as cell suppressive activity with a reciprocal increase in SMAD7. These results indicate that IL-7 induces CXCR3 ligand-dependent T cell antitumor reactivity in lung cancer.
Malignant Pleural Effusions (MPE) may be useful as a model to study hierarchical progression of cancer and/or intratumoral heterogeneity. To strengthen the rationale for developing the MPE-model for these purposes, we set out to find evidence for the presence of cancer stem cells (CSC) in MPE and demonstrate an ability to sustain intratumoral heterogeneity in MPE-primary cultures. Our studies show that candidate lung CSC-expression signatures (PTEN, OCT4, hTERT, Bmi1, EZH2 and SUZ12) are evident in cell pellets isolated from MPE, and MPE-cytopathology also labels candidate-CSC (CD44, cMET, MDR-1, ALDH) subpopulations. Moreover, in primary cultures that use MPE as the source of both tumor cells and the tumor microenvironment (TME), candidate CSC are maintained over time. This allows us to live-sort candidate CSC-fractions from the MPE-tumor mix on the basis of surface markers (CD44, c-MET, uPAR, MDR-1) or differences in xenobiotic metabolism (ALDH). Thus, MPE-primary cultures provide an avenue to extract candidate CSC populations from individual (isogenic) MPE-tumors. This will allow us to test whether these cells can be discriminated in functional bioassays. Tumor heterogeneity in MPE-primary cultures is evidenced by variable immunolabeling, differences in colony-morphology, and differences in proliferation rates of cell subpopulations. Collectively, these data justify the ongoing development of the MPE-model for the investigation of intratumoral heterogeneity, tumor-TME interactions, and phenotypic validation of candidate lung CSC, in addition to providing direction for the pre-clinical development of rational therapeutics.
Myeloid-derived suppressor cells (MDSCs) are important regulators of immune responses. These cells suppress the cytotoxic activities of natural killer (NK)-cell and T-cell effectors and promote tumor growth. We demonstrated that depleting MDSCs improve therapeutic responses to vaccination in a murine model of lung cancer. This approach may prove beneficial against tumors in which MDSC exert prominent immunosuppressive effects.
The dose-dependent toxicities of doxorubicin (DOX) limit its clinical applications, particularly in drug-resistant cancers, such as liver cancer. In this study, we investigated the role of quercetin on the antitumor effects of DOX on liver cancer cells and its ability to provide protection against DOX-mediated liver damage in mice.
The role of c-Crk (CRK) in promoting metastasis is well described however the role of CRK phosphorylation and the corresponding signaling events are not well explained. We have observed CRK-II serine 41 phosphorylation is inversely correlated with p120-catenin and E-cadherin expressions in non-small cell lung cancer (NSCLC) cells. Therefore, we investigated the role of CRK-II serine 41 phosphorylation in the down-regulation of p120-catenin, cell motility and cell invasiveness in NSCLC cells. For this purpose, we expressed phosphomimetic and phosphodeficient CRK-II serine 41 mutants in NSCLC cells. NSCLC cells expressing phosphomimetic CRK-II seine 41 mutant showed lower p120-catenin level while CRK-II seine 41 phosphodeficient mutant expression resulted in higher p120-catenin. In addition, A549 cells expressing CRK-II serine 41 phosphomimetic mutant demonstrated more aggressive behavior in wound healing and invasion assays and, on the contrary, expression of phosphodeficient CRK-II serine 41 mutant in A549 cells resulted in reduced cell motility and invasiveness. We also provide evidence that PAK1 mediates CRK-II serine 41 phosphorylation. RNAi mediated silencing of PAK1 increased p120-catenin level in A549 and H157 cells. Furthermore, PAK1 silencing decreased cell motility and invasiveness in A549 cells. These effects were abrogated in A549 cells expressing phosphomimetic CRK-II serine 41. In summary, these data provide evidence for the role of PAK1 in the promotion of cell motility, cell invasiveness and the down regulation of p120-catenin through CRK serine 41 phosphorylation in NSCLC cells.
Radiotherapy (RT) is a major modality of cancer treatment. However, tumors often acquire radioresistance, which causes RT to fail. The exact mechanisms by which tumor cells subjected to fractionated irradiation (FIR) develop an adaptive radioresistance are largely unknown. Using the radioresistant KYSE-150R esophageal squamous cell carcinoma (ESCC) model, which was derived from KYSE-150 parental cells using FIR, the role of Bmi-1 in mediating the radioadaptive response of ESCC cells to RT was investigated. The results showed that the level of Bmi-1 expression was significantly higher in KYSE-150R cells than in the KYSE-150 parental cells. Bmi-1 depletion sensitized the KYSE-150R cells to RT mainly through the induction of apoptosis, partly through the induction of senescence. A clonogenic cell survival assay showed that Bmi-1 depletion significantly decreased the radiation survival fraction in KYSE-150R cells. Furthermore, Bmi-1 depletion increased the generation of reactive oxygen species (ROS) and the expression of oxidase genes (Lpo, Noxo1 and Alox15) in KYSE-150R cells exposed to irradiation. DNA repair capacities assessed by ?-H2AX foci formation were also impaired in the Bmi-1 down-regulated KYSE-150R cells. These results suggest that Bmi-1 plays an important role in tumor radioadaptive resistance under FIR and may be a potent molecular target for enhancing the efficacy of fractionated RT.
Myeloid derived suppressor cells (MDSC) are important regulators of immune responses. We evaluated the mechanistic role of MDSC depletion on antigen presenting cell (APC), NK, T cell activities and therapeutic vaccination responses in murine models of lung cancer.
Modifications of adjuvants that induce cell-mediated over antibody-mediated immunity is desired for development of vaccines. Nanocapsules have been found to be viable adjuvants and are amenable to engineering for desired immune responses. We previously showed that natural nanocapsules called vaults can be genetically engineered to elicit Th1 immunity and protection from a mucosal bacterial infection. The purpose of our study was to characterize immunity produced in response to OVA within vault nanoparticles and compare it to another nanocarrier.
Chronic neuroinflammation is an important component of Alzheimers disease and could contribute to neuronal dysfunction, injury and loss that lead to disease progression. Multiple clinical studies implicate tumor necrosis factor-? as an inflammatory mediator of neurodegeneration in patients with Alzheimers because of elevated levels of this cytokine in the cerebrospinal fluid, hippocampus and cortex. Current Alzheimers disease interventions are symptomatic treatments with limited efficacy that do not address etiology. Thus, a critical need exists for novel treatments directed towards modifying the pathophysiology and progression.
The presence of regional metastases in patients with head and neck squamous cell carcinoma (HNSCC) is a common and adverse event associated with poor prognosis. The authors recent work on human HNSCC tissues underlies Snails role as a molecular prognostic marker for HNSCC. Snail positivity is significantly predictive of poorly differentiated, lymphovascular invasive, and regionally metastatic tumors. Here, the authors investigate the capacity of Snail to drive epithelial-mesenchymal transition (EMT) in human oral epithelial cell lines and its ability to confer drug resistance.
Many tumors, including lung cancers, promote immune tolerance to escape host immune surveillance and facilitate tumor growth. Tumors utilize numerous pathways to inhibit immune responses, including the elaboration of immune-suppressive mediators such as PGE2, TGF-?, IL-10, VEGF, GM-CSF, IL-6, S100A8/A9 and SCF, which recruit and/or activate myeloid-derived suppressor cells (MDSCs). MDSCs, a subset of heterogeneous bone marrow-derived hematopoietic cells, are found in the peripheral blood of cancer patients and positively correlate to malignancy. Solid tumors contain MDSCs that maintain an immune-suppressive network in the tumor microenvironment. This review will focus on the interaction of tumors with MDSCs that lead to dysregulation of antigen presentation and T-cell activities in murine tumor models. Specific genetic signatures in lung cancer modulate the activities of MDSCs and impact tumor progression. Targeting MDSCs may have a long-term antitumor benefit and is at the forefront of anticancer therapeutic strategies.
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