Malignant Pleural Effusion Formation Research Articles

Switching off malignant pleural effusion formation-fantasy or future?

Malignant pleural effusion (MPE) is common and difficult to treat. In the vast majority of patients the presence of MPE heralds incurable disease, associated with poor quality of life, morbidity and mortality. Current therapeutic approaches are inefficient and merely offer palliation of associated symptoms. Recent scientific progress has shed light in the biologic processes governing the mechanisms behind the pathobiology of MPE. Pleural based tumors interfere with pleural fluid drainage, as well as the host vasculature and immune system, resulting in decreased fluid absorption and increased pleural fluid production via enhanced plasma extravasation into the pleural space. In order to achieve this feat, pleural based tumors must elicit critical vasoactive events in the pleura, thus forming a favorable microenvironment for tumor dissemination and MPE development. Such properties involve specific transcriptional signaling cascades in addition to secretion of important mediators which attract and activate host cell populations which, in turn, impact tumor cell functions. The dissection of the biologic steps leading to MPE formation provides novel therapeutic targets and recent research findings provide encouraging results towards future therapeutic innovations in MPE management.

Effect of Endostar combined with angiopoietin-2 inhibitor on malignant pleural effusion in mice.

The present study was designed to investigate the synergetic effect of Endostar combined with an angiopoietin-2 specific inhibitor L1-10 on malignant pleural effusion (MPE) mouse model. A MPE mouse model was established by injecting Lewis lung carcinoma (LLC) cells into pleural cavity of C57BL/6 mice. The mice were randomly divided into four treatment groups: saline, Endostar, L1-10, and Endostar+L1-10. In the present study, we reported for the first time that Endostar combined with L1-10 had significant synergistic effects on the formation of MPE and tumor growth. Moreover, Endostar combined with L1-10 had additive effect on the attenuation of pleural inflammation, inhibition of tumor angiogenesis and pleural vascular hyperpermeability, which are central to the inhibition of MPE. Finally, these studies also found that Endostar combined with L1-10 could have complementary actions by reducing VEGF and IL-6 local release and downregulating VEGF expression in pleural tumors, which involved in the pathogenesis of MPE. Therefore, combined therapy with Endostar and L1-10 may be an encouraging strategy for the treatment of MPE.

Interplay of Th1 and Th17 Cells in Murine Models of Malignant Pleural Effusion

IFN-γ-producing CD4(+) T (Th1) cells and IL-17-producing CD4(+) T (Th17) cells have been found to be involved in multiple malignancies; however, the reciprocal relationship between Th1 and Th17 cells in malignant pleural effusion (MPE) remains to be elucidated. To explore the differentiation and immune regulation of Th1 and Th17 cells in the development of MPE in murine models. The distribution and differentiation of Th1 and Th17 cells in MPE were investigated in IFN-γ(-/-), IL-17(-/-), and wild-type mice. The effects of Th1 and Th17 cells on the development of MPE and the survival of mice bearing MPE were also investigated. We have demonstrated that increased Th1 and Th17 cells could be found in MPE as compared with blood and spleen. Compared with wild-type mice, Th17 cells were markedly augmented in MPE from IFN-γ(-/-) mice, and improved survival could be seen in IFN-γ(-/-) mice. Th1 cell numbers were elevated in MPE from IL-17(-/-) mice, and decreased survival could be seen in IL-17(-/-) mice. The in vitro experiments showed that IFN-γ deficiency promoted Th17-cell differentiation by suppressing the STAT3 pathway and that IL-17 deficiency promoted Th1-cell differentiation by suppressing the STAT1 pathway. In mouse models of MPE, IFN-γ inhibited Th17-cell differentiation, whereas IL-17 inhibited Th1-cell differentiation. IL-17 inhibited the formation of MPE and improved the survival of mice bearing MPE; in contrast, IFN-γ promoted MPE formation and mouse death.

Beneficial Impact of CCL2 and CCL12 Neutralization on Experimental Malignant Pleural Effusion

Using genetic interventions, we previously determined that C-C motif chemokine ligand 2 (CCL2) promotes malignant pleural effusion (MPE) formation in mice. Here we conducted preclinical studies aimed at assessing the specific therapeutic potential of antibody-mediated CCL2 blockade against MPE. For this, murine MPEs or skin tumors were generated in C57BL/6 mice by intrapleural or subcutaneous delivery of lung (LLC) or colon (MC38) adenocarcinoma cells. Human lung adenocarcinoma cells (A549) were used to induce MPEs in severe combined immunodeficient mice. Intraperitoneal antibodies neutralizing mouse CCL2 and/or CCL12, a murine CCL2 ortholog, were administered at 10 or 50 mg/kg every three days. We found that high doses of CCL2/12 neutralizing antibody treatment (50 mg/kg) were required to limit MPE formation by LLC cells. CCL2 and CCL12 blockade were equally potent inhibitors of MPE development by LLC cells. Combined CCL2 and CCL12 neutralization was also effective against MC38-induced MPE and prolonged the survival of mice in both syngeneic models. Mouse-specific CCL2-blockade limited A549-caused xenogeneic MPE, indicating that host-derived CCL2 also contributes to MPE precipitation in mice. The impact of CCL2/12 antagonism was associated with inhibition of immune and vascular MPE-related phenomena, such as inflammation, new blood vessel assembly and plasma extravasation into the pleural space. We conclude that CCL2 and CCL12 blockade are effective against experimental MPE induced by murine and human adenocarcinoma in mice. These results suggest that CCL2-targeted therapies may hold promise for future use against human MPE.

The role of vascular endothelial growth factor in the pathogenesis, diagnosis and treatment of malignant pleural effusion.

Malignant pleural effusions (MPEs) are a significant source of cancer-related morbidity. Over 150,000 patients in the United States suffer from breathlessness and diminished quality of life due to MPE each year. Current management strategies are of mostly palliative value and focus on symptom control; they do not address the pathobiology of the effusion, nor do they improve survival. Further elucidation of the pathophysiological mechanisms, coupled with the development of novel treatments such as intrapleural chemotherapeutics targeting this process, has the potential to greatly improve the efficacy of our current management options. Vascular endothelial growth factor-A (VEGF-A) has been implicated as a critical cytokine in the formation of malignant pleural effusions. Elevated levels of VEGF produced by tumor cells, mesothelial cells, and infiltrating immune cells result in increased vascular permeability, cancer cell transmigration, and angiogenesis. Therefore antiangiogenic therapies such as Bevacizumab, a monoclonal antibody targeting VEGF-A, may have a potential role in the management of malignant pleural effusions. Herein we review the pathogenesis and potential treatment strategies of malignant pleural effusions, with a focus on angiogenesis and antiangiogenic therapeutics.

Establishment of a malignant pleural effusion mouse model with lewis lung carcinoma cell lines expressing enhanced green fluorescent protein.

Malignant pleural effusion (MPE) is a poor prognostic factor in patients with advanced lung cancer. The aim of this study is to establish a mouse model of MPE using Lewis lung carcinoma (LLC) cell lines expressing enhanced green fluorescent protein (EGFP). The mouse model was created by injecting LLC-EGFP cells directly into the pleural cavity of nude mice under the guidance of stereomicroscope and then mice were sacrificed periodically. The dynamic growth and metastasis of tumor cells were screened using in vivo fluorescence imaging. The remaining mice were subjected to transverse computed tomography (CT) periodically to analyze the rate of MPE formation. The survival rate and tumor metastasis were also observed after modeling. Pleural fluid was gently aspirated using a 1 mL syringe and its volume was measured. When two or more mice bore MPE at the same time, we calculated the average volume. The correlation of MPE with the integrated optical density (IOD) were analyzed. Four days after the inoculation of LLC-EGFP cells, green fluorescence was observed by opening the chest wall. The tumor formation rate was 100%, and the IOD gradually increased after inoculation. The metastatic foci were mediastinal, contralateral pleural and pericardial. The metastasis rates were 87%, 73%, and 20%, respectively. CT imagings revealed that the rates of MPE formation on days 7, 14 and 21 were 13%, 46%, and 53%. The mean survival time of nude mice was 28.8 days. The average MPE volume increased obviously on day 10 and peaked on day 16 with a value of 0.5 mL. The MPE volume and IOD were significantly correlated (r=0.91, P<0.0001). This study was the first to establish a mouse model of MPE by injecting LLC-EGFP into the pleural cavity under the guidance of a stereomicroscope. The model can enable dynamic observations of the biological behavior of tumor cells in the pleural cavity. It might be helpful for basic research on advanced lung cancer as well as anti-tumor drug development.

Malignant pleural effusion: tumor-host interactions unleashed.

Malignant pleural effusion (MPE) poses a significant clinical problem. Current nonetiologic management is suboptimal in terms of efficacy and safety. In light of recent research progress, we propose herein a new view of MPE development, which may rapidly translate into meaningful changes in therapeutics. In addition to tumor-induced impairment of pleural fluid drainage, pertinent findings point toward another pathway to MPE formation: a vicious loop of interactions between pleural-based tumor cells and the host vasculature and immune system that results in increased net fluid production via enhanced plasma extravasation into the pleural space. The ability of tumor cells to trigger this cascade likely rests on a specific and distinct transcriptional repertoire, which results in important vasoactive events in the pleural space. Although the characterization of tumor-derived factors responsible for MPE development is in the making, an additional, indirect path to MPE was recently demonstrated: tumor cells recruit and co-opt host cells and mediators, which, in turn, amplify tumor cell-primed fluid leakage and impact tumor cell functions. Importantly, recent evidence suggests that the biologic events that culminate in clinical MPE are likely amenable to therapeutic inhibition and even prevention. In this perspective, the scientific basis for an update of current concepts of MPE formation is highlighted. Key questions for future research are posed. Finally, a vision for novel, effective, safe, and convenient treatment modalities that can be offered to outpatients with MPE is set forth.

Secreted phosphoprotein-1 directly provokes vascular leakage to foster malignant pleural effusion

Secreted phosphoprotein-1 (SPP1) promotes cancer cell survival and regulates tumor-associated angiogenesis and inflammation, both central to the pathogenesis of malignant pleural effusion (MPE). Here, we examined the impact of tumor- and host-derived SPP1 in MPE formation and explored the mechanisms by which the cytokine exerts its effects. We used a syngeneic murine model of lung adenocarcinoma-induced MPE. To dissect the effects of tumor- versus host-derived SPP1, we intrapleurally injected wild-type and SPP1-knockout C57/BL/6 mice with either wild-type or SPP1-deficient syngeneic lung cancer cells. We demonstrated that both tumor- and host-derived SPP1 promoted pleural fluid accumulation and tumor dissemination in a synergistic manner (P<0.001). SPP1 of host origin elicited macrophage recruitment into the cancer-affected pleural cavity and boosted tumor angiogenesis, whereas tumor-derived SPP1 curtailed cancer cell apoptosis in vivo. Moreover, the cytokine directly promoted vascular hyper-permeability independently of vascular endothelial growth factor. In addition, SPP1 of tumor and host origin differentially affected the expression of proinflammatory and angiogenic mediators in the tumor microenvironment. These results suggest that SPP1 of tumor and host origin impact distinct aspects of MPE pathobiology to synergistically promote pleural fluid formation and pleural tumor progression. SPP1 may present an attractive target of therapeutic interventions for patients with MPE.

A sulindac analogue is effective against malignant pleural effusion in mice

Objectives To examine whether a sulindac derivative (C-18) with previously reported anti-angiogenic properties limits malignant pleural effusion (MPE) formation in mice. Methods MPE was generated by intrapleural injection of murine adenocarcinoma cells in C57BL/6 mice. Animals were divided into three groups, a control group and two treatment groups receiving intraperitoneally a daily dose of either 1 mg or 2 mg of C-18 for a total of 12 doses. Mice were sacrificed on day 14. Measurements and main results Pleural fluid volume and the number of pleural tumor implantations were measured. Tumor angiogenesis, pleural vascular permeability and the host inflammatory response were also assessed. C-18 significantly limited pleural fluid formation and inhibited intrapleural tumor dissemination. The mean ± SEM pleural fluid volume was 758 ± 63 μl for the control group, compared to 492 ± 120 μl ( p = 0.042) and 279 ± 77 μl ( p < 0.001) for the low dose and high dose group of C-18, respectively. Control group animals had 6.2 ± 1 intrapleural tumors, while C-18 treated animals had 3.1 ± 0.8 ( p = 0.014) and 3 ± 0.7 ( p = 0.009) for the low and high dose respectively. In addition C-18 significantly suppressed pleural vascular permeability. No significant difference in tumor angiogenesis and inflammatory response was observed, while there was also no measurable effect in tumor cell apoptosis and proliferation in vitro and in vivo. Conclusions C-18 halted experimental MPE formation and intrapleural tumor dissemination, through down-regulation of pleural vascular permeability.

Malignant pleural effusion cells show aberrant glucose metabolism gene expression

Malignant pleural effusion (MPE) accompanying lung adenocarcinoma indicates poor prognosis and early metastasis. This study aimed to identify genes related to MPE formation. Three tissue sample cohorts, seven from healthy lungs, 18 from stage I-III lung adenocarcinoma with adjacent healthy lung tissue and 13 from lung adenocarcinomas with MPE, were analysed by oligonucleotide microarray. The identified genes were verified by quantitative real-time PCR (qRT-PCR), immunohistochemical staining, and immunofluorescence confocal microscopy. 20 up- or down-regulated genes with a two-fold change in MPE cancer cells compared to healthy tissues were differentially expressed from early- to late-stage lung cancer. Of 13 genes related to cellular metabolism, aldolase A (ALDOA), sorbitol dehydrogenase (SORD), transketolase (TKT), and tuberous sclerosis 1 (TSC1) were related to glucose metabolism. qRT-PCR validated their mRNA expressions in pleural metastatic samples. Immunohistochemical staining confirmed aberrant TKT, ALDOA, and TSC1 expressions in tumour cells. Immunofluorescence confirmed TKT co-localisation and co-distribution of ALDOA with thyroid transcription factor 1-positive cancer cells. TKT regulated the proliferation, vascular endothelial growth factor secretion in vitro and in vivo vascular permeability of cancer cell. Glucose metabolic reprogramming by ALDOA, SORD, TKT and TSC1 is important in MPE pathogenesis.

Abstract 71: Metformin inhibits lung cancer cell growth and angiogenesis in vitro and in vivo: Effect on IL6-Stat3 pathway

Abstract The usage of metformin as a first line treatment for type 2 diabetes associates with reduced cancer risk. Animal studies have shown that metformin inhibits breast cancer growth mainly via AMPK-mTOR pathway. Associated clinical trial is conducting. However, the application of metformin in other cancers and other mechanisms remain not clarified. In our previous study, we demonstrated the down-regulation of TSC1 (tuberous sclerosis 1) in lung cancer cells from malignant pleural effusion (MPE) and verified with qRT-PCR and immunohistochemistry. TSC1/TSC2 complex can suppress the mTOR pathway and receive input from LKB1-AMPK pathway and we found that metformin induced TSC1expression in vitro. Previous studies demonstrate that TNFα-sphingomyelinase and IL6-Stat3 contribute the formation of MPE via activating VEGF secretion. TNF-α can also induce IL6 secretion in many tumors. In our recent study, we first investigated the effect of metformin on proliferation of lung cancer cell. Lung cancer lines including PC14PE6/AS2 (AS2), A549 and CL 1-0 were treated with various dosage of metformin. After treatment for 96 hours, metformin inhibited proliferation of AS2, A549 and CL1-0 lung cancer cells dose-dependently. The IC50 of all cells are around 2.5-5 mM, measured by MTT assays. We also investigated the addictive cytotoxicity of metformin to cisplatin. Metformin co-incubation with cisplatin for 72 hr in AS2 cells displayed an increased cytotoxicity as compared to each agent alone. As for the association with angiogenesis, by analyzing the supernatant with ELISA assay, the VEGF secretion also decreased after treatment with metformin in AS2 and A549 cells. And Western blotting was conducted to access the effect on Stat3 and AMPK phosphorylation of metformin. In AS2 cell, not only the phosphorylation of AMPK increased after metformin treatment, the Stat3 phosphorylation was also suppressed. In addition, stimulation with TNF-α led to substantial secretion of IL6, which was inhibited by co-incubation with metformin. In vivo, the AS2 cells were injected i.p. into the unanesthetized nude mice and MQ-water or metformin (1mg) were injected 2 days later. Around 2 to 4 weeks, the mice developed peritoneal carcinomatosis and malignant ascites. In the metformin group, though the same number of mice developed peritoneal carcinomatosis and ascites, the amount of ascites and VEGF level are less than the control group. In conclusion, metformin inhibit lung cancer cell proliferation and angiogenesis in vitro and in vivo. And the IL6-Stat3-VEGF pathway may also involve in the anticancer effect of metformin. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 71.

The angiogenetic pathway in malignant pleural effusions: Pathogenetic and therapeutic implications

Increased permeability of the pleural microvasculature is generally attributed to the substances that are released in inflammatory and malignant pleural effusions, although the exact pathogenetic mechanisms of malignant pleural effusions are unclear. Current therapies used to prevent the re-accumulation of pleural fluid and relieve symptoms are of variable efficacy and may cause serious adverse effects. Understanding the mechanisms of fluid accumulation would hopefully permit the development of more specific, effective and safer treatment modalities. Angiogenesis, pleural vascular increased permeability and inflammation are considered central to the pathogenesis of malignant pleural effusions. Vascular endothelial growth factor (VEGF) is a member of the VEGF/platelet-derived factor gene family and consists of at least six isoforms. Since it was shown that VEGF contributes to the formation of malignant pleural effusions, there have been some attempts to implicate, therapeutically, this finding using different molecules (ZD6474, PTK 787 and bevacizumab). However, the role of the biological axis of VEGF and angiopoietins needs further investigation in both the pathogenesis and the treatment of malignant pleural effusion. In both non-small-cell lung carcinoma and breast cancer, it has been shown that the ligand for CXCR4, CXCL12 or SDF-1α, exhibited peak levels of expression in organs that were the preferred destination for their respective metastases. Recent findings imply that new therapeutic strategies aimed at blocking the SDF-1-CXCR4 axis may have significant applications for patients by modulating the trafficking of hemato/lymphopoietic cells and inhibiting the metastatic behavior of tumor cells as well. The purpose of this report is to review novel pathogenetic and therapeutic implications regarding the angiogenetic pathways in malignant pleural effusions.

The angiogenetic pathway in malignant pleural effusions: Pathogenetic and therapeutic implications (Review)

Increased permeability of the pleural micro-vasculature is generally attributed to the substances that are released in inflammatory and malignant pleural effusions, although the exact pathogenetic mechanisms of malignant pleural effusions are unclear. Current therapies used to prevent the re-accumulation of pleural fluid and relieve symptoms are of variable efficacy and may cause serious adverse effects. Understanding the mechanisms of fluid accumulation would hopefully permit the development of more specific, effective and safer treatment modalities. Angiogenesis, pleural vascular increased permeability and inflammation are considered central to the pathogenesis of malignant pleural effusions. Vascular endothelial growth factor (VEGF) is a member of the VEGF/platelet-derived factor gene family and consists of at least six isoforms. Since it was shown that VEGF contributes to the formation of malignant pleural effusions, there have been some attempts to implicate, therapeutically, this finding using different molecules (ZD6474, PTK 787 and bevacizumab). However, the role of the biological axis of VEGF and angiopoietins needs further investigation in both the pathogenesis and the treatment of malignant pleural effusion. In both non-small-cell lung carcinoma and breast cancer, it has been shown that the ligand for CXCR4, CXCL12 or SDF-1α, exhibited peak levels of expression in organs that were the preferred destination for their respective metastases. Recent findings imply that new therapeutic strategies aimed at blocking the SDF-1-CXCR4 axis may have significant applications for patients by modulating the trafficking of hemato/lymphopoietic cells and inhibiting the metastatic behavior of tumor cells as well. The purpose of this report is to review novel pathogenetic and therapeutic implications regarding the angiogenetic pathways in malignant pleural effusions.

VEGF-C and lung cancer

Vascular endothelial growth factor-C (VEGF-C) binds to special receptors on the vessel and lymphatic endothelium to accelerate the formation and metastasis of lung cancer. It shows specific action on malignant pleural effusion formation and immuno-regulation factor expression. It is reported that inhibition of VEGF-C would be a target point in lung cancer therapy. Key words: Vascular endothelial growth factor C ; Lung neoplasms; Pleural effusion

Therapeutic effects of recombinant human endostatin adenovirus in a mouse model of malignant pleural effusion

Malignant pleural effusion (MPE) is a common clinical problem in patients with advanced cancer. Evidence suggests that tumor-mediated angiogenesis and enhanced vascular permeability in the pleural wall are due to high levels of vascular endothelial growth factor (VEGF), which plays an important role in the pathogenesis of MPE. The present study was designed to test whether the recombinant adenovirus-mediated delivery of human endostatin (Ad-hEndo), one of the potent inhibitors of angiogenesis, would inhibit the formation and progression of MPE. We developed a novel mouse model of MPE by injecting Lewis lung carcinoma (LLC) cells directly into pleural cavity of C57BL/6 mice. To evaluate the therapeutic effects of endostatin in this MPE model, we injected the Ad-hEndo into the pleural cavity of MPE-bearing mice three times with the 3-day interval. We found that this treatment resulted in significant reduction in pleural effusion volume, the number of pleural tumor foci, microvessel density, and vascular permeability, while it significantly prolonged the survival time. In addition, VEGF level of MPE in the group administered with the Ad-hEndo was obviously decreased as compared with that in the two control groups administered with null-adenovirus (Ad-null) or normal saline. Our work provides a rationale for future studies toward evaluating the effectiveness of the adenovirus-based endostatin therapy for MPE.

A Central Role for Tumor-derived Monocyte Chemoattractant Protein-1 in Malignant Pleural Effusion

Tumor cells in malignant pleural effusions (MPEs) are an important source of monocyte chemoattractant protein (MCP)-1. However, the role of tumor-derived MCP-1 in the pathogenesis and progression of MPE has not been determined. B16 mouse skin melanoma cells, which are deficient in MCP-1 expression, and mouse Lewis lung cancer (LLC) cells, which express high levels of MCP-1, were engineered to stably express MCP-1 and short hairpin RNAs (shRNAs) targeting the MCP-1 transcript, respectively. Cells were injected into the pleural cavities of syngeneic immunocompetent mice, and MPE volume and pleural tumors were quantified at necropsy (day 14). MCP-1 and other mediators were determined by cytometric bead array and enzyme-linked immunosorbent assay, and mononuclear and endothelial cells were identified by immunolabeling of F4/80 and factor VIII-related antigen respectively. Mouse survival was assessed using Kaplan-Meier analysis. Vascular permeability in mice with MPE was assessed using albumin-binding Evans blue. Statistical tests were two-sided. LLC cells expressing shRNA against MCP-1 elaborated less than 5% of the MCP-1 level in cells expressing nonspecific shRNA (control cells), and intrapleural delivery of these cells resulted in less MPE (mean MPE volume = 86 and 585 muL, respectively; difference = 499 muL; 95% confidence interval [CI] = 331 to 669 muL; P < .001), reduced MCP-1 levels in the pleural fluid, and lower mortality than when control cells were delivered. Overexpression of MCP-1 in intrapleurally injected B16 melanoma cells led to increased MPE and reduced survival. In mice with MPE, MCP-1 was a potent inducer of vascular permeability, mononuclear recruitment, and, in pleural tumors, of angiogenesis. MCP-1 produced by tumor cells is an important determinant of their capacity to induce the formation of MPE and may be a useful target for the treatment of malignant pleural disease.

Osteopontin is involved in the formation of malignant pleural effusion in lung cancer

Malignant pleural effusion (MPE) is associated with advanced-stage lung cancer and is a poor prognostic sign for these patients. Osteopontin (OPN) is a multifunctional cytokine that is involved in the tumor progression and angiogenesis of lung cancer cells. The purpose of this study is to investigate and provide evidence for the role of OPN in the formation of MPE associated with lung cancer. In this study, we established an OPN knockdown murine lung cancer cell line, 3LL cells, utilizing the small interfering RNA (siRNA) technique. To reveal the effect of OPN on the formation of MPE associated with lung cancer, we directly injected OPN knockdown 3LL cells, 3LL/OPN siRNA, or control cells, 3LL/control siRNA, into the pleural space of C57BL/6 mice. OPN knockdown significantly reduced the formation of MPE, but did not inhibit in vivo tumor growth of 3LL cells in mice. Vascular endothelial growth factor (VEGF) concentration in MPE was markedly decreased in the 3LL/OPN siRNA in comparison with that of the 3LL/control siRNA. In vitro, recombinant OPN protein enhanced VEGF secretion from human umbilical vein endothelial cell (HUVEC) or human mesothelial cell line, Met5A cells, in a concentration-dependent manner. These results suggest that OPN is positively involved in the formation of MPE of lung cancer presumably by promoting VEGF secretion from vascular endothelial cells or mesothelial cells. OPN could be an effective target molecule for reducing MPE in lung cancer patients.

Tumor Necrosis Factor-α Promotes Malignant Pleural Effusion

Tumor necrosis factor (TNF)-alpha is present in the microenvironment of human tumors, including malignant pleural effusion (MPE). Although the cytokine is produced in the pleural cavity by both tumor and host cells, its effects on MPE formation are unknown. In these studies, we sought to determine the role of TNF-alpha in the pathogenesis of MPE and to assess the therapeutic effects of its neutralization in a preclinical model. For this, MPEs were generated in immunocompetent mice using intrapleural injection of mouse lung adenocarcinoma cells. The roles of tumor- and host-derived TNF-alpha were assessed using combined experimentation with TNF-alpha gene-deficient mice and in vivo TNF-alpha neutralization. To expand the scope of preclinical data, TNF-alpha and vascular endothelial growth factor (VEGF) expression were determined in human cancer cell lines and human MPE. In the MPE model, TNF-alpha of host and tumor origin was present. TNF-alpha neutralization significantly limited tumor dissemination, effusion formation, vascular hyperpermeability, TNF-alpha and VEGF expression, and angiogenesis, thereby improving survival. In contrast, these variables were not different between TNF-alpha gene-sufficient and TNF-alpha gene-deficient mice. In mouse cancer cells, TNF-alpha functioned via nuclear factor-kappaB- and neutral sphingomyelinase-dependent pathways to induce TNF-alpha and VEGF, respectively. These results were recapitulated in human cancer cells, and a correlation was detected between TNF-alpha and VEGF content of human MPE. We conclude that tumor-derived TNF-alpha is important in the development of MPE in mice, and provide preclinical evidence supporting the efficacy of TNF-alpha blockade against malignant pleural disease.

Autocrine IL-6-induced Stat3 activation contributes to the pathogenesis of lung adenocarcinoma and malignant pleural effusion

Malignant pleural effusion (MPE) is a poor prognostic sign for patients with non-small-cell lung cancer (NSCLC). The generation of MPE is largely regulated by vascular endothelial growth factor (VEGF), and upregulation of VEGF by Stat3 has been observed in several types of tumor cells. In this study, we demonstrate constitutively activated Stat3 in several human lung cancer cell lines and in tumor cells infiltrated in the pleurae of patients with adenocarcinoma cell lung cancer (ADCLC) and MPE. The observations suggest that activated Stat3 plays a role in the pathogenesis of ADCLC. In PC14PE6/AS2 cells, a Stat3-positive human ADCLC cell line, autocrine IL-6 activated Stat3 via JAKs, not via Src kinase. PC14PE6/AS2 cells express higher VEGF mRNA and protein than do Stat3-negative PC14PE6/AS2/dnStat3 cells. In an animal model, PC14P6/AS2/dnStat3 cells produced no MPE and less lung metastasis than did PC14P6/AS2 cells. PC14PE6/AS2 cells also expressed higher VEGF protein, microvessel density, and vascular permeability in tumors than did PC14P6/AS2/dnStat3 cells. Therefore, we hypothesize that autocrine IL-6 activation of Stat3 in ADCLC may be involved in the formation of malignant pleural effusion by upregulating VEGF. Higher levels of IL-6 and VEGF were also found in the pleural fluids of patients with ADCLC than in patients with congestive heart failure. The autocrine IL-6/Stat3/VEGF signaling pathway may also be activated in patients with ADCLC and MPE. These findings provide novel targets for the management of MPE.

P-131 Inhibition of lung metastases and formation of malignant pleural effusion of U14 uterine cervical cancer by 20(S)-ginsenoside Rh2 in mice

P-131 Inhibition of lung metastases and formation of malignant pleural effusion of U14 uterine cervical cancer by 20(S)-ginsenoside Rh2 in mice