5743

PTGS2

prostaglandin-endoperoxide synthase 2 (prostaglandin G/H synthase and cyclooxygenase);PTGS2;prostaglandin-endoperoxide synthase 2 (prostaglandin G/H synthase and cyclooxygenase)

PGH synthase 2|PHS II|cyclooxygenase 2b|prostaglandin G/H synthase 2|prostaglandin H2 synthase 2

1q25.2-q25.3

protein-coding

BACKGROUND/AIMS: c-met, c-erbB-2, interleukin (IL)-6, and cyclooxygenase (COX)-2 expressions are considered to be implicated in the carcinogenesis and progression of cholangiocarcinoma, but the molecular pathogenesis of cholangiocarcinoma is still poorly understood. We aimed to analyze the expressions of each marker and their relationships with clinicopathologic factors. METHODS: One hundred and fourteen tissue samples were obtained from surgically resected specimens from patients with biliary tract cancer. The expressions of c-met, c-erbB-2, COX-2, and IL-6 were examined by immunohistochemically. The expression of each marker and correlations between these markers and clinicopathologic factors were analyzed. RESULTS: The expression rates of each maker were as follows: c-met 34/112 (30.4%), c-erbB-2 5/112 (4.5%), COX-2 53/113 (46.9%), and IL-6 68/113 (60.2%), respectively. c-met expression was more frequently observed in cases with invasion through the adjacent connective tissues (p=0.0263). IL-6 overexpression was more frequently observed in cases with absent lymph node metastasis (p=0.0325). Either c-erbB-2 expression or COX-2 expression was significantly associated with lymph node metastasis (p=0.0442). CONCLUSIONS: The expression of c-met was closely related to the invasiveness of cholangiocarcinoma. Co-expression of c-met, COX-2 and, IL-6 showed a significant correlation with invasiveness and lymph node metastasis and these could be useful marker to guide clinical outcome in patients with cholangiocarcinoma.

OBJECTIVE: To study the expression of cyclooxygenase-2 (COX-2) gene in breast cancer and its clinicopathologic characteristics. METHODS: With beta-actin gene as reference, the COX-2 mRNA was monitored in 30 specimens of breast cancer tissue and adjacent normal breast tissue by reverse transcription-polymerase chain reaction (RT-PCR). RESULTS: The COX-2 mRNA expression was significantly upregulated in most breast cancer tissues with range of 0.05 - 0.91 (median 0.56), which was rare in normal breast tissue with range of 0 - 0.09 (median 0). The difference of COX-2 mRNA expression between cancer and normal breast tissue was significant (rank sum test, P < 0.05). COX-2 overexpression in breast cancer was related to its lymph node metastasis (P < 0.05) but not to age, tumor size, pathologic grade or pathologic type (P > 0.05). CONCLUSION: The level of COX-2 mRNA expression is obviously higher in the breast cancer tissue than that in normal breast tissue. COX-2 overexpression may play a crucial role in the carcinogenesis, development of cancer and lymph node metastasis in breast cancer patients.

Cyclooxygenase-2 (COX-2), an inducible enzyme involved in prostaglandin (including PGE(2)) biosynthesis, is overexpressed in several epithelial malignancies including breast cancer. We tested the hypothesis that COX-2 overexpression in breast cancer cells results in increased cell motility and invasion. COX-2 overproducing cells were generated by stable transfection of several human breast cancer cells with pSG5-COX2 vector. We confirmed the overexpression of COX-2 protein by western blotting, and by measuring PGE(2) in the medium with an immunoassay. We measured cell motility by counting the number of cells crossing an 8-micron pore size PET membrane, and cell invasion by counting the number of cells invading through a Matrigel-coated membrane that simulates basement membrane. COX-2 transfected MDA-231 cells produced 30-43-fold more PGE2 as compared to parental cells. COX-2 overexpression increased cell migration approximately 2.2-fold and cell invasion through Matrigel approximately 5.1-fold. Addition of 50 microM NS-398, a COX-2 inhibitor, inhibited Matrigel invasion of MDA-231 cells by 54% as compared to solvent confirming the role of COX-2 in cell invasion. It is known that an increase in cell migration and invasion can be brought about by cytoskeletal alterations and basement membrane degradation due to increased expression of pro-urokinase plasminogen activator (pro-uPA). To investigate the mechanism of our observed increase in cell invasion by COX-2, we found by western blotting that the level of pro-uPA was significantly higher (approximately 5-fold) in COX-2 transfected MDA-231 cells than untransfected MDA-231 cells. Similar to our observations in cell culture, we found evidence that increased COX-2 activity correlates with uPA in a mouse model of breast cancer metastasis to bone. In this study, we conclude that COX-2 overexpression in human breast cancer cells enhances cell motility and invasiveness thus suggesting a mechanism of COX-2 mediated metastasis.

metastasis entails numerous biological functions that collectively enable cancerous cells from a primary site to disseminate and overtake distant organs. Using genetic and pharmacological approaches, we show that the epidermal growth factor receptor ligand epiregulin, the cyclooxygenase COX2, and the matrix metalloproteinases 1 and 2, when expressed in human breast cancer cells, collectively facilitate the assembly of new tumour blood vessels, the release of tumour cells into the circulation, and the breaching of lung capillaries by circulating tumour cells to seed pulmonary metastasis. These findings reveal how aggressive primary tumorigenic functions can be mechanistically coupled to greater lung metastatic potential, and how such biological activities may be therapeutically targeted with specific drug combinations.

Cyclooxygenases (COX) are rate-limiting enzymes involved in the conversion of PLA(2)-mobilized arachidonic acid into prostaglandins and thromboxanes. COX-2 is a key mediator of inflammation during both physiologic and pathologic responses to endogenous stimuli and infectious agents. Its overexpression has been detected in different cancers, including that of the breast. Using RNA interference, we have reduced the expression of COX-2 in the highly malignant breast cancer cell line MDA-MB-231 below detectable levels in response to interleukin-1 beta or 12-O-tetradecanoylphorbol-13-acetate treatment. Microarray analysis showed that COX-2 silencing resulted in the loss of mRNA expression of several oncogenic markers, such as matrix metalloproteinase-1, chemokine (C-X-C motif) receptor 4, and interleukin-11, which have been correlated with poor disease outcome, and in the up-regulation of antimetastatic transcripts, such as thrombospondin-1 and Epstein-Barr-Induced 3. Cells lacking COX-2 were less able to invade reconstituted extracellular matrix than parental cells in vitro. Consistent with these changes, loss of COX-2 resulted in the abolition or the significant delay of tumor onset when the cells were injected in the mammary fat pad of severe combined immunodeficient mice. Finally, silencing of COX-2 resulted in the inhibition of metastasis to the lungs of severe combined immunodeficient mice after intravenous injection. These data show that silencing of COX-2 abolishes the metastatic potential of MDA-MB-231 cells in vivo.

BACKGROUND: Lymphangiogenesis has become a new research frontier in tumor metastasis since the discovery of reliable lymphatic markers that have allowed observation and isolation of lymphatic endothelium. Cyclooxygenase-2 (COX-2) has been reported to be involved in the critical steps in carcinogenesis. However, possible role of COX-2 in lymphangiogenesis and lymphatic metastasis is still poorly understood. In present study, we aimed to investigate the relationship between vascular endothelial growth factor-C (VEGF-C) and COX-2 in human breast cancer, and correlations with lymphangiogenesis and prognosis. METHODS: Tissue samples of primary tumors from 70 patients undergoing intentionally curative surgical resections for breast cancer were immunohistochemically examined for VEGF-C, COX-2, and D2-40 expressions. The association between COX-2 and VEGF-C expressions and clinicopathological parameters as well as prognosis were analysised. To demonstrate the presence of proliferating lymphatic endothelial cells, 10 random cases with high LVD counts were selected for D2-40/Ki-67 double immunostaining. RESULTS: A significant correlation was found between the expression of VEGF-C and COX-2 (r = 0.529, P < 0.001), and both elevated VEGF-C expression and elevated COX-2 expression were associated with higher lymph vessel density (LVD), lymph node metastasis and D2-40 positive lymphatic invasion (LVI) as well as worse disease free survival (DFS) and overall survival (OS) in a univariate analysis. In the double immunostain for the lymph vessel marker D2-40 and the proliferation marker Ki-67, the results confirmed Ki-67-positive nuclei in a proportion of lymph vessel endothelial cells. CONCLUSION: There is indeed lymphangiogenesis in breast cancer, the most compelling evidence being the presence of proliferating lymphatic endothelial cells. VEGF-C and COX-2 are coexpressed and significantly associated with lymphangiogenesis and prognosis in invasive breast cancer. Suggesting COX-2 may up-regulate VEGF-C expression and thus promote lymph node metastasis via lymphangiogenesis pathway in human breast cancer.

PURPOSE: To evaluate the prognostic significance of cyclooxygenase-2 (COX-2) in breast cancer patients treated with conservative surgery and radiation therapy (CS+RT). METHODS AND MATERIALS: Between 1975 and 2003, we retrieved specimens from 504 breast cancer patients treated with CS+RT. The specimens were constructed into tissue microarrays processed and stained for estrogen receptor (ER), progesterone receptor, Her2/neu, and COX-2. Each core was scored as positive or negative. All data including demographics, clinical, pathologic, staging, and outcome variables were entered into a computerized database. RESULTS: expression of COX-2 was positive in 58% of cases and correlated with younger age (p = 0.01) and larger tumor size (p = 0.001). expression of COX-2 was predictive of local relapse (relative risk[RR], 3.248; 95% confidence interval [CI], 1.340-7.871; p = 0.0091), distant metastasis (RR, 2.21; 95% CI, 1.259-3.896; p = 0.0058), and decreased survival (RR, 2.321; 95% CI, 1.324-4.071; p = 0.0033). Among ER-positive patients, COX-2 expression was predictive of worse local control (85% vs. 93%, p = 0.04), distant metastasis (75% vs. 95%, p = 0.002) and worse survival (65% vs. 94%, p = 0.002). Among ER-negative tumors COX-2 expression was not significantly correlated with local control (87 vs. 95%, p = 0.12), distant metastasis (73% vs. 78%, p = 0.39), or survival (77% vs. 87%, p = 0.15). CONCLUSIONS: In breast cancer patients treated with CS+RT, COX-2 expression is associated with younger age, larger tumor size, worse local control, distant metastasis, and worse overall survival. The significance is limited to hormone receptor-positive tumors, consistent with the known effect of COX-2/PGE2 on aromatase activity. Use of COX-2 inhibitors in estrogen-dependent breast cancers warrants further investigation.

OBJECTIVE: COX-2 is implicated in carcinogenesis and tumour progression in many cancers, including breast cancer. Recently, it has been reported that human breast carcinomas aberrantly express COX-2, and that raised tissue levels of COX-2 may have prognostic value. Patients expressing high levels of COX-2 can develop local recurrence, and have reduced disease-free and disease-related overall survival. The aim of this study was to investigate COX-2 expression in human ductal and lobular breast cancers and its possible association with clinicopathological features and prognostic molecular markers. RESEARCH DESIGN AND METHODS: Cytoplasmic COX-2 expression was detected by means of immunohistochemistry in a series of 91 breast carcinomas with ductal (n = 60) and lobular (n = 31) patterns. COX-2 expression was investigated by multivariate analyses and compared with clinicopathological features. RESULTS AND CONCLUSIONS: COX-2 immune positivity and percentage of positive cells correlated significantly with the size, grading, extent of primary tumour and vascular invasion of carcinoma but not with biological parameters (estrogen receptor, progesterone receptor and human EGF receptor 2). The findings of the present study suggest that COX-2 overexpression in lobular and ductal breast cancers, which correlates with traditional clinico-pathological parameters, may be considered as a negative prognostic marker.

The aim of this article is to identify nuclear co-localization of COX-2 and HIF-1alpha in human-bone metastasis of breast cancer, index of transcriptionally activated cells and functional for gene expression. In particular, we verified whether hypoxia exerted a direct role on metastasis-gene expression or through COX-2 signaling, due to the relevance for clinical implications to individuate molecular targets for diagnosis and therapy. The experiments were performed in vitro with two metastatic clones, 1833 and MDA-231BO, and the parental MDA-MB231 cells, in vivo (1833-xenograft model), and in human-bone metastasis specimens. In 1833 cells in vitro, COX-2 signaling pathway was critical for nuclear HIF-1alpha-protein expression/translocation, mechanisms determining HIF-1 activity and gene expression. The data were corroborated by immunohistochemistry in human-bone metastasis specimens. COX-2 and HIF-1alpha showed wide co-localization in the nucleus, indicative of COX-2-nuclear import in transcriptionally activated metastatic cells and consistent with COX-2-HIF-1alpha functional interaction. A network of microenvironmental signals controlled COX-2 induction and HIF-1 activation downstream. In fact, hypoxia through HGF and TGF-beta1 autoregulatory loops triggered a specific array of transcription factors responsible for COX-2 transactivation. The novelty was that HGF and TGF-beta1 biological signals were produced by hypoxic metastatic cells and, therefore, the microenvironment seemed to be modified by metastatic-cell engraftment in the bone. In agreement, HIF-1alpha expression in bone marrow supportive cells occurred in metastasis-bearing animals. Altogether, the data supported the pre-metastatic-niche theory. Our observations might be useful to design therapies against bone metastasis, by affecting the phenotype changes of metastatic cells occurring at the secondary growth site through COX-2-HIF-1 interaction.

BACKGROUND: A major problem with the use of current chemotherapy regimens for several cancers, including breast cancer, is development of intrinsic or acquired drug resistance, which results in disease recurrence and metastasis. However, the mechanisms underlying this drug resistance are unknown. To study the molecular mechanisms underlying the invasive and metastatic activities of drug-resistant cancer cells, we generated a doxorubicin-resistant MCF-7 breast cancer cell line (MCF-7/DOX). METHODS: We used MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) assays, flow cytometry assays, DNA fragmentation assays, Western blot analysis, cell invasion assays, small interfering RNA (siRNA) transfection, reverse transcription-polymerase chain reaction, experimental lung metastasis models, and gelatin and fibrinogen/plasminogen zymography to study the molecular mechanism of metastatic activities in MCF-7/DOX cells. RESULTS: We found that MCF-7/DOX acquired invasive activities. In addition, Western blot analysis showed increased expression of epidermal growth factor receptor (EGFR) and Cox-2 in MCF-7/DOX cells. Inhibition of Cox-2, phosphoinositide 3-kinase (PI3K)/Akt, or mitogen-activated protein kinase (MAPK) pathways effectively inhibited the invasive activities of MCF-7/DOX cells. Gelatin and fibrinogen/plasminogen zymography analysis showed that the enzymatic activities of matrix metalloproteinase-2 (MMP-2), MMP-9, and urokinase-type plasminogen activator were markedly higher in MCF-7/DOX cells than in the MCF-7 cells. In vitro invasion assays and mouse models of lung metastasis demonstrated that MCF-7/DOX cells acquired invasive abilities. Using siRNAs and agonists specific for prostaglandin E (EP) receptors, we found that EP1 and EP3 played important roles in the invasiveness of MCF-7/DOX cells. CONCLUSIONS: We found that the invasive activity of MCF-7/DOX cells is mediated by Cox-2, which is induced by the EGFR-activated PI3K/Akt and MAPK pathways. In addition, EP1 and EP3 are important in the Cox-2-induced invasion of MCF-7/DOX cells. Therefore, not only Cox-2 but also EP1 and EP3 could be important targets for chemosensitization and inhibition of metastasis in breast cancers that are resistant to chemotherapy.

BACKGROUND: In breast cancer cyclooxygenase-2 overexpression is related with poor outcomes, but the clinical relevance of cyclooxygenase-2 is still unclear. Progesterone receptor is a regulator of cyclooxygenase-2 expression. We postulate that progesterone receptor was associated to cyclooxygenase-2 expression and clinicopathologic factors in breast cancer. METHODS: Cyclooxygenase-2 and progesterone receptor expression was analyzed, by immunohistochemistry, in 31 cases of invasive ductal carcinoma. RESULTS: The expression of cyclooxygenase-2 and progesterone receptor was observed in 64.5% and 45.2% of the tumors, respectively. We found 11 tumors with both cyclooxygenase-2 and progesterone receptor overexpression and they are related with tumor lower size and lower axillary node metastasis. CONCLUSION: These results suggest that progesterone receptor has a protective role in breast cancer by inhibiting chronic inflammatory pathway.

Up-regulation of cyclooxygenase-2 (COX-2) is frequently found in human cancers and is significantly associated with tumor metastasis. Our previous results demonstrate that COX-2 and its metabolite prostaglandin E2 (PGE2) stimulate the expression of CCR7 chemokine receptor via EP2/EP4 receptors to promote lymphatic invasion in breast cancer cells. In this study, we address the underlying mechanism of COX-2/PGE2-induced CCR7 expression. We find that COX-2/PGE2 increase CCR7 expression via the AKT signaling pathway in breast cancer cells. Promoter deletion and mutation assays identify the Sp1 site located at the -60/-57 region of CCR7 gene promoter is critical for stimulation. Chromatin immunoprecipitation (ChIP) assay confirms that in vivo binding of Sp1 to human CCR7 promoter is increased by COX-2 and PGE2. Knockdown of Sp1 by shRNA reduces the induction of CCR7 by PGE2. We demonstrate for the first time that AKT may directly phosphorylate Sp1 at S42, T679, and S698. Phosphorylation-mimic Sp1 protein harboring S42D, T679D, and S698D mutation strongly activates CCR7 expression. In contrast, change of these three residues to alanine completely blocks the induction of CCR7 by PGE2. Pathological investigation demonstrates that CCR7 expression is strongly associated with phospho-AKT and Sp1 in 120 breast cancer tissues. Collectively, our results demonstrate that COX-2 up-regulates CCR7 expression via AKT-mediated phosphorylation and activation of Sp1 and this pathway is highly activated in metastatic breast cancer.

BACKGROUND/AIM: Heparanase (HPA) contributes to breast cancer metastasis by facilitating the breakdown of the basement membrane and extracellular matrix. High expression of HPA is thought to be associated with increased nodal involvement and poor survival in patients with breast cancer. Overexpression of cyclooxygenase-2 (COX-2) in breast cancer is associated with indicators of poor prognosis such as lymph node metastasis, poor differentiation, and large tumor size. The underlying mechanism by which HPA and COX-2 overexpression increases the metastatic potential of breast cancer is not fully-understood. To enhance our understanding over these mechanisms, we aimed to investigate the relationship between the size of the tumor and HPA expression, tumor grade as well as lymph node status in patients with breast cancer. MATERIALS AND METHODS: Immunohistochemical analysis of HPA and COX-2 expression was performed on 246 breast tumor samples. The expression of HPA was correlated with COX-2 expression, tumor grade, lymph node status, oestrogen receptor status. RESULTS: The overexpression of HPA and COX-2 was associated with increased likelihood of lymph node positivity in large, high-grade tumors. High-grade tumors with size greater than 20 mm, that overexpressed HPA, were 4-times more likely to be associated with lymph node involvement (OR 4.71,

OBJECTIVE: To investigate the expressions of COX-2 and MMP-9 in cervical carcinoma and their clinical significance. METHODS: Immunohistochemical SP method was used to detect the expressions of COX-2 and MMP-9 in 72 cases of invasive carcinoma of cervix (ICC) and 16 cases of normal cervical epithelium remote from tumor (NCE), and the relationship between the expressions of COX-2 and MMP-9 in ICC and some factors relating to clinical pathology of cervical carcinoma such as histopathological grading, lymph node metastasis, stroma involvement and FIGO staging were analyzed statistically. RESULTS: The rates of positive expression of COX-2 and MMP-9 in ICC were significantly higher than those in NCE. COX-2: 88.9% in group ICC and 12.5% in group NCE, P = 0.000. MMP-9: 94.4% in group ICC and 43.8% in group NCE, P = 0.000. The expression of COX-2 was positively correlated with lymph node metastasis and stroma involvement (r = 0.296, P = 0.012 in group ICC and r = 0.257, P = 0.029 in group NCE, respectively). The expression of MMP-9 was positively correlated with FIGO staging (r = 0.329, P = 0.005) and histopathological grading (r = 0.351, P = 0.003). The expression of COX-2 was positively correlated with the expression of MMP-9 in ICC (r = 0.297, P = 0.011). CONCLUSION: The overexpressions of COX-2 and MMP-9 are closely related to the invasion and growth of cervical carcinoma. The tissue with overexpression of COX-2 has strong invasion ability. COX-2 and MMP-9 have synergistic effect on proliferation, invasion and metastasis of cancer cells. Detecting the expression of both COX-2 and MMP-9 may be of value in further understanding the biological behavior and predicting the prognosis of cervical carcinoma.

COX-2 overexpression is recognized in various cancers, but the role of COX-2 in the progression of cancer, including the liver metastasis of colon cancer, is not clearly understood. We examined the role of COX-2 in the mechanism of liver metastasis of colon cancer, using a highly metastasizable colon carcinoma cell line, LM-H3. A COX-2 inhibitor, JTE-522, inhibited cell proliferation and invasion of LM-H3 in vitro and clearly reduced the number of metastatic nodules on the surface of nude mouse livers in vivo. We also examined the effects of JTE-522 on the production of growth factors and MMPs through the use of ELISA and gelatin zymography, respectively. JTE-522 downregulated PDGF production by LM-H3 but had no influence on VEGF production. JTE-522 also inhibited MMP-2 secretion by LM-H3. JTE-522 downregulated PGE(2) production, but the associated changes in PGE(2) did not affect PDGF and VEGF production by LM-H3. We conclude that JTE-522 downregulated the cell proliferation and invasive potential of LM-H3 by reducing the production of PDGF and MMP-2 and hypothesize that these inhibitory effects on the production of PDGF and MMP-2 can lead to inhibition of liver metastasis of colon cancer. These data indicate that the COX-2 inhibitor JTE-522 has a high potential for use as a clinical agent for the treatment of liver metastasis of colon cancer.

The aim of the present study is to investigate the relation of cyclooxygenase (COX)-2 with liver metastasis. Pieces of human colon cancer xenograft were implanted orthotopically (CI), as well as intraperitoneally, and subcutaneously. Liver metastasis developed most frequently, and the COX-2 expression of both mRNA and protein of the tumors was the most dominant in the CI group. In contrast, the expression of COX-1 or vascular endothelial cell growth factor mRNAs had no significant differences among the groups. The intensity of COX-2 mRNA was negatively correlated with the apoptotic index. In conclusion, COX-2 plays an important role in organ-specific metastasis of the colon cancer.

PURPOSE: Several lines of experimental evidence indicated that over-expression of vascular endothelial growth factor-C and cyclooxygenase-2 genes promotes angiogenesis and lymphangiogenesis, both of which are essential for the growth and spreading of tumor cells. This study was designed to evaluate the coexpression of vascular endothelial growth factor-C and cyclooxygenase-2 in human colorectal carcinoma to determine their relationships and correlations with lymph node metastasis and prognosis. METHODS: Tissue samples of primary tumors and metastatic lymph nodes from 150 patients undergoing intentionally curative surgical resections for colorectal adenocarcinoma were immunohistochemically examined for vascular endothelial growth factor-C, cyclooxygenase-2, and CD34 expressions. Then, we analyzed their relationships and correlations with clinicopathologic findings and patients' survival time. RESULTS: The positivity rate of vascular endothelial growth factor-C and cyclooxygenase-2 in the primary tumor was 68 and 72.7 percent, respectively, and in the metastatic lymph nodes was 93.3 and 80 percent, respectively. A significant correlation was found between the expression scores of vascular endothelial growth factor-C and cyclooxygenase-2 (P < 0.0001), and both also were correlated to microvessels density and several clinicopathologic parameters, including primary tumor size, lymph node metastasis, lymphatic invasion, and TNM stage. Patients with vascular endothelial growth factor-C-positive and/or cyclooxygenase-2-positive tumors had a significant shorter survival time than those with negative tumors did. However, in a multivariate analysis, only cyclooxygenase-2 expression was recognized as an independent prognostic factor (P = 0.0412; relative risk ratio, 3.067; 95 percent confidence interval, 1.046-8.994). CONCLUSIONS: These data show that in human colorectal carcinoma, vascular endothelial growth factor-C and cyclooxygenase-2 are coexpressed and significantly associated with lymph node metastasis and prognosis.

PURPOSE: Recently, an objective response rate of 12% was reported in a phase II study of cetuximab in patients with epidermal growth factor receptor (EGFR)-expressing metastatic colorectal cancer (mCRC) refractory to fluoropyrimidine-, oxaliplatin-, and irinotecan-based chemotherapy (IMC-0144). In this large molecular correlates study, we tested whether K-ras mutation status and polymorphisms in genes involved in the EGFR-signaling pathway were associated with clinical outcome in IMC-0144. EXPERIMENTAL DESIGN: We analyzed all available tissue samples from 130 of 346 mCRC patients enrolled in the IMC-0144 phase II clinical trial of cetuximab. Genomic DNA was extracted from formalin-fixed paraffin-embedded tumor tissues, and K-ras mutation status and the genotypes were analyzed using PCR-RFLP, direct DNA-sequencing, and 5-end [gamma-33P] ATP-labeled PCR-protocols. RESULTS: The PFS of patients with cyclooxygenase-2 (COX-2) -765 G>C [C/C; risk ratio (RR), 0.31; 95% confidence interval (95% CI), 0.12-0.84; P = 0.032], COX-2 +8473 T>C (C/C; RR, 0.67; 95% CI, 0.40-1.13; P = 0.003), EGF +61 A>G (G/G; RR, 0.57; 95% CI, 0.34-0.95; P = 0.042), and EGFR +497 G>A (A/G; RR, 0.82; 95% CI, 0.56-1.20; P = 0.017) genotypes was significantly longer compared with those with other genotypes. In addition, patients whose tumors did not have K-ras mutations showed better RR, PFS, and overall survival than patients with K-ras mutations. In multivariable analysis, COX-2 +8473 T>C (adjusted P = 0.013) and EGFR +497 G>A (adjusted P = 0.010) remained significantly associated with progression-free survival, independent of skin rash toxicity, K-ras mutation status, and Eastern Cooperative Group performance status. CONCLUSIONS: Polymorphisms in COX-2 and EGFR may be useful independent molecular markers to predict clinical outcome in patients with mCRC treated with single-agent cetuximab, independent of skin rash toxicity, K-ras mutation, and Eastern Cooperative Oncology Group performance status.

Several new drugs that are targeted towards various angiogenic factors have shown considerable potential for controlling tumor proliferation and metastases. expression levels of the targeted genes in primary tumors and metastases should be understood to maximize the use of such drugs. The present study aimed to clarify associations between mRNA levels of cyclooxygenase 2 (COX-2) and angiogenic factors [vascular endothelial growth factor (VEGF) and interleukin-8 (IL-8)] in primary colorectal cancer and in corresponding liver metastasis. We also compared these gene expressions of primary colorectal cancer between patients with and without liver metastasis. In 31 pairs of formalin-fixed and paraffin-embedded primary and metastatic liver tumors as well as 27 specimens of consecutive stage II patients without recurrence, mRNA was quantified by real-time reverse transcription-polymerase chain reaction following the laser capture microdissection. We found a significantly positive correlation in IL-8 between primary tumors and matched liver metastases (p=0.034, rs=0.39) and in VEGF (p=0.0083, rs=0.48), but not in COX-2, which was associated with both VEGF (p=0.044, rs=0.37) and IL-8 (p=0.0004, rs=0.64) in primary colorectal cancers. Multiple regression analysis revealed that COX-2 was independently associated with IL-8 (p<0.0001). There were no differences in mRNA levels between patients with and without liver metastasis. The mRNA levels of VEGF and IL-8 in liver metastasis can be predicted from those in primary colorectal cancer. COX-2 might exert angiogenic activity more through the IL-8, than the VEGF pathway. These angiogenic factors were sufficiently up-regulated before hematogenous metastasis. These preliminary data merit further validation studies.

BACKGROUND: The role of peroxisome proliferator-activated receptor delta (PPAR delta) in the development and progression of colorectal cancer (CRC) remains controversial. AIMS: We investigated the impact of PPAR delta expression in tissues on liver metastasis of CRC. METHODS: We analyzed samples of primary CRC and matched normal adjacent tissues from 52 patients for the expression of PPAR delta, cyclooxygenase (COX)-2, vascular endothelial growth factor (VEGF)-A, and CXC chemokine receptor 4 (CXCR4). Correlations of the molecules expressions with clinical characteristics and prognosis of patients were studied. RESULTS: The number of patients positive for PPAR delta, COX-2, CXCR4, and VEGF-A was 25, 33, 18, and 19, respectively. Among the PPAR delta (+)/COX-2 (+), PPAR delta (-)/COX-2 (+), PPAR delta (+)/COX-2 (-), and PPAR delta (-)/COX-2 (-) patient groups, PPAR delta (+)/COX-2 (+) patients had the highest incidence of liver metastasis (p<0.01). PPAR delta (+)/COX-2 (+) expression was a significant independent prognostic factor (HR=7.108, 95% CI 1.231-41.029, p=0.0283) by Cox proportional analysis. PPAR delta (+)/COX-2 (+) patients had the highest positivity for CXCR4 or VEGF-A in tissues (p<0.01). Among the patients in the CXCR4 (+)/VEGF-A (+), CXCR4 (+)/VEGF-A (-), CXCR4 (-)/VEGF-A (+), and CXCR4 (-)/VEGF-A (-) groups, CXCR4 (+)/VEGF-A (+) patients had the highest incidence of liver metastasis (p<0.01). CONCLUSIONS: The expression of both PPAR delta and COX-2 in tissues may lead to liver metastasis and consequent poor prognosis in CRC patients.

Increased prostaglandins (PGs) are associated with many inflammatory pathophysiological conditions; and are synthesized from arachidonic acid by either of 2 enzymes, cyclooxygenase-1 (COX-1) or -2 (COX-2). Recent epidemiologic, expression, and pharmacologic studies suggest COX-2 derived metabolites also play a functional role in the maintenance of tumor viability, growth and metastasis. Archival and/or prospectively collected human tissues were prepared for immunohistochemistry, and representative cases assayed via Western blot, RT-PCR, or TAQman analysis. Consistent overexpression of COX-2 was observed in a broad range of premalignant, malignant, and metastatic human epithelial cancers. COX-2 was detected in ca. 85% of the hyperproliferating, dysplastic, and neoplastic epithelial cells, and in the existing and angiogenic vasculature within and adjacent to hyperplastic/neoplastic lesions. These data collectively imply COX-2 may play an important role during premalignant hyperproliferation, as well as the later stages of invasive carcinoma and metastasis in various human epithelial cancers.

BACKGROUND: Vascular endothelial growth factor-C (VEGF-C) is a specific growth factor of lymphatics, which is known to play some role in tumor growth and metastasis to lymph nodes and distant organs in various malignancies. The purpose of the present study was to investigate the expression of VEGF-C in human esophageal squamous cell carcinomas (ESCC) to elucidate its role in tumor progression and lymph node metastasis. Another aim of the study was to investigate the relation between VEGF-C and cyclooxygenase-2 (COX-2) in ESCC. METHODS: The expression of VEGF-C and COX-2 in ESCC was evaluated in 13 endoscopic mucosal resection specimens and in 21 surgical specimens by immunohistochemical staining. Clinical data were obtained from medical records. RESULTS: The degree of VEGF-C expression increased as the depth of primary tumor progressed (r = 0.521, P = 0.002), the stage progressed (r = 0.572, P < 0.001), and the degree of COX-2 expression increased (r = 0.387, P = 0.024). The VEGF-C positive rate was different between early cancers in which regional lymph node metastasis was thought to be absent and advanced cancers in which regional lymph node metastases were confirmed after surgery (20.0% vs 100.0%; P < 0.001). CONCLUSIONS: The VEGF-C expression in ESCC is related to COX-2 expression, and VEGF-C is also associated with the depth of primary tumor, the stage, and probably lymph node metastasis. Thus the investigation of VEGF-C expression in ESCC may assist in management planning.

BACKGROUND & OBJECTIVE: Molecular biological studies indicated that abnormal expression of cyclooxygenase-2(COX-2), p53, proliferating cell nuclear antigen(PCNA), and nm23 correlate with the development of gastric cancer, but the relationship between abnormal expression of above gene proteins and the biological behavior of gastric cancer was not yet clear. This study was designed to investigate the relationship between expressions of COX-2, p53, PCNA, and nm23 and the clinicopathological behavior in gastric cancer. METHODS: The expressions of COX-2, p53, PCNA, and nm23 were detected by using immunohistochemical(SP) method. RESULTS: (1) The overexpression rate of COX-2 in gastric cancer was 70.4% (50/71), and the overexpression of COX-2 was correlated to tumor size, gross morphology, histological type, lymph node metastases, and clinicopathological stage of gastric cancer (P < 0.05, or P < 0.01). (2) The overexpression rates of p53 and PCNA in 71 surgically removed specimens from the patients with gastric cancer were 74.6% and 78.9%, respectively. And the expressions of p53 (86.7%) and PCNA (88.8%) in gastric cancer patients with lymph node metastasis were significantly higher than those in patients without lymph node metastasis (53.8% and 61.5%) (both, P < 0.05), respectively. Moreover, expressions of p53(52.2%) and PCNA(60.9%) in stage I + II patients were lower than those(85.4% and 87.5%) in stage III + IV (both, P < 0.05). (3) The low-expression of nm23 in gastric cancer was 71.8%, and its low-expression rate of nm23 in patients without lymph node metastasis(88.5%) and in stage I + II (91.3%) were much higher than those in the patients with lymph node metastasis (62.2%) and in stage III + IV (62.5%) (respectively, P < 0.05). (4) The differences of histological types, depth of invasion, lymph node metastasis and clinical stage were significantly different(P < 0.05 or P < 0.001), between the patients with two or more genes expression of COX-2, p53, PCNA, and nm23 and those with less one gene expression. CONCLUSION: Our results suggest that abnormal expressions of COX-2, p53, PCNA, and nm23 associate with malignant potential, lymph node metastasis and clinical stage, and they might therefore play a role in development of gastric cancer.

Epidemiological studies have shown that the use of nonsteroid anti-inflammatory drugs (NSAIDs) is associated with reduced risk of gastric cancer. The best-known target of NSAIDs is the cyclooxygenase (Cox) enzyme. Two Cox genes have been cloned, of which Cox-2 has been connected with gastric carcinogenesis. expression of Cox-2 is elevated in gastric adenocarcinomas, which correlates with several clinicopathological parameters, including depth of invasion and lymph node metastasis. This suggests that Cox-2-derived prostanoids promote aggressive behavior of adenocarcinomas of the stomach. Cox-2 expression is especially prominent in intestinal-type gastric carcinoma and it is already present in dysplastic precursor lesions of this disease, which suggests that Cox-2 contributes to gastric carcinogenesis already at the preinvasive stage. Our most recent data show that Cox-2 is expressed in gastric adenomas of trefoil factor 1 deficient mice. Treatment of these mice with a Cox-2 selective inhibitor, celecoxib, reduced the size of the adenomas. Taken together these data support efforts to initiate clinical studies to investigate the effect of Cox-2 inhibitors as chemotherapeutic agents and as adjuvant treatment modalities against gastric neoplasias.

AIM: To investigate the expression of cyclooxygenase-2 (COX-2) in gastric cancer and its relation with the liver metastasis and prognosis. METHODS: expression of COX-2 mRNA and protein was examined in gastric cancer and its paired substantial normal tissue by semi-quantitative reverse transcription-polymerase chain reaction and immunohistochemistry. The relation between COX-2 expression and prognosis was investigated in 195 cases. RESULTS: The expression of COX-2 mRNA in gastric cancer tissue was significantly higher than that in normal tissue in 47 cases (w = 792, P<0.01). The COX-2 mRNA in pT3-4 tissue expressed higher than that in pT1-2 tissue (w = 204, P<0.05). The positive expression rate of COX-2 protein was 57.9% (113/195). The COX-2 expression was significantly related to histological type, lymphnode metastasis, venous invasion and liver metastasis (P<0.05). No relation was found between COX-2 expression and invasion depth, peritoneal metastasis and International Union against cancer TNM stage. The multiple regression analysis showed that the COX-2 expression and venous invasion were obviously associated with liver metastasis (P<0.05). However, there was no significant correlation between COX-2 immunoreactivity and prognosis. CONCLUSION: COX-2 may play an important role in the development of gastric cancer, and the over-expression of COX-2 protein may be a high risk factor for liver metastasis.

Osteopontin (OPN), cyclooxygenase-2 (COX-2) and vascular endothelial growth factor (VEGF) are overexpressed in various experimental models of malignancy. However, the correlation and role of the three molecules in gastric cancer is unclear. In the present study, we found that OPN, COX-2 and VEGF were overexpressed in 53 cancerous tissues with gastric cancer compared with 40 normal mucosa tissues by immunohistochemistry method. Moreover, the results indicated co-expression of OPN, COX-2, and VEGF in gastric cancer. Levels of OPN, COX-2, and VEGF were all significantly correlated with TNM stage, lymph node metastasis and distant metastasis (P < 0.05), while not related to prognosis of patients. In addition, individual levels of OPN, COX-2, and VEGF were all significantly correlated with microvessel density (MVD), valued by CD34 staining directly with r-values of 0.416, 0.400, and 0.566, respectively (P < 0.01). Both OPN and COX-2 levels showed a positive correlation with VEGF (P < 0.05). Meanwhile, expression of COX-2 is in relation to OPN (P < 0.01). Overall, survival for patients with high MVD was significantly lower than for patients with low MVD (P < 0.05). Our findings indicate that OPN, COX-2, and VEGF synergically promote angiogenesis and metastasis in gastric cancer. It may be an important and useful strategy to target these molecules for prevention and therapy of tumor.

Cycloxygenase-2 catalyzes the synthesis of prostaglandins from arachidonic acid and this enzyme has been implicated in the metastasis of gastric cancer. In order to examine the significance of cycloxygenase-2 (Cox-2) in the survival and proliferation of gastric cancer cells, we have stably overexpressed an antisense Cox-2 in two gastric cancer cell lines, SGC7901 and AGS, in order to reduce the expression of this protein. The sense and antisense Cox-2 expression vectors were created by cloning COX-2 cDNA, in pIRES2-EGFP plasmid. Cox-2 gene expression was monitored by RT-PCR and Western blotting and the results indicated that cells with antisense Cox-2 construct had significantly reduced Cox-2 expression in comparison to the cells that received sense-Cox-2 plasmid. Reduction of Cox-2 expression in SGC7901 and AGS gastric cancer cells led to markedly decreased proliferation. The metastatic capability of the two cell lines, as assessed by in vitro colony formation assay, is also significantly compromised by lowered Cox-2 expression. Thus, this study demonstrates that Cox-2 activity is necessary for the proliferation and metastasis of gastric cancer cells.

OBJECTIVE: To investigate the expression of cyclooxygenase-2 in laryngeal squamous cell carcinoma and the correlation with tumor angiogenesis. METHOD: The COX-2 and FVIII of 42 cases of laryngeal squamous cell carcinoma were detected by immunohistochemical SABC method,and the correlation between the expression rates, clinical significance and microvessel density(MVD) was analyzed. RESULT: The positive rate of COX-2 in laryngeal squamous cell carcinoma tissues and adjacent normal tissues were 71.43% and 19.05% respectively. The expression of COX-2 were not associated with age, sex, tumor site and T stage. The tumors of grade III approximately IV showed a higher COX-2 expression than tumors of grade I approximately II did and the expression of COX-2 in the cervical lymph node metastasis group was higher than that in the non-metastasis group. The expression of COX-2 was closely associated with microvessel density(MVD) of laryngeal squamous cell carcinoma. CONCLUSION: The expression of COX-2 may play a crucial role in the carcinogenesis and development of laryngeal squamous cell carcinoma and cervical lymph node metastasis in patients with laryngeal squamous cell carcinoma. COX-2 may be a important factor in tumor angiogenesis.

OBJECTIVE: To investigate the protein and mRNA expression of HIF-1alpha and COX-2 in laryngeal squamous carcinoma, and their correlation with clinicopathologic parameters. METHOD: The protein and mRNA expression of HIF-1alpha and COX-2 in fresh laryngeal squamous carcinoma tissues and adjacent normal tissues from 76 patients were examined by immunohistochemistry and reverse transcription polymerase chain reaction. RESULT: 1) The protein positive expression rate of HIF-1alpha and COX-2 in tumor samples were significantly higher than that in normal tissues (P < 0.05); The protein positive expression rate of HIF-1alpha were higher in moderately and poorly differentiated tumors than in well differentiated tumors (P < 0.05); While the protein positive expression rate of COX-2 were not associated with T stage (P > 0.05); 2) The mRNA positive expression rate of HIF-1alpha and COX-2 were 67.11% and 75.00% respectively, their expression were significantly higher in tumors than that in normal tissues (P < 0.05). The mRNA expression of COX-2 were not detected in normal tissues. The mRNA positive expression rate of HIF-1alpha were associated with T stage, but were not associated with differentiated clinicopathologic stage. 3) The protein positive expression rate of HIF-1alpha were associated with that of COX-2. CONCLUSION: HIF-1alpha and COX-2 could up regulate each other to promote the development and metastasis of laryngeal squamous carcinoma.

The mechanisms underlying the effects of COX-2 on tumor lymphangiogenesis remain largely undefined. Here, the human lung cancer cell lines A549, 95D, Anip973, and AGZY83-a with different metastatic capacities were investigated by immunostaining, western blotting, and real-time RT-PCR. We observed increased expressions of COX-2 and VEGF-C in the three highly metastatic cell lines compared with the less metastatic AGZY83-a cell line. The COX-2-specific inhibitor Celecoxib suppressed VEGF-C expression whereas the main COX-2 metabolite PGE(2) elevated VEGF-C expression in Anip973 and AGZY83-a cells in positive and negative experiments. To determine the functional link to COX-2 more specifically and elucidate the mechanistic pathway, we used a siRNA to knock down the high COX-2 expression in Anip973 cells and transfected a COX-2 cDNA to enhance the low COX-2 expression in AGZY83-a cells, and then treated the cells with EP1/EP4 agonists or antagonists, respectively. The results revealed that the EP1/EP4 agonists significantly increased VEGF-C production in the COX-2-knockdown Anip973 cells. In contrast, the EP1/EP4 antagonists diminished VEGF-C production in the COX-2-overexpressing AGZY83-a cells. Furthermore, animal models provided evidence that Celecoxib decreased VEGF-C expression, lymphangiogenesis, and lymph node metastases in Anip973 cells, whereas PGE(2) treatment increased the same factors in the parental AGZY83-a cells. A positive correlation between COX-2 and VEGF-C was also confirmed in vivo. The present data suggest that COX-2 regulates VEGF-C expression via the PGE(2) pathway, and that EP1/EP4 receptors are involved in PGE(2)-mediated VEGF-C production. Thus, COX-2 may represent a candidate gene for blocking tumor lymphangiogenesis and lymph node metastasis.

BACKGROUND AND OBJECTIVE: The incidence of lung adenocarcinoma increases rapidly, and hypoxia-inducible factor-1alpha (HIF-1alpha), cyclooxygenase-2 (COX-2), E-cadherin play an important role in the proliferation and differentiation of cancer cell. The aim of this study is to investigate the clinical significance of the expression of HIF-1alpha, COX-2, E-cadherin in patients with lung adenocarcinoma and the internal relationship among them. METHODS: The expression levels of HIF-1alpha, COX-2 and E-cadherin were determined in 10 cases of normal lung issue and in 45 cases of issue of lung adenocarcinoma by immunohistochemical method. RESULTS: The positive expression rate of HIF-1alpha and COX-2 was 60% (27/45) and 40% (18/45) respectively in 45 cases of lung adenocarcinoma, was null in 10 cases of normal lung issue. The positive expression rate of E-cadherin was 48.9% (22/45) in 45 cases of lung adenocarcinoma, was 100% in 10 cases of normal lung issue. The expression of HIF-1alpha was positively correlated with the size of the cancer (P < 0.05), but not with age, smoking, lymphatic metastasis, differentiated degree and surgical-pathologic staging (P > 0.05). The expression of COX-2 was positively correlated with the size of the cancer, lymphatic metastasis, surgical-pathologic and expression of HIF-1alpha (P < 0.05), but not with age, smoking and differentiated degree (P > 0.05). The expression of E-cadherin was positively correlated with differentiated degree and lymphatic metastasis (P < 0.05), but not with age, smoking, the size of the cancer, surgical-pathologic and expression of HIF-1alpha (P > 0.05). CONCLUSIONS: The overexpression of HIF-1alpha and COX-2 and the downregulation of E-cadherin is found in lung adenocarcinoma. The overexpression of HIF-1alpha may induce the overexpression of COX-2, and is non-correlated with expression of E-cadherin.

Matrix metalloproteinases (MMPs) have been implicated in a variety of pathophysiological conditions, of which MMP-7 is expressed by tumor cells of epithelial and mesenchymal origin. However, the function of MMP-7 in human lung adenocarcinoma (LAC) is unclear. In the present study the expression of MMP-7 in LAC was examined by immunohistochemical assay using a tissue microarray procedure. A loss-of-function experiment was performed to explore the effects and molecular mechanisms of lentiviral vector-mediated MMP-7 siRNA (siMMP-7) on cell proliferation and invasive potential in LAC A549 cells, measured by MTT and Transwell assays, respectively. It was found that, the expression of MMP-7 protein in LAC was significantly increased compared with that in adjacent non-cancerous tissues (ANCT) (76.0% vs 44.0%, P<0.001), and positively correlated with lymph node metastases of the tumor (P=0.014). Furthermore, targeted inhibition of cyclooxygenase-2 (COX-2) by siRNA downregulated the expression of MMP-7 and inhibited invasion of LAC cells, and knockdown of MMP-7 suppressed tumor proliferation and invasion in LAC cells. Taken together, our findings indicate that increased expression of MMP-7 is associated with lymph node metastasis and upregulated by COX-2, and promotes the tumorigenesis of LAC, suggesting that MMP-7 may be a potential therapeutic target for the treatment of cancer.

Autotaxin, a lysophospholipase D producing lysophosphatidic acid, augments invasive and metastatic potential of tumor cells. Current investigations have focused on understanding the molecular mechanisms by which autotaxin regulates the expression of a major mediator of tumor invasion and metastasis, urokinase-type plasminogen activator (uPA) in human A2058 melanoma cells. Autotaxin induced uPA expression in a dose-dependent manner that was inhibited by pharmacological inhibitors for Gi (pertussis toxin), phosphoinositide 3-kinase (PI3K, LY294002), Akt inhibitor (AktI), proteosome activity and IkappaB phosphorylation (pyrrolidine dithiocarbamate), and by a dominant negative mutant (DN) of Akt. Autotaxin phosphorylated Akt and induced the translocation of nuclear [corrected] factor-kappaB (NF-kappaB) to the nucleus that were inhibited by AktI or by overexpressing DN-Akt. Consistently, green fluorescence protein-tagged p65 of NF-kappaB accumulated in the nucleus by autotaxin that was abrogated when the cells were transfected with DN-Akt. Moreover, autotaxin increased the DNA binding ability of NF-kappaB and promoter activity of uPA. Collectively, these data strongly suggest autotaxin induces uPA expression via the Gi-PI3K-Akt-NF-kappaB signaling pathway that might be critical for autotaxin-induced tumor cell invasion and metastasis.

In a previous study, we demonstrated that elevation of COX-2 is significantly associated with lymph node metastasis in squamous cell carcinoma (SCC) of cervix. The main objective of this study is to characterize the relationship between elevation of COX-2 and its possible clinical role in adenocarcinoma (AC) of cervix. Using immunohistochemistry, we examined COX-2 expression levels in 84 patients with AC of uterine cervix [71 ACs, 13 adenosquamous carcinomas (ASCs)]. Elevation of COX-2 was correlated with clinicopathological variables and p53 expression, as detected by immunohistochemistry. Elevation of COX-2 was detected in 13.0% (11 of 84) of the tumors. Elevation of COX-2 was significantly correlated with histologic type (AC 8.5% vs. ASC 38.5%, P=0.011). Both tumor stage and lymph node metastasis were correlated with elevation of COX-2 with a borderline significance (P=0.062 and 0.068, respectively). Elevation of p53 was not associated with elevation of COX-2. The association between lymph node metastasis and elevation of COX-2 was stronger in cases of AC than in cases of ASC (28.4 vs. 4.3%, P=0.023). According to the results of univariate analysis, elevation of COX-2 was significantly associated with decreased overall survival (P=0.003, log-rank test). However, multivariate analyses revealed that only tumor stage was independently associated with overall survival, suggesting that elevation of COX-2 itself may not be an independent prognostic factor. The present study shows that elevation of COX-2 may contribute to lymph node metastasis in AC of uterine cervix. This suggests that the potential therapeutic role of COX-2 inhibitors should be validated, not only in SCC, but also in AC of uterine cervix.

metastasis entails numerous biological functions that collectively enable cancerous cells from a primary site to disseminate and overtake distant organs. Using genetic and pharmacological approaches, we show that the epidermal growth factor receptor ligand epiregulin, the cyclooxygenase COX2, and the matrix metalloproteinases 1 and 2, when expressed in human breast cancer cells, collectively facilitate the assembly of new tumour blood vessels, the release of tumour cells into the circulation, and the breaching of lung capillaries by circulating tumour cells to seed pulmonary metastasis. These findings reveal how aggressive primary tumorigenic functions can be mechanistically coupled to greater lung metastatic potential, and how such biological activities may be therapeutically targeted with specific drug combinations.

To understand the role of cyclooxygenase (COX)-2 in metastatic potential of oral cancer, COX-2 overexpressing KB/COX-2 cells were inoculated orthotopically into the masseter muscle or injected into the left cardiac ventricle of nude mice. KB/COX-2 showed about 4-fold increase of COX-2 protein expression as compared to KB/Neo which was a mock transfected control. In orthotopic inoculation, metastasis to the regional lymph nodes occurred in 2 out of 15 mice, and metastasis to the lung in 3 out of 15 mice. On the other hand, in intra-cardiac injection, hematogenous metastasis to the lung and bone occurred in 8 out of 10 mice in KB/COX-2, but no metastasis occurred except for only one metastasis to the femur bone out of 10 mice in KB/Neo. Treatment of KB/COX-2 with COX-2 small interfering RNA (siRNA) inhibited the colony formation but not cell growth in vitro, and suppressed tumorigenicity and hematogenous metastasis in nude mice. When expression of adhesion molecules such as E-cadherin, alpha-catenin, beta-catenin and CD44 was examined, there was no difference in alpha- and beta-catenin between the cells. However, expression of E-cadherin was detected in KB/Neo, but not in KB/COX-2. In contrast, expression of CD44 was markedly increased in KB/COX-2 as compared to KB/Neo. Treatment with COX-2 siRNA resulted in suppression of CD44 expression and detectable expression of E-cadherin in KB/COX-2. These findings suggested that overexpression of COX-2 increased hematogenous metastasis, at least in KB cells, via down-regulating E-cadherin and up-regulating CD44 expression.

OBJECTIVE: To study the expression of cyclooxygenase-2 (COX-2) and matrix metalloproteinase-2 (MMP-2) in nasopharyngeal carcinoma (NPC) and their relationship with lymph node metastasis. METHOD: The expression of COX-2 and MMP-2 in 86 NPC tissues and 30 normal nasopharyngeal tissues were evaluated by immunohistochemical analysis. RESULT: The positive rate of COX-2 and MMP-2 in NPC tissue was 75.58% and 66.28% respectively, which was significantly higher than that in normal nasopharyngeal (P < 0.01). The expression of both COX-2 and MMP-2 were positively correlated with lymph node metastasis (P < 0.01). There was a positive correlation between the expression of COX-2 and MMP-2 (P < 0.01). CONCLUSION: COX-2 and MMP-2 expression are increased in NPC, and they may cooperate in the course of lymph metastasis of NPC.

OBJECTIVE: To discuss the expression and correlation of COX2 and LMP1 in NPC. METHOD: Fifty-three nasopharyngeal biopsy specimens of NPC patients who had been diagnosed definitely with pathology in our department from 2000 to 2005, and 8 nasopharyngeal biopsy specimens of chronic nasopharyngitis patients were collected. The expression of COX2, LMP1 were detected with streptavidin peroxidase immunohistochemistry staining. All the datas were analyzed with SPSS 12.0. RESULT: The positive expression rate of COX2 was 71.70% (38/53), and that of LMP1 was 66.04% (35/53) of Nasopharyngeal carcinoma tissue. The higher expression rate was detected in neck lymph nodes metastasis group of nasopharyngeal carcinoma (P<0.05). While the expression rate of COX2, LMP1 was no significant relation with age, gender, clinical stage and pathological classification (P>0.05). The expression of COX2 and LMP1 showed positive correlation (gamma = 0.797, P<0.01). CONCLUSION: COX2 and LMP1 were highly expressed in nasopharyngeal carcinoma cells. The expressions of COX2 and LMP1 was significantly associated with neck lymph nodes metastasis, and was none-significant relation with age, gender, clinical stage and pathological classification. The expression of LMP1 showed a significant positive correlation with that of COX2. LMP1 could enhance NPC neck lymph nodes metastasis by up-regulating the expression of COX2.

OBJECTIVE: To evaluate the expression of vascular endothelial growth factor C (VEGF-C), Epstein-Barr virus (EBV) latent membrane protein 1 (LMP1), cyclooxygenase-2 (COX-2) in nasopharyngeal carcinoma (NPC). METHOD: LMP1, COX-2 and VEGF-C were detects by immunohistochemical staining for 57 case NPC tissue. RESULT: The positives rates of LMP1, COX-2 and VEGF-C detected by immunohistochemical staining were 49.1% (28/57), 75.4% (43/57) and 59.6% (34/57), respectively. The expression of LMP1, COX-2 and VEGF-C were correlated to each other in NPC (P < 0.05). CONCLUSION: LMP1 and COX-2 may induce expression of VEGF-C directly or LMP1 induce expression of VEGF-C by induce COX-2 expression, may contribute to lymph metastasis and develop NPC.

BACKGROUND & OBJECTIVE: The overexpression of P-glycoprotein (P-gp) plays an important role in the chemoresistance of relapsed lymphoma. It is up-regulated in a cyclooxygenase-2 (COX-2)-dependent manner. This study was to observe the expression and correlation of COX-2 and P-gp in B-cell non-Hodgkin's lymphoma (B-NHL), and to investigate their correlations to clinicopathologic features of B-NHL. METHODS: The expression of COX-2 and P-gp in 43 specimens of naive B-NHL and 27 specimens of relapsed B-NHL were detected by SP immunohistochemistry and reverse transcription-polymerase chain reaction (RT-PCR). RESULTS: The positive rates of COX-2 and P-gp were significantly lower in naive B-NHL than in relapsed B-NHL (23.3% vs. 74.1%, 11.6% vs. 70.4%, P<0.01). COX-2 expression was positively correlated to P-gp expression in relapsed B-NHL (r=0.998, P<0.01). The positive rates of COX-2 and P-gp were significantly higher in aggressive B-NHL than in indolent B-NHL (64.3% vs. 10.7%, P=0.002; 52.4% vs. 7.1%, P=0.012). CONCLUSION: COX-2 and P-gp are highly expressed in relapsed B-NHL, and they may play a role in tumor metastasis of B-NHL.

To investigate the components of the prostaglandin biosynthetic pathway, the characteristics of cyclooxygenase (COX)-1, COX-2, hematopoietic prostaglandin D synthase (hPGDS), microsomal prostaglandin E synthase (mPGES), and thromboxane synthase (TXS) were examined for autopsied cases with small cell lung cancer (SCLC) and non-small cell lung cancer (NSCLC). Autopsied patients (n=120) with lung cancer were examined, of whom 25 had SCLC and 95 had NSCLC. Immunostaining was performed for COX-1, COX-2, hPGDS, mPGES, TXS, CD34, vascular endothelial growth factor (VEGF), and basic fibroblastic growth factor (bFGF). The immunostaining scores of the prostaglandin biosynthetic pathway markers were significantly higher for adenocarcinoma than for SCLC (p<0.0001). In addition, there were significant correlations between two markers of the prostaglandin biosynthetic pathway for cases with SCLC and NSCLC. For NSCLC, the mean immunostaining scores for the prostaglandin biosynthetic pathway markers were significantly higher for cases with high count groups than low count groups for MVD, VEGF and bFGF, except COX-2 and MVD, and COX-2 and bFGF. The mean immunostaining scores for COX-1, COX-2, mPGES, and TXS were significantly higher for cases with more metastatic organs who had NSCLC. Prostaglandin biosynthetic pathway markers may act synergistically and enhance tumor angiogenesis, the expression of angiogenic factors, and metastases in patients with NSCLC.

PURPOSE: The aim of this study was to quantitate circulating COX-2 levels in patients with tobacco-related intraoral cancer and to evaluate antitumor activities of COX-2 peptide inhibitors in vitro on KB cell lines. PATIENTS AND METHODS: We used a novel biosensor-based surface plasmon resonance (SPR) technique for estimation of circulating COX-2 levels in 76 patients with oral cancer and 43 normal individuals. Antitumor activities of five COX-2 inhibitory peptides were evaluated using propidium iodide labeling and flow cytometry, alamar blue, MTS, and annexin-V binding assays. RESULTS: Patients with oral cancer showed threefold increase in serum COX-2 level when compared to normal controls (P < 0.0001). Further, late-stage tumors and lymph node metastasis were associated with significant increase in serum COX-2 levels. Patients with higher circulating COX-2 also showed higher immunoreactivity to anti-COX-2 antibody in the lesions. The peptides significantly reduced viability and inhibited growth/proliferation, induced cytotoxicity and apoptosis in tumor cells. However, no such effect was observed either on normal human leukocytes or on MCF-7 cell line that did not over express COX-2. CONCLUSION: Our results indicate that SPR may be a useful proteomic technique for quantitative assessment of COX-2 and to identify patients with high-risk oral premalignant or occult cancer, as well as in monitoring response to novel COX-2 targeting strategies. Furthermore, COX-2 peptide inhibitors appear to be a new class of potent anticancer agent for human oral carcinoma.

Oral squamous cell carcinoma (OSCC) is the sixth most common cancer and frequently metastasizes to the cervical lymph nodes, leading to poor survival of patients with OSCC. However, the mechanism of lymph node metastasis is not fully understood. To clarify the molecular mechanism underlying OSCC metastasis to regional lymph nodes, the highly metastatic fluorescent labeled OSCC cell line SAS-LM3 was successfully established allowing us to monitor the progression of lymph node metastases in a non-invasive manner. SAS-LM3 tumors showed increased lymphangiogenesis and elevated expression of VEGF-C, a potent stimulator of lymphangiogenesis, compared to parental SAS tumors. SAS-LM3 showed high expression of cyclooxygenase-2 (COX-2) compared to parental SAS cells and immunohistochemical analysis demonstrated intense COX-2 expression at the primary site. Inactivation of COX-2 by knockdown or the COX-2 inhibitor NS-398 decreased VEGF-C expression. Administration of COX-2 inhibitor NS-398 in SAS-LM3 tumor-bearing mice suppressed tumor lymphangiogenesis and lymphatic metastases. Collectively, our results indicate that COX-2 promotes tumor lymphangiogenesis and lymph node metastasis of OSCC. COX-2 ablation holds promise as a potential therapeutic approach for lymph node metastasis in OSCC.

PURPOSE: Cervical lymph node metastasis in oral squamous cell carcinoma (OSCC) is recognized as a poor prognostic factor, although its mechanism remains unclear. Recently, cyclo-oxygenase-2 (COX-2) level has been found to correlate highly with vascular endothelial growth factor C (VEGF-C) and lymph node metastasis, as in other solid tumors. However, there has been no report of this correlation in OSCC. Therefore, the aim of this study was to investigate whether COX-2 immunohistochemical expression in OSCC was associated with VEGF-C expression, histopathologic parameters, and lymph node metastasis. MATERIALS AND METHODS: Lymphatic vessel density, VEGF-C, and COX-2 immunohistochemical expression were examined pathologically in 60 specimens of invasive OSCC. Relations of histopathologic parameters to lymph node metastasis were analyzed. RESULTS: expression levels of VEGF-C and COX-2 and lymphatic vessel density in the lymph node metastatic group were significantly higher than in the nonmetastatic group (P < .01). A significant correlation was found between the expression levels of VEGF-C and COX-2 (r = 0.512; P < .001). COX-2 expression was significantly related to lymph node metastasis (P = .004) and VEGF-C expression (P = .005). Univariate analysis showed that survival time was impaired by higher COX-2 and VEGF-C expression levels. Multivariate survival analysis showed that COX-2 expression was an independent prognostic factor. CONCLUSION: This study showed that VEGF-C expression was upregulated by COX-2 in OSCC. High VEGF-C expression appears to promote peritumoral lymphangiogenesis. These data indicated that lymph node metastasis is promoted by COX-2 and VEGF-C in OSCC.

Lysophosphatidic acid (LPA) is a bioactive phospholipids and involves in various cellular events, including tumor cell migration. In the present study, we investigated LPA receptor and its transactivation to EGFR for cyclooxygenase-2 (COX-2) expression and cell migration in CAOV-3 ovarian cancer cells. LPA induced COX-2 expression in a dose-dependent manner, and pretreatment of the cells with pharmacological inhibitors of Gi (pertussis toxin), Src (PP2), EGF receptor (EGFR) (AG1478), ERK (PD98059) significantly inhibited LPA- induced COX-2 expression. Consistent to these results, transfection of the cells with selective Src siRNA attenuated COX-2 expression by LPA. LPA stimulated CAOV-3 cell migration that was abrogated by pharmacological inhibitors and antibody of EP2. Higher expression of LPA2 mRNA was observed in CAOV-3 cells, and transfection of the cells with a selective LPA2 siRNA significantly inhibited LPA-induced activation of EGFR and ERK, as well as COX-2 expression. Importantly, LPA2 siRNA also blocked LPA-induced ovarian cancer cell migration. Collectively, our results clearly show the significance of LPA2 and Gi/Src pathway for LPA-induced COX-2 expression and cell migration that could be a promising drug target for ovarian cancer cell metastasis.

OBJECTIVE: To evaluate the expression of cyclooxygenase (COX)-2 in ovarian serous carcinomas (OSC) and its correlation with microvessel density (MVD), nm23 expression, clinicopathologic prognostic factors and survival. METHODS: Specimens from 44 patients with OSC were evaluated by immunohistochemistry for COX-2 and nm23 expression. tumor MVD was assessed with CD34 immunostaining. The survival data of the patients were found from data files. RESULTS: 40 specimens (90.1%) showed positive COX-2 staining. Patients with high COX-2-expressed tumors had shorter overall survival, but it was not statistically significant. No correlation was found between COX-2 expression and clinicopathologic variables. There was no significant correlation between COX-2 and nm23 expression or MVD. CONCLUSIONS: COX-2 is frequently expressed in OSC. Although we could not confirm the prognostic significance of Cox-2 expression in the present cohort of OSC patients, the p value for overall survival was just slightly greater than alpha, and this result can be referred as almost significant. We considered that the limited number of cases in our study might affect the statistical analysis of our results. Further studies involving a larger number of patients are needed to clarify the prognostic significance of COX-2 expression in ovarian carcinomas.

BACKGROUND AND AIMS: Recent observations suggest an implication of cyclooxygenase-2 (COX-2) in tumor lymphangiogenesis through an upregulation of vascular endothelial growth factor-C (VEGF-C) expression. However, it is unclear whether COX-2 is also associated with VEGF-C expression, tumor lymphangiogenesis and lymph node metastasis in human prostate cancer. METHODS: COX-2 and VEGF-C expression were examined in tumor tissues from 58 prostate cancer patients using immunohistochemical staining. We also analyzed the association of COX-2 and VEGF-C expression with tumor lymphangiogenesis quantified as lymphatic vessel density (LVD), lymph node metastasis, and patients' biochemical progression-free survival (b-PFS). RESULTS: High expression of either COX-2 or VEGF-C was correlated with tumor lymphangiogenesis and lymph node metastasis, as well as poor b-PFS. Moreover, a strong correlation was found between expression of COX-2 and VEGF-C (r = 0.631, p <0.001). CONCLUSIONS: COX-2 is positively associated with VEGF-C expression, tumor lymphangiogenesis and lymphatic metastasis in prostate cancer. These findings suggest that COX-2 may play a pivotal role in lymphangiogenesis and lymph node metastasis of prostate cancer via the regulation of VEGF-C expression.

CpG-oligonucleotides (CpG-ODNs), mimicking bacterial DNA, have recently been shown to stimulate prostate cancer invasion in vitro via Toll-like receptor 9 (TLR9). Since cyclooxygenase 2 (COX-2), frequently overexpressed in multiple tumor types including prostate cancer, is a causal factor for tumor development, invasion and metastasis, an interesting question is raised whether TLR9 regulates COX-2 expression in prostate cancer cells. To address this question, herein we examined COX-2 expression in PC-3 cells stimulated with different doses and time courses of CpG-ODNs. The regulatory role of NF-kappaB in TLR9-mediated COX-2 expression was also investigated. CpG-ODN was found to up-regulate the expression of COX-2 in PC-3 cells in a dose- and time-dependent manner, but have little impact on COX-1 expression. Moreover, CpG-ODN also promoted nuclear translocation and activation of NF-kappaB, which appeared to be required for COX-2 induction by CpG-ODN. Overall, TLR9 up-regulates COX-2 expression in prostate cancer cells, at least partially through the activation of NF-kappaB, which may be implicated in tumor invasion and metastasis.

OBJECTIVE: To investigate the expression of cyclooxygenase-2 (COX-2) in different prostate cancer (PCa) cell lines and its role in the acquisition of invasive and metastatic potentials of PCa. METHODS: We detected the expressions of COX-2 in prostate cancer cell lines LNCaP, C4-2, IF11, IA8 and PC-3 with different metastatic potentials by Western blotting and RT-PCR, and analyzed their roles in the invasion and metastasis of different PCa cell lines. RESULTS: Western blotting and RT-PCR showed that the expression of the COX-2 protein was high in PC-3, but absent in IF11, IA8, LNCaP and C4-2 (P < 0.05), and it was consistent with the expression of COX-2 mRNA. CONCLUSION: COX-2 expresses differently in PCa cell lines with different metastatic potentials. The overexpression of COX-2 may be associated with the high invasion and metastasis of PC-3, but not with the metastasis of other cell lines.

OBJECTIVE: The aim of this study was to determine reliable predictive biomarkers for patients with metastatic renal cell carcinomas (RCCs) who had received cytokine therapy. METHODS: Tissue specimens were obtained from 62 patients with metastatic RCCs between 1995 and 2006. Paraffin wax embedded tissues were immunostained for carbonic anhydrase IX (CAIX), cyclooxygenase-2 (COX-2), and vascular endothelial growth factor (VEGF). RESULTS: Fifty-two specimens (84%) were assessed as clear cell type, with 5, 3, and 2 tumors showing sarcomatoid, papillary, and undifferentiated features, respectively. With a median 54 months of follow-up, 15/18 responding patients (83%) exhibited high CAIX staining compared with only 24/44 (55%) nonresponding patients (odds ratio, OR, 4.1; 95% confidence interval,

BACKGROUND: Prognosis of renal cell carcinoma (RCC) differs within the same stage and grade. Our aim was to investigate the incidence of COX-2 in primary RCC tumors at different stages according to the occurrence of metastasis, and the impact of this biomarker on the survival of RCC patients. PATIENTS AND METHODS: The cytoplasmic/membranous COX-2 protein expression was examined by immunohistochemistry in RCC tumors from 102 patients. The patients were divided into those with: no metastasis during 7.5 years' follow-up (nm), no metastasis at the time of nephrectomy but who later developed metastases (lm), and those with metastasis at presentation (pm). The immunoreactivity of COX-2 was classified as none (absent/weak intensity in fewer than 10% of the cancer cells), low (weak intensity in over 10% of the cancer cells) or high immunostaining (strong intensity in the majority of the cancer cells). In addition p53 and Ki-67 immunostaining was also assessed in tumors. RESULTS: Percentages of COX-2 reaction were (no/low/high): 78/16/7 in the nm, 53/28/19 in the lm, 92/8/0 in the pm groups (p=0.014). Median metastasis-free survival was shorter in lm patients with COX-2-negative tumors when compared to those with COX-2-positive ones (15 vs. 46 months; p=0.020). Median overall survival was shorter in pm/lm patients with COX-2-negative tumors when compared to those with COX-2-positive ones (28 vs. 94 months; p=0.027), and with COX-2-negative/Ki-67-positive tumors when compared to COX-2-positive/Ki-67-negative ones (19 vs. 97 months; p=0.004). Findings for patients with COX-2-negative/p53-positive tumors were similar, with shorter survival compared to those with COX-2-positive/p53-negative ones (19 vs. 97; p=0.006). CONCLUSION: COX-2 protein expression is associated with slow development of metastases, and favourable prognosis in metastatic RCC.

OBJECTIVE: To investigate the expression of cyclooxygenase-2 (COX-2) and vascular endothelial growth factor-C (VEGF-C) in papillary thyroid carcinoma and their relationship to cervical lymph metastases. METHOD: In this study, the expressions of COX-2 and VEGF-C were examined by immunohistochemistry in papillary thyroid carcinoma tissues of 40 patients, and analysis was performed on the correlation of cervical lymph metastases with COX-2 and VEGF-C expression. RESULT: Positive expressions of COX-2 and VEGF-C were 70.0% (28/40) and 75.0% (30/40) respectively in papillary thyroid carcinoma. The positive rates of COX-2 and VEGF-C expression were 80.8% (21/26) and 84.6% (22/26) respectively in patients with cervical lymph metastases, and 50.0% (7/14) and 57.1% (8/14) respectively in patients without cervical lymph metastases, with a statistically significant difference between two groups (P<0.05, all). COX-2 was positively correlated to VEGF-C expression in papillary thyroid carcinoma (r=0.378, P<0.05). CONCLUSION: The results suggest that COX-2 and VEGF-C were highly expressed in papillary thyroid carcinoma, with possible interaction of their expressions, and may play a critical role in the cervical lymph metastases of papillary thyroid carcinoma.