FGF18 as a potential biomarker in serous and mucinous ovarian tumors.

Fibroblast growth factor 18 (FGF18) has been suggested to play important roles in promoting progression of ovarian high-grade serous carcinoma. Our aim was to investigate FGF18 expression in the whole spectrum of serous and mucinous ovarian tumors, highlighting differences in expression within the adenoma-carcinoma sequence and differences between type I and type II tumors. We also aimed to test the prognostic significance of this expression and its relation to microvessel density (MVD). We evaluated the immunohistochemical expression of FGF18 and CD31 in 103 ovarian tumors and statistically analyzed their association with clinicopathological variables and patients' outcome. FGF18 score increased significantly within the adenoma-carcinoma sequence for serous and mucinous tumors. MVD increased significantly only among serous tumors. FGF18 and MVD correlated significantly (overall and among serous tumors only) and were significantly higher in type II than type I tumors. Cox regression models were built. Independent predictors could not be determined due to multicollinearity between the predictors. However, the combination of International Federation of Gynecology and Obstetrics (FIGO) stage, ovarian carcinoma type, and/or FGF18 score achieved the highest predictability of poor prognosis. FGF18 could play a role within the adenoma-carcinoma sequence in type I tumors and might modulate angiogenesis among serous tumors. Our findings further augment the differences between type I and type II tumors. The combination of FIGO stage, ovarian carcinoma type, and/or FGF18 score could predict poor prognosis among ovarian carcinoma patients. Our work identifies FGF18 in ovarian neoplasia as a promising field of research, although evaluation of the performance of the developed models is still needed.

Antitumor effect of FP3 on a patient-derived tumor tissue xenograft model of rectal carcinoma.

BACKGROUND/AIMS: FP3 is an engineered protein which contains the extracellular domain 2 of VEGF receptor 1 (Flt-1) and extracellular domain 3 and 4 of VEGF receptor 2 (Flk-1, KDR) fused to the Fc portion of human immunoglobulin G1. Previous studies demonstrated its antiangiogenic and antitumor effects in vitro and in vivo. METHODOLOGY: In this study, a PDTT xenograft model of rectal carcinoma was established for assessment of the antitumor activity of FP3. Xenografts were treated with FP3 or bevacizumab (Avastin). After tumor growth was confirmed, volume and microvessel density in tumors were evaluated. Levels of VEGF and PCNA in the tumor were examined by immunohistonchemical staining and western blotting. RESULTS: FP3 showed significant antitumor activity in the PDTT xenograft model of rectal carcinoma. The microvessel density in tumor tissues treated with FP3 was lower than that of the control. Antitumor activity of FP3 was similar to that of bevacizumab in the PDTT xenograft model of rectal carcinoma. CONCLUSIONS: This study indicated that FP3 could be used as an effective antiangiogenic and antitumor agent in treatment of colorectal carcinoma.

Adnexal mass vascularity assessed by 3-dimensional power Doppler: does it add to the risk of malignancy index in prediction of ovarian malignancy?: four hundred-case study.

The Risk of Malignancy Index (RMI) is used for the prediction of ovarian malignancy. It includes menopausal status, carbohydrate antigen 125 serum levels, and ultrasound criteria. Three-dimensional power Doppler (3-DPD) is a reproducible investigation for assessment of tumor vascularity, classifying vascularity to avascular, parallel, and chaotic patterns. In this study; 3-DPD was added to RMI for prediction of malignancy in 400 cases of ovarian masses. Sensitivity of RMI for prediction of malignancy was 88%, with a cutoff value of 202.5 at 95% confidence interval. Sensitivity of 3-DPD for prediction of malignancy was 75%, adding 3-DPD to RMI increased its sensitivity to 99%. Considering the pilot nature of the study, further studies are needed to corroborate such findings.

The reduction in pigment epithelium-derived factor is a sign of malignancy in ovarian cancer expressing low-level of vascular endothelial growth factor.

BACKGROUND: Angiogenesis is a critical factor in the progression of solid tumors and metastasis. The aim of this study was to characterise the roles of angiogenic and anti-angiogenic factors on ovarian cancer. METHODS: The expression levels of vascular endothelial growth factor (VEGF, angiogenic factor) and pigment epithelial growth factor (PEDF, anti-angiogenic factor) were measured by real-time polymerase chain reaction and Western blotting in ovarian tumors. Microvessel density (MVD) was evaluated by the total microvessel length in high-power field of tumor tissue preparations. RESULTS: MVD correlated with tumor malignancy. The tissues with the highest expression levels of VEGF (VEGF-H) were malignant tumors. The VEGF expression levels in some malignant tumors (VEGF-L) were as low as that in benign tumors. Therefore, the expression of PEDF was examined. The PEDF expression levels in VEGF-L malignant tumors were significantly lower than those in benign tumors. On the other hand, the PEDF expression levels in VEGF-H malignant tumor tissues were not significantly different from those in benign tumors. CONCLUSION: The reduction in PEDF expression levels may be, in part, responsible for tumor malignancy in VEGF-L ovarian tumors. Furthermore, PEDF may be a useful marker of malignancy in VEGF-L ovarian tumors.

Inducible nitric oxide synthase expression, apoptosis, and angiogenesis in in situ and invasive breast carcinomas.

In this investigation, we studied the expression of inducible nitric oxide synthase (iNOS) and its association to apoptosis and angiogenesis in 43 in situ and 68 invasive breast carcinomas. Its expression was studied immunohistochemically using a polyclonal iNOS antibody, and the staining was evaluated both in tumor and stromal cells. Apoptosis was detected by 3' end labeling of fragmented DNA (terminal deoxynucleotidyl transferase-mediated nick end labeling method). Vascularization was detected immunohistochemically using an antibody to the FVIII-related antigen, and calculated microvessel densities were determined. In addition to strong iNOS expression in stromal cells, iNOS positivity was observed in tumor cells in 46.5% of in situ and 58.8% of invasive carcinomas. In invasive carcinomas, there were more cases with iNOS positivity both in tumor and stromal cells compared to in situ carcinomas (0.007). The proportion of cases with iNOS-positive tumor cells increased in in situ carcinomas from grade I to III (20.0%, 46.2%, and 73.3%). In invasive ductal carcinomas, there were more cases with iNOS-positive tumor cells than with in situ carcinomas (P = 0.04). Carcinomas with both iNOS-positive tumor and stromal cells had a higher apoptotic index (P = 0.02) and a higher calculated microvessel densities index (P = 0.02). A high number of iNOS-positive stromal cells associated with metastatic disease (P = 0.05). The results show that breast carcinoma cells, in addition to stromal cells, express iNOS and are capable of producing NO. Carcinomas with iNOS-positive tumor and stromal cells have a higher apoptotic indices and increased vascularization, suggesting that iNOS contributes to promotion of apoptosis and angiogenesis in breast carcinoma. The association of the number of iNOS-positive stromal cells with metastatic disease might be attributable to stimulation of angiogenesis, resulting in a higher vascular density and consequently a higher probability for tumor cells to invade.

Vascular stroma formation in carcinoma in situ, invasive carcinoma, and metastatic carcinoma of the breast.

The generation of vascular stroma is essential for solid tumor growth and involves stimulatory and inhibiting factors as well as stromal components that regulate functions such as cellular adhesion, migration, and gene expression. In an effort to obtain a more integrated understanding of vascular stroma formation in breast carcinoma, we examined expression of the angiogenic factor vascular permeability factor (VPF)/vascular endothelial growth factor (VEGF); the VPF/VEGF receptors flt-1 and KDR; thrombospondin-1, which has been reported to inhibit angiogenesis; and the stromal components collagen type I, total fibronectin, ED-A+ fibronectin, versican, and decorin by mRNA in situ hybridization on frozen sections of 113 blocks of breast tissue from 68 patients including 28 sections of breast tissue without malignancy, 18 with in situ carcinomas, 56 with invasive carcinomas, and 8 with metastatic carcinomas. A characteristic expression profile emerged that was remarkably similar in invasive carcinoma, carcinoma in situ, and metastatic carcinoma, with the following characteristics: strong tumor cell expression of VPF/VEGF; strong endothelial cell expression of VPF/VEGF receptors; strong expression of thrombospondin-1 by stromal cells and occasionally by tumor cells; and strong stromal cell expression of collagen type I, total fibronectin, ED-A+ fibronectin, versican, and decorin. The formation of vascular stroma preceded invasion, raising the possibility that tumor cells invade not into normal breast stroma but rather into a richly vascular stroma that they have induced. Similarly, tumor cells at sites of metastasis appear to induce the vascular stroma in which they grow. We conclude that a distinct pattern of mRNA expression characterizes the generation of vascular stroma in breast cancer and that the formation of vascular stroma may play a role not only in growth of the primary tumor but also in invasion and metastasis.

Thymidine phosphorylase-mediated angiogenesis regulated by thymidine phosphorylase inhibitor in human ovarian cancer cells in vivo.

Metastasis or progression of ovarian cancer cells is known to be due to the action of various angiogenic factors. We determined the expression of thymidine phosphorylase/platelet-derived endothelial cell growth factor (TP/PD-ECGF) and vascular endothelial growth factor (VEGF) in cell lines established from 3 serous adenocarcinomas, 3 clear cell carcinomas and 2 mucinous carcinomas of the human ovary. TP activity and the TP mRNA level were much higher in the serous adenocarcinoma cells than in the clear cells and mucinous carcinoma cells, and TP expression was extremely low in the clear cell carcinoma cells. Expression of VEGF mRNA was variable, but not significantly different between the 3 histological types of ovarian cancer. In vivo angiogenesis in the ovarian cancer cells was evaluated by the dorsal air sac assay and revealed that SHIN-3 and HRA serous adenocarcinoma cells, which have high levels of TP expression, induced angiogenesis, while KK clear cell carcinoma cells with low TP expression, did not. The degree of ovarian-cancer-induced angiogenesis seemed to be independent of expression of VEGF in the cells. To confirm that the serous adenocarcinoma-induced angiogenesis is dependent on TP levels, a potent and specific inhibitor of TP was administered orally to mice implanted with a chamber containing SHIN-3 or HRA cells. The TP inhibitor significantly inhibited the angiogenesis induced by the serous adenocarcinoma cells. These results suggest that the angiogenic potency of ovarian cancer cells differs with the histological type and is controlled by expression of TP/PD-ECGF, not by VEGF, and that TP-mediated angiogenesis may be the main factor responsible for progression or metastasis of ovarian serous adenocarcinomas.

Interleukin 10 expression is correlated with thrombospondin expression and decreased vascular involvement in colon cancer.

Interleukin 10 (IL-10) is an immuno-suppressive cytokine produced by T-lymphocytes, and a regulatory molecule for angiogenesis in various cancers. We examined IL-10 gene expression in 53 colon cancer patients who underwent surgical resection. IL-10 gene expression was correlated with TSP1 and TSP2 gene expression (P=0.0049, P=0.0285). Colon cancer with IL-10 gene expression (19/53) showed significantly decreased venous involvement (P=0.0433). The mean vessel counts in the colon cancers with IL-10 gene expression were significantly lower than those without IL-10 gene expression (P<0.001). These results suggested that IL-10 stimulates angiostatic factor gene expression, and results in suppression of venous involvement.

Mucinous colon carcinomas with microsatellite instability have a lower microvessel density and lower vascular endothelial growth factor expression.

We determined the association between cyclooxygenase-2 (COX-2), vascular endothelial growth factor (VEGF) expression, microvessel density (MVD) and microsatellite instability (MSI) or the histological type in colon adenocarcinomas. Sixty-six cases were studied, 28 MSI+ and 38 MSI-. MSI phenotype was determined using polymerase chain reaction. MVD was assessed after CD31 staining in ten x400 fields (0.96 mm(2)) in the most vascularized areas. VEGF and COX-2 expression were studied by means of immunohistochemistry. MVD positively correlated with the levels of VEGF expression (P=10(-4)) and also with the levels of COX-2 expression (P=0.007). MVD and VEGF expression were lower in MSI+ carcinomas (P=0.002 and P=0.03 respectively). When mucinous tumors were excluded from the statistical analysis, the association between low MVD, low VEGF and MSI status disappeared (P=0.5, P=1, respectively). MSI+ mucinous carcinomas had a lower MVD and VEGF expression than other MSI+ carcinomas (P=0.008 and P=0.004, respectively) and MSI- mucinous carcinomas (P=0.01 and P=0.001, respectively). COX-2 expression was lower in medullary carcinomas (P=0.001). In conclusion, mucinous MSI+ colon carcinomas represent a special group of colon adenocarcinomas relating to angiogenesis, with a lower MVD and VEGF expression than both MSI- mucinous carcinomas and MSI+ non-mucinous carcinomas. A low COX-2 expression could be related to the medullary phenotype. However, this has to be confirmed in a larger series. Finally, the low MVD of MSI+ mucinous colon adenocarcinomas could participate in their overall better prognosis.

The influence of theobromine on angiogenic activity and proangiogenic cytokines production of human ovarian cancer cells.

Angiogenesis plays an important role in ovarian cancer growth and metastasis formation. Adenosine is one of the most potent stimulator of neovascularisation. The aim of present study was to determine if theobromine, adenosine receptor antagonist, influences angiogenic activity and proangiogenic cytokines production. Theobromine caused significant inhibition of angiogenic activity of ovarian cancer cells. In in vivo and in vitro cultures theobromine diminished vascular endothelial growth factor (VEGF) production. Production of basic fibroblast growth factor (bFGF) and interleukin-8 (IL-8) was not altered by the examined drug. These findings suggest that theobromine might be a potent inhibitor of angiogenesis induced by ovarian cancer cells and its mechanism of action is related to inhibition of VEGF production.

Combretastatin A-1 phosphate potentiates the antitumour activity of cisplatin in a murine adenocarcinoma model.

BACKGROUND: The tubulin depolymerizing drug Combretastatin A-1 phosphate (CA1P), a water-soluble derivative of combretastatin A-1, has been recently shown to have a better efficacy in experimental models than the clinically active, close structural analogue, combretastatin A-4 phosphate (CA4P). Previous studies with CA4P in combination with standard anti-cancer agents have demonstrated improved efficacy relative to the standard agents. MATERIALS AND METHODS: In this study the synergistic effects of administering CA1P in combination with cisplatin (CPL) in a well-differentiated transplantable murine colon model (MAC 29) was evaluated. RESULTS: CA1P at 100 mgkg-1 significantly potentiated the anti-tumour effects of CPL. The effect with CPL was similar to that seen for CA1P at its maximum tolerated dose (MTD) alone. CONCLUSION: These data demonstrate that the combination of CA1P and CPL has significant preclinical antitumour activity against a transplantable murine adenocarcinoma model that is related to the antivascular effects of CA1P.

Immunohistopathological characterizatin of spontaneous metastases in a human lung mucoepidermoid adenocarcinoma (HLMC) xenograft.

The most common clinical form of lung cancer is a disseminated disease with distant metastases; several years of cancer progression precede presentation, and this ultimately limits the efficacy of curative therapy. In this immunohistochemical study, we examined a mucinous adenocarcinoma cell line, maintained by xenogeneic transplantation, and a spontaneous metastatic variant which produces distant tumors (in liver, spleen and kidney). The aim was to investigate possible parameters which characterize the metastatic process. Histopathological comparison between the two subcutaneous transplanted tumor lines showed that both lines presented a similar cellular morphology, a different pattern of cellular growth and an increased vascularization in the metastatic line with respect to its parent. All the tumor sections expressed differential immune reactivity with monoclonal antibodies against Lewis y (MAb C14), sialyl-Lewis x (MAb SNH3) and Lewis x (MAb FH2) determinants. Neither expressed MUC 1 mucins detectable with monoclonal antibodies reactive with the mucin protein core (MAbs C595 and SM3) nor was carcinoembryonic antigen (MAb C365) expressed. Neoplastic cells were reactive with an anti-pan cytokeratin monoclonal antibody confirming their epithelial histogenesis. Our findings have been evaluated with respect to defining metastatic phenotypes in lung cancer by examination of distinct histopathological and immunological parameters.

Relationship between inducible nitric oxide synthase expression and angiogenesis in primary gallbladder carcinoma tissue.

AIM: To explore the relationship between angiogenesis and biological behaviors of primary gallbladder carcinoma (PGBC), the relationship between the expression of inducible nitric oxide synthase (iNOS) and biological behaviors of PGBC and its relationship with the expression of iNOS and angiogenesis of PGBC. METHODS: The expression of iNOS and micro-vessel density (MVD) were assessed by immunohistochemical method and image analysis system in 40 specimens of PGBC and in 8 specimens of normal gallbladder. The immunostaining results and related clinicopathologic materials were analyzed by statistical methods. RESULTS: MVD in PGBC was significantly higher than that in normal gallbladder tissue (46+/-14 vs 14+/-6, P<0.05), and was not related with age, gender, tumor size and histological type. MVD of poorly and undifferentiated tumor tissues was higher than that of moderately-differentiated and well-differentiated tumor tissues (52+/-9 vs 43+/-9 vs 33+/-6, P<0.01). MVD of Nevin IV and V stages was higher than that of Nevin I, II and III stages (52+/-8 vs 37+/-13, P<0.01). MVD of cases with lymphatic or liver metastasis was significantly higher than that without liver metastasis (55+/-6 vs 42+/-10, P<0.05)or lymphatic metastasis (53+/-8 vs 38+/-8, P<0.01). The positive level index (PLI) of iNOS in PGBC was 0.435+/-0.134, and was not related with age, gender, tumor size, histological type, differentiation and clinical stage of PGBC. The PLI of iNOS in cases with lymphatic metastasis was higher than that without lymphatic metastasis (0.573+/-0.078 vs 0.367+/-0.064, P<0.01). The PLI of iNOS in cases with liver metastasis was higher than that without liver metastasis (0.533+/-0.067 vs 0.424+/-0.084, P<0.05). There was a significant correlation between PLI of iNOS and MVD in PGBC (P<0.05). CONCLUSION: Angiogenesis of PGBC is significantly related to the biological behaviors of PGBC. The expression of iNOS is related to the biological behaviors of PGBC. The detection of MVD and the expression of iNOS in PGBC can be used as parameters to determine the degree of malignancy and prognosis.

[Inhibitory action of Chinese compound prescription for reinforcing vital energy and invigorating blood circulation on cancer cell multiplication].

Experiments have proved that the Chinese compound prescription for reinforcing vital energy and invigorating blood circulation markedly helps to improve the morphology of cancer cell nucleolus and membrane surface microvillus, amend the composition of microtubulin and facilitate intercellular gap junctional communication. Cell cycle kinetics shows that the multiplication of cancer cells in human stomach is checked mainly at G2M stage.

STAT5b as molecular target in pancreatic cancer--inhibition of tumor growth, angiogenesis, and metastases.

The prognosis of patients suffering from pancreatic cancer is still poor and novel therapeutic options are urgently needed. Recently, the transcription factor signal transducer and activator of transcription 5b (STAT5b) was associated with tumor progression in human solid cancer. Hence, we assessed whether STAT5b might serve as an anticancer target in ductal pancreatic adenocarcinoma (DPAC). We found that nuclear expression of STAT5b can be detected in approximately 50% of DPAC. Blockade of STAT5b by stable shRNA-mediated knockdown showed no effects on tumor cell growth in vitro. However, inhibition of tumor cell motility was found even in response to stimulation with epidermal growth factor or interleukin-6. These findings were paralleled by a reduction of prometastatic and proangiogenic factors in vitro. Subsequent in vivo experiments revealed a strong growth inhibition on STAT5b blockade in subcutaneous and orthotopic models. These findings were paralleled by impaired tumor angiogenesis in vivo. In contrast to the subcutaneous model, the orthotopic model revealed a strong reduction of tumor cell proliferation that emphasizes the meaning of assessing targets in an appropriate microenvironment. Taken together, our results suggest that STAT5b might be a potential novel target for human DPAC.