Purpose: The forkhead box transcription factor FoxP3 is specifically expressed in T cells with regulatory properties (Treg). Recently, high numbers of Treg were described to be associated with poor survival in different malignancies. The aim of the presented study was determine the prognostic effect of FoxP3 mRNA expression (reflecting the tissue content of Treg) in ovarian carcinoma and its relation with cytokines, such as IFN-g. Experimental Design: Total RNA was isolated from 99 ovarian carcinoma and from 14 healthy ovarian biopsies. Real-time PCR for FoxP3 was done and correlated with IFN-g-, CD3-, IRF-1-, SOCS-1-, HER-2-, and iNOS expression as well as patients'outcome.The mRNA data was corroborated by FoxP3 immunohistochemistry. Results: Quantitation of FoxP3 expression identified a patient subgroup (>81th percentile), which is characterized by a significantly worse prognosis in terms of overall survival (27.8 versus 77.3 months, P = 0.0034) and progression-free survival (18 versus 57.5 months; P = 0.0041). FoxP3 expression correlated with IFN-g, IRF-1, and CD3 expression. High FoxP3 expression represents an independent prognostic factor for overall survival (P = 0.004) and progression-free survival (P = 0.004). Conclusions: High expression levels of FoxP3 might represent a surrogate marker for an immunosuppressive milieu contributing to tumor immune escape. Strategies selectively depleting Treg might improve the antitumor activity of endogenously arising tumor-reactiveTcells and immunotherapies using vaccines or antibodies.Cancer patients can harbor significant numbers of CD8 and CD4 T cells specific for tumor antigens. In most cases, tumorreactive T cells fail to eradicate the tumor in vivo because they seem to be actively maintained in an unresponsive state. Several immune-evasion strategies of malignant tumors have been described thus far (reviewed in ref. 1). Recently, regulatory T cells (Treg), which are characterized by coexpression of CD4 and CD25, have also been attributed to contribute to cancer-related immunosuppression (2 -5). Treg represent 5% to 10% of total CD4 + T cells and are crucial for the repression of autoimmune disorders and transplant rejection (6). The activation of Treg is antigen specific; however, inhibition of CD4 + and CD8 + T cells seems to be antigen nonspecific. Very recent evidence showed that Treg abrogate CD8 + T cell -mediated tumor rejection in a transforming growth factor-h -dependent manner (7). The suggestion of a role for Treg in cancer-induced immunosuppression in humans arises from the observation that patients suffering from a variety of cancer types have an enlarged pool of Treg in the peripheral blood, tumor-draining lymph nodes, and in the tumor itself (8 -12). Exact characterization of Treg has been hampered by the lack of specific cell surface markers. The observation that autoimmune diseases occur in both humans and mice lacking functional FoxP3 (13) indicates that this transcription factor plays a crucial role in the regulation of Treg function. ...

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CD4؉ CD25 ؉ regulatory T cells (

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Serum levels of vascular endothelial growth factor (VEGF-S) have been reported to correlate with tumor stage and prognosis in various human malignancies. The source of soluble VEGF in peripheral blood remains obscure. We therefore measured the concentration of immunoreactive VEGF in 241 serum samples and 61 plasma samples (VEGF-P) from 20 subjects undergoing myeloablative chemotherapy and from 3 normal platelet donors. A significant correlation between the peripheral blood platelet count (PC) and VEGF-S (r = 0.86) but not VEGF-P was found. VEGF-S levels were 58.43 ± 42.50 pg/ml (mean ± SD) in patients with a PC < 50 × 10⁹/l, 203.29 ± 176.56 pg/ml for a PC of 50–150 × 10⁹/l, and 457.42 ± 475.41 pg/ml for a PC > 150 × 10⁹/l. Interestingly, VEGF-P levels were substantially lower than the corresponding VEGF-S values, namely below the detection limit in most cases. Supernatants from platelet-rich plasma contained no VEGF, but after in vitro lysis of the platelets very high VEGF levels were found. The VEGF content per 10⁹ platelets was calculated at 2.51 ± 2.39 pg and was dependent on the mean platelet volume. In summary, VEGF release from platelets during blood clotting was found to be the main source of VEGF in serum samples. Cancer patients in clinical remission have negligible amounts of soluble VEGF in peripheral blood, and myeloablative chemotherapy causes a significant drop in VEGF-S levels corresponding to the decrease in PC. Thus, studies addressing the diagnostic and prognostic value of VEGF-S in cancer patients must be interpreted with caution. Our data provide the basis for predicting VEGF-S in relation to PC in vivo, and for reevaluating former studies of VEGF-S in patients with malignant or nonmalignant disease.

Recent studies have demonstrated cyclooxygenase 2 (COX-2) overexpression in various human malignancies, especially in breast cancer, where COX-2 turned out to be a predictor of poor survival. To evaluate the relation of COX-2 and Ep-CAM overexpression and its prognostic significance, we performed a retrospective study on 212 breast cancer patients with a median follow-up time of 10.5 years. Overexpression of COX-2 in tumour tissue samples was assessed by immunohistochemistry. COX-2 overexpression was found in 48.6% of the tumour samples and was predictive for poor disease-free and overall survival. Univariate analysis revealed a strong correlation between COX-2 and Ep-CAM overexpression (P ¼ 0.009). Concurrent COX-2 and Ep-CAM overexpression was present in 21.7% of tumour specimens and had an additive negative impact on disease-free and overall survival. Determination of both tumour markers should help in guiding new therapeutic strategies in patients with invasive breast cancer. Cyclooxygenase-2 (COX-2) is a prostaglandin synthase that catalyses the synthesis of prostaglandin G 2 (PGG 2 ) and PGH 2 from arachidonic acid. Recent studies have led to the recognition of the importance of COX-2 in tumorigenesis of different tumour types. It has been shown that COX-2 is involved in tumour angiogenesis (Tsujii et al, 1998;Gately, 2000), in suppression of apoptosis (Sheng et al, 1998) and in the promotion of invasiveness (Tsujii et al, 1997). COX-2 overexpression was found in pancreatic (Molina et al, 1999;Okami et al, 1999;Kokawa et al, 2001), oesophageal (Zimmermann et al, 1999), prostate (Yoshimura et al, 2000), lung (Khuri et al, 2001), head and neck cancers (Chan et al, 1999) and in malignant gliomas (Shono et al, 2001). Tsujii et al reported that COX-2 overexpression in intestinal epithelial cells leads to downregulation of adhesion molecules (i.e. cadherins), resulting in an enhanced tumorigenic potential (Tsujii and DuBois, 1995).Enhanced COX-2 expression in breast cancer was first indicated by reports of elevated prostaglandin levels in breast carcinomas (Bennett et al, 1977), particularly in patients with metastatic disease (Rolland et al, 1980). A key role of COX-2 for the initiation and progression of breast cancer is suggested by the finding that mere overexpression of COX-2 can be sufficient for inducing mammary gland tumorigenesis in transgenic mice (Liu et al, 2001). Notably, in human breast cancer cell lines, a positive correlation was found between invasiveness, metastatic potential and prostaglandin production (Liu and Rose, 1996). Different groups have described the prognostic significance of COX-2 overexpression in breast cancer (Hwang et al, 1998;Ristimaki et al, 2002;Soslow et al, 2000).We have recently described the prognostic significance of Ep-CAM overexpression in patients with invasive breast cancer (Gastl et al, 2000). Ep-CAM (also called 17-1A, ESA, EGP40, 323/A3) is a 40-kDa transmembrane glycoprotein expressed on most human epithelial cells (Gottlinger et al, 1986). The Ep-CAM glycoprote...

A critical event in tumor growth and progression is the upregulation of angiogenesis. Thus, targeting angiogenesis has become an attractive treatment modality in cancer medicine. Our study analyzed various solid tumor types for the expression of Dkk-3, a cystein-rich, N-glycosylated secreted member of the dickkopf protein family that has been proposed as a tumor suppressor gene. Tissue microarrays of gliomas (n 5 30), high-grade non-Hodgkin's lymphomas (NHL, n 5 80), colorectal cancer (n 5 35) and melanoma (n 5 30) were immunohistochemically analyzed for Dkk-3 and CD31 expression. Moreover, the effects of Dkk-3 were studied in vitro in primary endothelial colony-forming cells (ECFC) and in vivo in a mouse melanoma model. In comparison to normal tissue, the number of blood vessels expressing Dkk-3 was increased in glioma, high-grade NHL, melanoma and colorectal carcinoma. Confocal laser scanning microscopy revealed that Dkk-3 vesicles localized also in Weibel Palade bodies. In vitro cell proliferation and migration of ECFC was not significantly affected by adenoviral overexpression or siRNA-mediated downregulation of Dkk-3. Interestingly, tube formation in matrigel decreased after downregulation of Dkk-3 and increased after adenoviral overexpression. Stable overexpression of murine Dkk-3 in B16F10 cells significantly increased microvessel density in the C57/BL6 melanoma model. Thus, we postulate a novel function of Dkk-3 in endothelial cells as a differentiation factor involved in remodeling the tumor vasculature. ' 2007 Wiley-Liss, Inc.Key words: angiogenesis; glioma; lymphoma; melanoma; colorectal cancer; Dickkopf-3 Growth, progression and metastasis of solid tumors are strongly dependent on the supply of oxygen and nutrients. 1 Without adequate blood supply tumors become hypoxic, acidotic and necrotic. Therefore, growing tumors induce an angiogenic switch 2 and produce a variety of growth factors that are responsible either for remodeling of the pre-existing vascular network by means of angiogenic sprouting or intus-susceptive growth or for neovascularization by recruiting circulating endothelial cells. 3 Capillaries of the tumor vascular bed show a variety of structural abnormalities, such as the lack of a basement membrane, low numbers of surrounding pericytes, mosaic vessel structure and altered expression patterns of leukocyte adhesion molecules. 4 On the molecular level, tumor endothelial cells show a distinct gene expression profile 5 that differs from that of normal endothelial cells. 6 Thus, markers specifically expressed or overexpressed on tumor endothelial cells or secreted into the perivascular space are potential candidates for vascular targeting. 7 Among the dickkopf family of secreted modulators of wnt signalling, Dkk-3 is the most divergent member with still unknown signal transduction and function. 8 Dkk-1 and Dkk-2 have been shown to bind to Kremen receptors, resulting in down-regulation of the Wnt-coreceptor lipoprotein-related protein from the cell surface. 9-11 Dkk-1 knock-out mice show se...