Thymidine phosphorylase expression in endometrial carcinomas

Thymidine phosphorylase (TP) is a potent angiogenic molecule shown to induce endothelial cell migration and proliferation. We investigated the expression of TP in a series of 156 endometrial carcinomas, using immunohistochemical methods. Histopathological parameters of known prognostic significance and the molecular factors of p53, bcl-2 and angiogenesis were also assessed. Thymidine phosphorylase was expressed in cancer cells, stromal fibroblasts and myometrial cells. The pattern of TP staining was nuclear or mixed nuclear/cytoplasmic, and only exceptionally was purely cytoplasmic. An exclusively cytoplasmic staining was documented for the tumour-associated foamy macrophages. Cancer cell reactivity was rather limited; only 3.2% of endometrial carcinomas expressed TP in more than 50% of the neoplastic cell population and only 12% expressed the enzyme in more than 10% of the cancer cells. By contrast, TP reactivity was frequent in the fibroblasts of the tumour supporting stroma and the fibroblasts/myometrial cells at the invading tumour front, where approximately 1/3 of the cases expressed TP in more than 50% of the respective constituent cells. A high TP reactivity in the stromal fibroblasts was significantly associated with the presence of foamy macrophages and an intense lymphocytic response. A high TP reactivity at the invading tumour front was significantly associated with an intense lymphocytic response and the adverse prognostic parameters of high tumour grade, deep myometrial invasion, advanced stage of disease and the non-endometrioid carcinomas. There was no significant association of cancer cell TP reactivity with any of the parameters studied, including nuclear p53 accumulation, cytoplasmic/perinuclear bcl-2 expression, microvessel density (MVD) and prognosis. Similarly, no relationship was established between fibroblastic or fibroblastic/myometrial TP reactivity and MVD. It is concluded that TP is not a major angiogenic factor in endometrial carcinomas. However, a prominent TP activity at the invading tumour front, which is probably induced by cytokines of histiocytic and lymphocytic origin, may promote tumour invasion and progression.     

Endometrial carcinoma: association of steroid hormone receptor expression with low angiogenesis and bcl-2 expression

In endometrial tissues, malignant change may be accompanied by a loss of hormone dependence which is, usually, reflected in a parallel loss of oestrogen and progesterone receptors (ER and PR). In this study, the steroid receptor status of 164 endometrial carcinomas was related to intratumoural angiogenesis and the apoptotic proteins bcl-2 and p53. Relationships to conventional histopathological features and patient survival were also sought. Immunohistochemistry was performed on formalin-fixed, paraffin-embedded tissues. The mean follow-up was 55 months (range 19-167 months). Specific nuclear staining for ER and PR was detected in 35% and 32% of endometrial carcinomas, respectively, and was very commonly co-expressed (P<0.0001). The failure of demonstrating a steroid receptor complement in endometrial neoplasms was, in general terms, an adverse prognostic sign. Thus, ER or PR loss was significantly associated with non-endometrioid carcinomas (ER P=0.01; PR P=0.004) and with deep myometrial invasion (ER P<0.0002), high intratumoural angiogenesis (PR P<0.01) and the absence of bcl-2 expression (PR P<0.005). There was a trend for patients with ER or simultaneous ER/PR expression to have an improved survival, but this association did not reach the level of statistical significance. In multivariate analysis (all stages), tumour cell type (endometrioid versus non-endometrioid carcinomas) and stage of disease were the only variables associated with prognosis (P=0.01 and P<0.0001, respectively), with tumour cell type retaining its independent prognostic value and within stage-I endometrial carcinomas (P=0.02). It is suggested that the loss of steroid hormone receptors in endometrial carcinomas is associated with a more aggressive phenotype and the switching-on of angiogenic pathways.     

Macrophage infiltration is associated with VEGF and EGFR expression in breast cancer

Angiogenesis is esential for tumour growth and metastasis. Vascular endothelial growth factor (VEGF) is a potent endothelial cell mitogen and is an important component of the angiogenic stimulus in a range of human neoplasias. In addition to its mitogenic activities, VEGF has also been found to stimulate migration in macrophages via the flt-1 VEGF receptor. It has previously been shown that increased focal tumour macrophage infiltration is associated with increased angiogenesis and worsened relapse-free and overall survival in breast cancer. Macrophages are able to stimulate angiogenesis by their production of a range of factors including VEGF, tumour necrosis factor-alpha (TNF-alpha), and thymidine phosphorylase (TP). Thus, in breast cancer, VEGF could have a dual role in the regulation of angiogenesis, by direct mitogenic stimulation of endothelial cells, and also indirectly by attracting macrophages into avascular tumours. The purpose of this study was to localize VEGF protein in a series of 96 consecutive primary breast carcinomas and to determine its relationship to focal macrophage infiltration (macrophage index). These two variables were also compared with the pathological features of the tumours, as well as oestrogen receptor (ER), epidermal growth factor receptor (EGFR), microvessel density, macrophage index, and survival. An inverse relationship (p=0.0006) was noted between VEGF and EGFR, with high VEGF expression correlating with low EGFR levels. In the EGFR-negative group of cases (n=56), positive associations were observed between VEGF expression and macrophage index (p=0.005), ER (p=0.05), p53 (p=0. 006), tumour grade (p=0.02), and tumour necrosis (p=0.03). Macrophage counts were higher in EGFR-positive tumours (p=0.0006) and no associations were found between VEGF expression and increased microvessel density. These results show that in breast cancers there are two types of macrophage infiltrates, one associated with the presence of EGFR and low VEGF expression in tumours and the other with high VEGF expression in EGFR-negative tumours. VEGF expression may be an important factor in the recruitment of tumour-associated macrophages into breast carcinomas and may thus have an additional, indirect, pathway of angiogenic stimulation in this type of tumour.     

HER2 is unlikely to be involved in directly regulating angiogenesis in human breast cancer.

Angiogenesis is a fundamental component of oncogenesis. Angiogenic factors such as vascular endothelial growth factor (VEGF) and platelet derived-endothelial cell growth factor/thymidine phosphorylase (PD-ECGF/TP) are generated from tumor cells to provide tumor growth and are thought to be regulated via the HER2 oncogene, whose amplification is the most common genetic alteration in breast cancer. The present study aimed to evaluate the immunoreactivity of angiogenic factors (VEGF, PD-ECGF/TP) and microvessel density (MVD) via epidermal growth factor receptor (EGFR) and HER2, and to correlate their expression with clinicopathologic features. Two hundred one invasive human breast cancer specimens were tested immunohistochemically for the expression of these proteins. In addition, MVD was examined using computerized image analysis. VEGF could be an additional interesting prognostic variable, as it was significantly associated with tumor grade (P=0.002), stage (P=0.018), and negative estrogen receptor status (P=0.011). EGFR was significantly related to invasive ductal carcinoma (P=0.030), tumor grade (P=0.009), VEGF expression (P=0.013), PD-ECGF/TP expression (P=0.024), and MVD (P=0.050). The finding that VEGF is not correlated to MVD does not rule out a crucial role of VEGF as a key factor in angiogenesis. HER2 could not be correlated to MVD, VEGF expression, or PD-ECGF/TP expression, indicating that this factor is unlikely to be involved in directly regulating angiogenesis, whereas the significant correlations between EGFR and histologic tumor type, tumor grade, the angiogenic factors VEGF and PD-ECGF/TP, and MVD point out that EGF is the major modulating growth factor for angiogenesis in breast cancer.     

Expression of vascular endothelial growth factor (VEGF) and its two receptors (VEGF-R1-Flt1 and VEGF-R2-Flk1/KDR) in non-small cell lung carcinomas (NSCLCs): correlation with angiogenesis and survival

The formation of new vessels (angiogenesis) is essential for primary tumour growth and metastasis and is induced by several angiogenic factors, including vascular endothelial growth factor (VEGF). The microvascular density (MVD) in tumours was assessed and the expression of VEGF and its receptors VEGF-R1-Flt1 and VEGF-R2-KDR/Flk1 was investigated in the different cellular compartments in vivo, in order to establish their interrelationship and their prognostic influence. Immunohistochemical study of 69 stage I–II non-small cell lung carcinomas (NSCLCs) was performed on paraffin sections with CD34 antibody to estimate MVD, using a Chalkley eye-piece graticule and VEGF, VEGF-R1, and VEGF-R2 antibodies. There was strong expression of VEGF and its receptors in tumour cells, endothelial cells, and stromal fibroblasts. In tumour cells, the level of VEGF was correlated with that of VEGF-R1 ( p = 0·018) but not that of VEGF-R2. In fibroblasts, high expression of VEGF was correlated with that of VEGF-R1 ( p = 0·0001) and VEGF-R2 ( p = 0·0001). In endothelial cells, expression of VEGF was correlated with that of VEGF-R1 ( p < 0·0001) and VEGF-R2 ( p = 0·04). The level of VEGF in fibroblasts was correlated with that of VEGF-R1 ( p = 0·0028) and VEGF-R2 ( p = 0·01) in endothelial cells. There was no correlation between the level of MVD and that of VEGF or VEGF-R1 or VEGF-R2. Neither the level of MVD, nor the level of expression of VEGF and VEGF receptors in any compartment influenced the patient's survival. In conclusion, although angiogenesis is essential for tumour growth, this study failed to demonstrate that MVD, VEGF, VEGF-R1, and VEGF-R2 are prognostic markers for stage I–II NSCLC. VEGF, however, might act as a direct autocrine growth factor for tumour cells via VEGF-R1 and angiogenesis could be promoted in a paracrine loop, where VEGF is produced by fibroblasts and tumour cells and then binds to endothelial cells via induced VEGF receptors. VEGF and its receptors thus appear as relevant therapeutic targets in NSCLC. Copyright © 1999 John Wiley & Sons, Ltd.     

Prognostic value of vascularity and vascular endothelial growth factor expression in non-small cell lung cancer

AIMS: High expression of the angiogenic factor vascular endothelial growth factor (VEGF) in tumours has been found to be associated with poor prognosis in some studies, but not in others. The aims of this study were to determine the prognostic value of VEGF in operable non-small cell lung cancer (NSCLC) and its possible association with vascularity. METHODS: Sections from 81 NSCLC archival specimens were stained with antibodies to von Willebrand factor (vWF) and VEGF. Vascularity was measured by the average density of vWF positive vessels. VEGF expression in tumour cells was assessed by consensus of two independent observers according to three indices, namely: (1) percentage of area stained, (2) intensity of staining, and (3) final score (product of area and intensity). RESULTS: VEGF immunoreactivity was present in all tumours and adjacent normal lung tissue. None of the three VEGF indices was associated with vascularity or the clinical parameters examined. Mean survival times were shorter in patients with high VEGF expression, but the difference was not significant. This applied to the full cohort of patients, or when analysed separately according to tumour type or stage. However, high VEGF expression was associated with poor survival in patients with high vascularity (p = 0.02). VEGF had no discriminant value among patients with low vascularity. Vascularity had no prognostic value, except for late stage patients (UICC stages II and IIIa combined; n = 36), where high vascularity was associated with longer survival (p = 0.01). CONCLUSIONS: VEGF on its own has no prognostic value in NSCLC, but may become a useful indicator when combined with vascularity. VEGF may play a physiological role in the normal lung.     

Effect of estrogen on angiogenesis in co-cultures of human endometrial cells and microvascular endothelial cells

BACKGROUND: We recently showed that vascular endothelial growth factor (VEGF) expression by endometrial glandular epithelial and stromal cells, and endometrial microvascular endothelial cell permeability, an early step in angiogenesis, were rapidly increased by estradiol (E(2)) administration to ovariectomized baboons. We proposed that estrogen promotes endometrial angiogenesis by regulating VEGF expression by glandular epithelial and stromal cells. In the present study, we developed a co-culture of human endometrial cells and microvascular endothelial cells to determine whether the regulatory role shown for estrogen on endometrial angiogenesis in vivo in the non-human primate would be demonstrable in vitro in the human. METHODS AND RESULTS: Human endometrial glandular epithelial and stromal cells were co-cultured with human myometrial microvascular endothelial cells (HMMECs) and E(2). HMMEC tube formation (means +/- SEM, % endothelial tube area/total endothelial cell area), an index of angiogenesis, was 65% (P < 0.05) and 2-fold (P < 0.01) greater in cells co-cultured with human glandular epithelial cells (54 +/- 7%) and glandular epithelial cells plus E(2) (66 +/- 11%), respectively, compared with medium (33 +/- 4%). In contrast, endothelial tube formation was not altered in HMMECs incubated with endometrial stromal cells (32 +/- 4%), stromal cells plus E(2) (36 +/- 2%) or E(2) (39 +/- 3%). CONCLUSIONS: We propose that estrogen, by regulating expression and secretion of angiogenic factors such as VEGF by glandular epithelial cells of the endometrium, regulates endometrial angiogenesis.     

Tumor specific activation of the VEGF/KDR angiogenic pathway in a subset of locally advanced squamous cell head and neck carcinomas

Vascular endothelial growth factor (VEGF) and its receptors, Flt-1 and flk-1(KDR), constitute an important angiogenic pathway which, under hypoxic conditions, is up-regulated in many solid tumours. We used the monoclonal antibody 11B5, specific for recognizing VEGF expression and the `VEGF/flk-1(KDR) complex' on tumour endothelium, to assess free VEGF protein expression and VEGF/receptor activated microvessel density (aMVD) in a series of 104 inoperable locally advanced squamous cell carcinomas of the head and neck, treated with chemo-radiotherapy. High VEGF expression in cancer cells was strongly associated with high VEGF/receptor expression in the vasculature. The high VEGF expression and the aMVD were not associated with the standard microvessel density (sMVD), as assessed with the monoclonal antibody anti-CD31 and, were not detected in normal tissue. An increased sMVD, however, was significantly related with the expression thymidine phosphorylase (TP), and also with the nuclear accumulation of the oncoprotein p53, but neither p53 nor TP was associated with VEGF expression by cancer cells or VEGF/receptor complex aMVD. In 35% of cancer cases examined, more than 20% of the microvessels assessed with anti-CD31 also expressed the VEGF/KDR complex. The vasculature of the normal head and neck mucosa did not express the VEGF/KDR complex. There was no association between VEGF expression or VEGF/receptor complex aMVD and response to chemo-radiotherapy or patient's survival. It is concluded that activation of the angiogenic pathway VEGF/flk-1(KDR) is tumor specific in a subgroup of locally advanced squamous cell carcinomas of the head and neck. Selective destruction of this type of vasculature, using immunoconjugates directed against the VEGF/receptor complex, may prove therapeutically useful for patients with a high tumoral VEGF/flk-1(KDR) activated microvessel fraction. This revised version was published online in July 2006 with corrections to the Cover Date.     

Expression of vascular endothelial growth factor in renal cell carcinomas

Vascular endothelial growth factor (VEGF) is an angiogenic factor that may be involved in tumor growth and metastasis. Only a few data concerning the role of VEGF in renal cell carcinomas (RCCs) are available, and no studies have yet evaluated its prognostic value. The aim of the present study was to assess VEGF expression in a large series of renal tumors with a long follow-up, correlated with the usual histoprognostic factors and survival. VEGF immunostaining was performed on formalin-fixed, paraffin-embedded archival tissue from 74 renal carcinomas (62 conventional renal cell and 12 papillary carcinomas). Positivity of immunostaining was semi-quantitatively scored by two pathologists. Angiogenesis was evaluated by immunostaining with anti-CD34 antibodies on serial sections. Cytoplasmic VEGF expression was detected in tumor cells in 35% (26/74) of RCCs, including 18 out of the 62 (29%) conventional RCCs and 8 out of the 12 (67%) papillary carcinomas (P=0.02). In the group of conventional RCCs, VEGF expression was positively correlated with both nuclear grade (P=0.05) and size of the tumor (P=0.05). Furthermore, a significant correlation was observed between VEGF expression and microvascular count (P=0.04). Finally, cumulative survival rate was significantly lower in the group of patients with conventional RCCs expressing VEGF (log rank test, P=0.01). In the Cox model, VEGF expression was a significant independent predictor of outcome, as well as stage and nuclear grade. This study suggests that VEGF is involved in angiogenesis in conventional RCCs and appears to be a potential prognostic factor in these tumors. Received: 9 July 1999 / Accepted: 18 October 1999     

Clinical prognostic values of vascular endothelial growth factor, microvessel density,and p53 expression in esophageal carcinomas

Vascular endothelial growth factor (VEGF) is known to play a key role in tumor angiogenesis. The tumor-suppressor gene p53 has been thought to regulate VEGF. We investigated the effect of VEGF on esophageal carcinoma and the correlation between VEGF and p53. Tissue samples were taken from 81 patients with esophageal carcinoma after surgery. VEGF and p53 expressions were examined by immunohistochemical staining. Microvessels in the tumor stained for CD34 antigen were also counted. VEGF and p53 expressions were observed in 51.3% (41/80) and 51.9% (41/79), respectively. The microvessel density was 70.9+/-6.7 (mean+/-SE) in VEGF-positive group and 68.7+/-5.1 in VEGF-negative group. However, no correlation was noted between VEGF and p53 expression. Whereas the tumor size, nodal status, depth of invasions, and tumor stage were associated with poor overall survival, VEGF expression or p53 expression was not. These results indicate that VEGF and p53 are highly expressed in esophageal carcinomas. Since the VEGF expression is not correlated with the p53 expression, microvessel density or clinicopathological findings, further studies with other angiogenic molecules are needed to determine the role in esophageal carcinomas.     

Increased expression of hypoxia-inducible factor 1alpha in type I and type II endometrial carcinomas.

Hypoxia-inducible factor 1alpha (HIF-1alpha) is a nuclear protein that is upregulated in many tumors and triggers biologic events intimately associated with aggressive tumor behavior. The aim of this study was to analyze the expression of HIF-1alpha, vascular endothelial growth factor (VEGF), Ki-67 and p53 in type I and type II endometrial adenocarcinoma. In total, 149 patients diagnosed with endometrial adenocarcinoma in our institute from 1995 to 2001 were included in this study, of which 108 were type I and 41 were type II endometrial adenocarcinoma. Patient demographics, clinical and pathological data were reviewed. Tissue microarrays were prepared from the paraffin blocks and immunohistochemistry was performed for antibodies against HIF-1alpha, VEGF, Ki-67 and p53. High expression of HIF-1alpha, VEGF, Ki-67 and p53 were significantly more frequent in type II than type I endometrial adenocarcinoma (P<0.001). HIF-1alpha expression was highly correlated with VEGF expression in the tumor cells (P=0.001). In type I endometrial adenocarcinoma, high expression of HIF-1alpha showed a significant correlation with higher grade of the tumor, depth of myometrial invasion, adnexal invasion and clinical stage. A similar correlation was not observed in type II endometrial adenocarcinoma. Surgical stage was the only independent prognostic marker for survival. In conclusion, high expression of HIF-1alpha is more frequent in type II than in type I endometrial adenocarcinoma. In type I endometrial adenocarcinoma, HIF-1alpha expression correlates with morphologic features of aggressiveness. In type II endometrial adenocarcinoma, there is no correlation between HIF-1alpha expression and these features. Thus, HIF-1alpha may play an important role in endometrial adenocarcinoma progression, particularly in type I endometrial adenocarcinoma. Additional investigations of HIF-1alpha as a biomarker of aggressive potential and as a novel target for therapeutics in endometrial adenocarcinoma are warranted.     

Expression of angiogenic factors and apoptotic factors in leiomyosarcoma and leiomyoma.

Angiogenesis is essential for tumor growth and metastasis. Some angiogenic factors, such as vascular endothelial growth factor (VEGF), platelet-derived endothelial cell growth factor (PD-ECGF), transforming growth factor-alpha (TGF-alpha) and basic fibroblast growth factor (bFGF) are involved in increased angiogenic activity and disease progression in many carcinomas. However, there is little information regarding the association between angiogenic factors and leiomyosarcoma. Although there are abundant vessels in the sarcoma which enable it to easily receive nutrition and medicinal components, chemotherapy cannot effectively treat leiomyosarcoma. This means the resistance to anticancer drugs in leiomyosarcoma is very strong. However, the resistant mechanism is still unclear. In this study, expressions of VEGF, PD-ECGF, TGF-alpha, bFGF, intratumoral microvessel density (IMVD), and p53, Bcl-2 and Bax were examined by immunohistochemistry in 30 patients with leiomyosarcoma and 21 patients with leiomyoma. With regard to angiogenesis, PD-ECGF and TGF-alpha were closely associated with an increase in IMVD (p=0.012, 0.0196, respectively), and VEGF and PD-ECGF were significantly expressed in leiomyosarcoma compared with leiomyoma (p=0.041, 0.041, respectively). Although p53 expression in leiomyosarcoma was significantly higher than in leiomyoma (p=0.016), the frequency of p53 positivity was not so high (47%). On the other hand, the ratio of Bcl-2/Bax in leiomyosarcoma was significantly higher than that in leiomyoma (p=0.033). The findings of this study suggest that in leiomyosarcoma, angiogenic factors, such as PD-ECGF, VEGF and TGF-alpha expression may be involved in tumor angiogenesis, and the frequently high ratio of Bcl-2/Bax and expression of p53 gene mutation might be related to chemoresistance mechanism.     

bcl-2 and c-erbB-2 proteins are involved in the regulation of VEGF and of thymidine phosphorylase angiogenic activity in non-small-cell lung cancer

Tumour angiogenesis has been recently recognised as one of the most important prognostic factors in lung cancer. Although a variety of angiogenic factors have been identified, the angiogenesis process remains poorly understood. Bcl-2, c-erbB-2 and p53 are well-known oncogenes involved in non-small-cell lung cancer pathogenesis. A direct correlation of thymidine phosphorylase (TP) and of vascular endothelial growth factor (VEGF) with intratumoural angiogenesis has been reported. In the present study we investigated the possible regulatory role if bcl-2, c-erB-2 proteins in angiogenesis and in VEGF and TP expression in non-small-cell lung cancer. Two hundred sixteen specimens from T1,2-N0,1 staged patients treated with surgery alone were immunohistochemically examined. Bcl-2 and c-erbB-2 were significantly inversely related to each other (P=0.04) and both were inversely associated with microvessel density (P<0.02). High TP and VEGF reactivity was statistically related to loss of bcl-2 expression (P<0.01). A significant co-expression of c-erbB-2 with TP was noted (P=0.01). However, TP expression was related to high angiogenesis only in cases with absence of c-erB-2 expression (P<0.0001). c-erbB-2 expression in poorly vascularised tumours was linked with poor outcome (P=0.03). The present study provides strong evidence that the bcl-2 gene has a suppressive function over genes involved in both angiogenesis (VEGF and TP) and cell migration (c-erbB-2) in NSCLC. TP and c-erbB-2 proteins are significantly, and often simultaneously, expressed in bcl-2 negative cases. However, expression of the c-erbB-2 abolishes the TP-related angiogenic activity. Whether this is a result of a direct activity of the c-erbB-2 protein or a consequence of a c-erbB-2-related immune response remains to be further investigated. This revised version was published online in July 2006 with corrections to the Cover Date.     

Thymidine phosphorylase expression in gallbladder adenocarcinomas

The expression of thymidine phosphorylase (TP), a potent chemotactic factor for endothelial cells, was studied in 60 adenocarcinomas of the gallbladder, by use of immunohistochemical techniques. Results on patterns of TP expression were correlated with angiogenesis (anti-CD31), histopathological variables, and patient survival. TP was frequently expressed in tumor cells, stromal cells, tumor-associated macrophages, and lymphocytes of gallbladder adenocarcinomas. The expression was mixed nuclear/cytoplasmic. However, only nuclear TP (TPnuc) expression by tumor cells was correlated with increased angiogenic activity. High angiogenesis, assessed as microvessel density (MVD), was the most significant prognostic factor. The subgroup of patients with TPnuc and medium/high MVD had the worst prognosis as evaluated by the survival curves. Furthermore, CD31+ lymphocytes, frequently seen in carcinomas with high-fibroblastic TP reactivity, were connected with an improved survival. It is concluded that angiogenesis, as verified by multivariate analysis, is the most important prognostic factor in gallbladder carcinomas. In these tumors, high histologic grade and low CD31+ lymphocytic infiltration are also independent predictors of poor prognosis. TP is associated with an aggressive phenotype apparently because of its anglogenic activity. Therapeutic strategies targeting TP may be of value in patients overexpressing this enzyme.     

Prognostic impact of VEGF, CD31, CD34, and CD105 expression and tumour vessel invasion after radical surgery for IB-IIA non-small cell lung cancer

AIMS: To evaluate the prognostic impact of tumour angiogenesis assessed by vascular endothelial growth factor (VEGF), microvessel density (MVD), and tumour vessel invasion in patients who had undergone radical resection for stage IB-IIA non-small cell lung cancer (NSCLC). METHODS: Fifty one patients (42 men, nine women; mean age, 62.3 years; SD, 6.9) undergoing complete surgical resection (35 lobectomy, 16 pneumonectomy) of pathological stage IB (n = 43) and IIA (n = 8) NSCLC were evaluated retrospectively. No patient underwent postoperative chemotherapy or neoadjuvant treatment. Tumour specimens were stained for VEGF and specific MVD markers: CD31, CD34, and CD105. RESULTS: VEGF expression significantly correlated with high CD105 expression (p < 0.0001) and tumour vessel invasion (p = 0.04). Univariate analysis showed that those patients with VEGF overexpression (p = 0.0029), high MVD by CD34 (p = 0.0081), high MVD by CD105 (p = 0.0261), and tumour vessel invasion (p = 0.0245) have a shorter overall survival. Furthermore, multivariate Cox regression analysis showed that MVD by CD34 (p = 0.007), tumour vessel invasion (p = 0.024), and VEGF expression (p = 0.042) were significant predictive factors for overall survival. Finally, the presence of both risk factors, tumour vessel invasion and MVD by CD34, was highly predictive of poor outcome (odds ratio, 3.4; 95% confidence interval, 1.7 to 6.5; p = 0.0002). CONCLUSIONS: High MVD by CD34 and tumour vessel invasion are more closely related to poor survival than the other neoangiogenetic factors in stage IB-IIA NSCLC. This may be because these factors are more closely related to the metastatic process.     

VEGF-B expression in human primary breast cancers is associated with lymph node metastasis but not angiogenesis

Angiogenesis is essential for tumour growth and metastasis. It is regulated by numerous angiogenic factors, one of the most important being vascular endothelial growth factor (VEGF). Recently VEGF-B, a new VEGF family member that binds to the tyrosine kinase receptor flt-1, has been identified. Although the importance of VEGF has been shown in many human tumour types, the contribution of VEGF-B to tumour neovascularization is unknown in any tumour type. This study therefore measured the mRNA level of VEGF-B and its receptor flt-1 by ribonuclease protection assay and the pattern of VEGF-B expression by immunohistochemistry in 13 normal breast samples and 68 invasive breast cancers. Flt-1 expression was significantly higher in tumours than in normal breast (p=0.02) but no significant difference was seen in VEGF-B between normal and neoplastic breast (p=0.3). There was a significant association between VEGF-B and node status (p=0.02) and the number of involved nodes (p=0.01), but not with age (p=0.7), size (p=0.6), oestrogen receptor (ER) (p=0.2), grade (p=0.5) or vascular invasion (p=0.16). No significant relationship was present between VEGF-B and flt-1 (p=0.2) or tumour vascularity (p=0.4). VEGF-B was expressed mostly in the cytoplasm of tumour cells, although occasional stromal components including fibroblasts and endothelial cells were also positive. No difference in VEGF-B expression was observed adjacent to regions of necrosis, in keeping with this VEGF family member not being hypoxically regulated. These findings suggest that VEGF-B may contribute to tumour progression by a non-angiogenic mechanism, possibly by increasing plasminogen activators and hence metastasis, as has been described in vitro. Measurement of VEGF-B together with other angiogenic factors may identify a poor prognostic patient group, which may benefit from anti-VEGF receptor therapy targeted to flt-1 (VEGFR1) as well as kdr (VEGFR2).     

Quantification of tumour and circulating vascular endothelial growth factor (VEGF) in patients with oesophagogastric cancer: a long-term follow-up study

Vascular endothelial growth factor (VEGF) is an angiogenic cytokine that regulates tumour angiogenesis. The prognostic significance of VEGF expression remains incompletely investigated for patients with oesophagogastric cancer. This study assesses the significance of tumour VEGF (T-VEGF) and circulating VEGF (C-VEGF) expression in a 10-year follow-up of patients with oesophagogastric cancer. Patients undergoing surgical resection were prospectively recruited between February 1999 and August 2000. Circulating VEGF, derived both from plasma (P-VEGF) and serum (S-VEGF), and T-VEGF were assessed using a commercial enzyme-linked immunosorbent assay (ELISA). As platelet count may contribute to C-VEGF, pre-operative platelet levels were also recorded to exclude a confounding effect. Patients were followed up over a 10-year period using the Northern Ireland Cancer Registry. Sixty-one patients were recruited (men=45) with a mean age of 65.7 years. The 10-year survival was 19.7% (n=12) with a median follow-up of 808 days (inter-quartile range [IQR]: 349.5-2358.5). Union for International Cancer Control (UICC) tumour staging was Stage I=9 (14.8%), Stage II=15 (24.6%), Stage III=33 (54.1%) and Stage IV=4 (6.6%). The only significant relationship between clinicopathological features and the study variables was for S-VEGF, which was elevated in patients with advanced T-stage (P = 0.05). Circulating VEGF did not correlate with platelet count. Although a trend towards decreased survival was observed for patients who had positive lymph nodes (P = 0.08) and advanced UICC stage (P = 0.09) on univariate analysis, only lymphovascular invasion significantly predicted poor prognosis in this cohort (P = 0.05). Therefore, ELISA quantification of circulatory or tumour VEGF does not appear to be a significant predictor of mortality in patients with oesophagogastric cancer.     

Sialosyl-Tn expression in normal and pathological conditions of human endometrium. An immunohistochemical study

To assess the clinico-prognostic relevance of the cell surface carbohydrate glycoprotein in normal and pathological conditions of human endometrium, Sialosyl-Tn (STn) antigen was immunohistochemically studied in normal (n = 10), hyperplastic (n = 18), and neoplastic (n = 60) endometrial lesions. There was no STn antigen reactivity in the proliferative endometrial slides, while weak staining was observed in all secretory endometria. STn expression was noted in 8/18 (44%) hyperplastic endometrial cases and in 40/60 (67%) endometrial carcinomas. Positive staining was observed throughout the cytoplasm of the glandular cancer cells, at the cell membranes, and in an intraluminar mucus. This antigen was mostly expressed heterogeneously as far as the distribution of positive cells is concerned. There was a statistically significant association between STn expression and the histological grading of cancer (p = 0.019). Advanced clinical stage (III-IV; p = 0.014) and infiltration of the myometrial wall (more than 1/2 of the myometrial wall; p = 0.004), but no STn immunoreactivity, were reported to be independent prognostic variables during follow-up. Our study shows that a) STn is not constantly expressed during the menstrual cycle, and is increased at the secretory phase of the cycle; b) Sialosyl-Tn reactivity decreases with the degree of tumor differentiation, but there was no relationship with other clinicopathological variables of cancer; c) this cell surface carbohydrate glycoprotein does not appear to predict the outcome of endometrial cancer patients.