The value of platelet-derived endothelial cell growth factor as a novel predictor of advancement of uterine cervical cancers

Serum platelet-derived endothelial cell growth factor (PD-ECGF) in patients with uterine cervical cancers revealed a significantly positive correlation with clinical stage and tumor size and with the advancement indicators lymph node metastasis, parametrial involvement, and vessel permeation in both squamous cell carcinomas and adenocarcinomas. The prognosis of the patients with high serum PD-ECGF was extremely poor, whereas the 36-month survival rate of the other patients with low serum PD-ECGF was 81.3% in squamous cell carcinomas and 80.0% in adenocarcinomas. Our data indicate that serum PD-ECGF levels reflect the status of advancement of uterine cervical cancers and thus may be recognized as a novel tumor marker for both squamous cell carcinomas and adenocarcinomas of the uterine cervix.     

Prediction of the effect of 5'-deoxy-5-fluorouridine by the status of angiogenic enzyme thymidine phosphorylase expression in recurrent breast cancer patients

The anti-cancer drug, 5'-deoxy-5-fluorouridine (5'DFUR) is known to have antitumor activity through the conversion to 5FU by thymidine phosphorylase (TP). Recently, TP was demonstrated to be identical to angiogenic molecule platelet-derived endothelial cell growth factor (PD-ECGF) by cDNA cloning and subsequent biochemical analyses. We have examined the relationship between the clinical response of 5'DFUR and TP/PD-ECGF expression determined by immunocytochemical analysis in 24 recurrent breast cancer patients. Of 24, 13 were TP/PD-ECGF positive and 11 were TP/PD-ECGF negative. In 13 TP/PD-ECGF positive patients, 4 showed objective response (OR) and 3 showed stable disease (SD) with 5'DFUR treatment, however only one case showed OR and no case showed SD in 11 TP/PD-ECGF negative patients, suggesting that 5'DFUR was likely to be effective for TP/PD-ECGF positive patients. In another group of recurrent breast cancer patients treated by adriamycin containing regimen, no significant correlation was observed between the response of 5'DFUR and the status of TP/PD-ECGF expression. It was indicated that an angiogenic enzyme TP/PD-ECGF expression might be a predictor of the effect of 5'DFUR treatment in human breast cancer.     

Expression of platelet-derived endothelial cell growth factor/thymidine phosphorylase in cervical intraepithelial neoplasia

Angiogenesis contributes to the growth and secondary spreading of solid tumors. Platelet-derived endothelial cell growth factor (PD-ECGF)/thymidine phosphorylase (TP) has been identified as such an angiogenic factor. In this study, the expression of PD-ECGF/TP and VEGF was evaluated by immunohistochemical staining of tumor specimens from 40 patients with cervical intraepithelial neoplasia (10 with moderate dysplasia; 10 with severe dysplasia; 10 with carcinoma in situ; 10 with invasive carcinoma). The microvessel density was assessed by immunostaining for factor VIII-related antigen in the most highly neovascularized area. In both the nucleus and cytoplasm, the intensity of PD-ECGF/TP expression in carcinoma in situ and invasive carcinoma was significantly stronger than that in moderate dysplasia. However, the intensity of VEGF expression was not significantly different in the various specimens. The microvessel density in mild dysplasia was significantly different from that in carcinoma in situ (p<0.05), and that in invasive carcinoma (p<0.05). There was no significant relationship between the microvessel density and the expression of PD-ECGF/TP or that of VEGF. These results show that the expression of PD-ECGF/TP appears to be involved in the promotion of angiogenesis in cervical intraepithelial neoplasia.     

Angiogenic switch occurs during the precancerous stage of human esophageal squamous cell carcinoma

We previously reported that vascular endothelial growth factor (VEGF) expression correlates with vessel density in human esophageal squamous cell carcinomas. However, tumor angiogenesis is not controlled simply by the presence of VEGF, and is likely regulated by several angiogenic factors produced by tumor and host cells. The goal of the present study was to determine the angiogenic profile of precancerous and cancerous lesions of the esophagus. Expression of mRNAs for VEGF, platelet derived endothelial cell growth factor (PD-ECGF), basic fibroblast growth factor (bFGF), and interleukin (IL)-8 was examined in six esophageal carcinoma cell lines and fresh biopsy specimens from 16 patients with invasive esophageal carcinoma by RT-PCR. Immunohistochemical analyses with antibodies against VEGF, PD-ECGF, bFGF, and IL-8 were performed on archival specimens of 60 normal esophageal mucosa, 11 dysplasias and 49 carcinomas of the esophagus. Microvessels were stained with anti-CD34 antibody and quantified by counting the number of vessels in a x200 field in the most vascularized areas of the tumor. Esophageal carcinoma cell lines and tumor tissues expressed mRNAs for one or more these angiogenic factors at various levels. An initial increase in vessel density and enhanced expression of PD-ECGF and VEGF were observed in dysplastic epithelium. Vessel density was significantly higher in more advanced lesions. bFGF and IL-8 were not expressed in dysplasias and mucosal carcinomas, but expression was increased in late stage squamous cell carcinoma. These findings suggest that the angiogenic switch is a very early event in the development of invasive carcinoma. Several different angiogenic factors produced by tumor cells and host cells may regulate angiogenesis during different steps of esophageal carcinogenesis.     

Expression of basic fibroblast growth factor is associated with a good outcome in patients with squamous cell carcinoma of the esophagus

Angiogenesis is an essential step in tumor growth and metastasis, but rather than being controlled by means of a simple mechanism, the control of tumor angiogenesis may be mediated by several angiogenic factors. We investigated the expression of basic fibroblast growth factor (b-FGF) and platelet-derived endothelial cell growth factor (PD-ECGF) in squamous cell carcinoma of the esophagus in order to clarify the mechanism of angiogenesis. Expression of b-FGF and PD-ECGF was immunohistochemically investigated in tissue specimens from the tumors of 79 patients with squamous cell carcinoma of the esophagus who underwent curative esophagectomy without preoperative chemotherapy or radiation therapy, and the relationship between expression of b-FGF/PD-ECGF, microvessel density (MVD), and clinicopathological background factors was assessed. Tumor cells that expressed b-FGF were found in 41 patients (51.9%), and tumor cells that expressed PD-ECGF were found in 57 patients (72.2%). Although the mean vascular density (47.9/mm(2)) of b-FGF-positive tumors was significantly lower than that (67.2/mm(2)) of b-FGF-negative tumors (p=0.014), the difference between the 56.0/mm2 in PD-ECGF-positive tumors and 60.3/mm2 in PD-ECGF-negative tumors was not significant. Although the survival rate of patients with b-FGF-positive tumors was significantly higher than those with b-FGF-negative tumors (p=0.033), there was no significant difference between the survival rates of patients with PD-ECGF-positive and -negative tumors (p=0.580). Expression of b-FGF may be associated with promotion of angiogenesis and a good prognostic factor in squamous cell carcinoma of the esophagus.     

TGF-alpha as well as VEGF, PD-ECGF and bFGF contribute to angiogenesis of esophageal squamous cell carcinoma

It has been demonstrated that vascular endothelial growth factor (VEGF) is associated with tumor progression as an angiogenic factor in esophageal squamous cell carcinoma (SCC)s. However, the role of other angiogenic factors such as transforming growth factor-alpha (TGF-alpha), platelet-derived endothelial cell growth factor (PD-ECGF), and basic fibroblast growth factor (bFGF) are still unknown in esophageal SCCs. In this study, we detected the expression of VEGF, TGF-alpha, PD-ECGF and bFGF in tissue specimens from 96 patients with SCC of the esophagus by immunohistochemical staining. To evaluate angiogenesis, endothelial cells were stained immunohistochemically and microvessel density (MVD) was counted in 24 cases. The positive rates for VEGF, TGF-alpha, PD-ECGF and bFGF were 65% (62/96), 67% (64/96), 66% (63/96), and 49% (47/96), respectively. Only TGF-alpha expression had a strong correlation with the average MVD (p=0.0059). However, the MVD increased as the number of positive factors for these 4 factors increased (p=0.0023). The expression of all of these factors significantly correlated to the depth of tumor invasion, and lymph node metastasis. Finally, survival analysis of the patients revealed that VEGF, TGF-alpha, and PD-ECGF were significant prognostic factors. However, multivariate analysis revealed that these factors were not prognostic. Thus, we suggest that TGF-alpha as well as VEGF, PD-ECGF and bFGF may be associated with angiogenesis, and the progression and metastasis of esophageal squamous cell carcinoma.     

Expression of platelet-derived endothelial cell growth factor (PD-ECGF) and its mRNA in uterine endometrial cancers

To determine the potential of growth, invasion and metastasis of uterine endometrial cancer cells associated with neovascularization, the expressions of platelet-derived endothelial cell growth factor (PD-ECGF) and its mRNA in uterine endometrial cancers and in normal uterine endometria as controls were determined and the relationship between their expressions and histological grades, grades of myometrial invasion and clinical stages of uterine endometrial cancers was analyzed. The levels of PD-ECGF were significantly higher in uterine endometrial cancers of well-differentiated grade (G1) with invasion to ≤1/2 myometrium (B) and of stage I than in those of moderately and poorly differentiated grades (G2 and G3, respectively) limited to endometrium (A) and with invasion to >1/2 myometrium (C) and of stages II and III/IV and in normal uterine endometria. There was no significant difference in the levels between uterine endometrial cancers of G2 and G3, A and C, or stages II and III/IV and normal uterine endometria. Therefore, the active availability of PD-ECGF might contribute to the acceleration of angiogenic activity in the early process of invasion of well-differentiated uterine endometrial cancers.     

Hypoxia correlates with angiogenesis in cervical cancers

Background Tissue hypoxia stimulates the induction of the angiogenic substances vascular endothelial growth factor and erythropoietin in the locus of the tissue. We have previously demonstrated that erythropoietin promotes angiogenesis by binding to its receptor in the endothelial cells of uterine and ovarian malignancies. In the present study, we examined whether malignant uterine cervix tissue showed hypoxia and whether hypoxia correlated with high vascular density through vascular endothelial growth factor.Methods To detect tissue hypoxia, we estimated the content of ATP in squamous cell carcinoma of the uterine cervix and in the normal cervix, using liquid chromatography columns. Surgically resected samples were fixed in Zamboni solution and processed for immunohistochemical microscopy to identify the endothelial cells and the location of vascular endothelial growth factor, with the use of anti-factor VIII and anti-vascular endothelial growth factor₁₆₅ antibody, respectively. The microvessels in a definite area were counted in sections of each specimen.Results Significantly lower ATP levels and significantly higher vascular density were seen in squamous cell carcinoma than in the controls (P < 0.05). The microvessel number in relation to ATP content was significantly higher in squamous cell carcinoma than in the controls (P < 0.001). Moreover vascular endothelial growth factor, the hyperplastic epithelium of the squamous cell carcinoma contained the immunoreactivity, with characteristic histopathological features suggesting retention of tissue fluid.Conclusion Squamous cell carcinoma of the uterine cervix showed hypoxia which correlated with abundant vascularity. Vascular endothelial growth factor expressed in the hyperplastic epithelium appears to promote angiogenesis in squamous cell carcinoma of the uterine cervix.     

Clinical implications of expression of ETS-1 related to angiogenesis in uterine endometrial cancers.

BACKGROUND: Angiogenesis is essential for development, growth and advancement of solid tumors. During angiogenesis, ETS-1 is strongly expressed in vascular endothelial cells and the adjacent interstitial cells, while the inhibition of ETS-1 expression leads to suppression of angiogenesis. This prompted us to study the clinical implications of ETS-1 in relation to angiogenesis in uterine endometrial cancers. PATIENTS AND METHODS: Sixty patients underwent resection for uterine endometrial cancers. From the tissues of 60 uterine endometrial cancers, the levels of ets-1 mRNA, vascular endothelial growth factor (VEGF), basic fibroblast growth factor (bFGF), platelet-derived endothelial cell growth factor (PD-ECGF) and interleukin (IL)-8 were determined by competitive RT-PCR using recombinant RNA and enzyme immunoassay, and the localization and counts of microvessel were determined by immunohistochemistry. RESULTS: There was a significant correlation between microvessel count and ets-1 gene expression levels in uterine endometrial cancers. Immunohistochemical staining revealed that the localization of ETS-1 was similar to that of vascular endothelial cells. The level of ets-1 mRNA tended to increase with increasing disease stage. Furthermore, the level of ets-1 mRNA correlated with levels of VEGF in well-differentiated adenocarcinomas (G1) and of bFGF in moderately differentiated adenocarcinomas (G2) and poorly differentiated adenocarcinomas (G3). CONCLUSIONS: ETS-1 is a possible angiogenic mediator in uterine endometrial cancers.     

Angiogenesis and platelet-derived endothelial cell growth factor/thymidine phosphorylase expression in cervical cancer.

The object of this study was to clarify the association of platelet-derived endothelial cell growth factor (PD-ECGF)/thymidine phosphorylase (dThdPase), separately assessed in cancer cells and in stroma cells, with clinicopathological factors including tumor angiogenesis and prognosis in cervical cancer. The expression of PD-ECGF was evaluated by immunohistochemical staining in 92 patients with stage Ib-II cervical cancer. The microvessel count was assessed by immunostaining for factor VIII-related antigen in the most neovascularized area. Microvessel count was significantly higher in tumors with non-squamous cell carcinoma. PD-ECGF expression in cancer cells was significantly higher in tumors with pelvic node metastasis and squamous cell carcinoma. Immunopositivity for PD-ECGF in stroma cells was significantly higher in tumors with large size and deep stromal invasion. The microvessel counts in cases with positive PD-ECGF expression in stroma cells were significantly higher than those in cases with negative PD-ECGF expression in stroma cells (p=0.048). Disease-free survival and overall survival were significantly worse in patients with deep stromal invasion, parametrial involvement, vaginal involvement, lymph-vascular space involvement, pelvic lymph node metastasis and high microvessel count. A multivariate analysis using Cox's proportional hazard model showed that high microvessel count independently predicted disease-free and overall survival. The expression of PD-ECGF in stroma cells may play a crucial role in the promotion of angiogenesis and tumor angiogenesis can be used as a useful prognostic marker for cervical cancer.     

Pd-ECGF positivity correlates with better survival,while iNOS has no predictive value for cervical carcinomas treated with radiotherapy

PURPOSE: Platelet-derived endothelial cell growth factor (PD-ECGF), which has angiogenic activity, is identical to thymidine phosphorylase. Tumor vascularization is considered to be an important prognostic factor. Nitric oxide synthases (NOSs) are a kind of enzyme that generates nitric oxide. Nitric oxide has not only a self defense against neoplastic cells but also tumor growth stimulation by promoting new blood vessel formation. Our purpose was to investigate the correlation between the expression of PD-ECGF or inducible NOS (iNOS) in cancer cells and prognosis. METHODS AND MATERIALS: Formaldehyde-fixed and paraffin-embedded biopsy specimens excised from 71 cervical squamous cell carcinoma patients who were treated with radiotherapy alone were investigated using an immunohistochemical method. RESULTS: Cancer cells that were positive for PD-ECGF showed intranuclear and cytoplasmic staining patterns. Of the 71 patients, 40 (56%) were positive for PD-ECGF and 31 (44%) were negative. The 5-year survival of the PD-ECGF-positive patients was significantly better than that of the PD-ECGF-negative patients (p = 0.026). Cancer cells that were positive for iNOS showed a cytoplasmic staining pattern. Twenty-seven patients (38%) were positive for iNOS and 44 (62%) were negative. No significant prognostic correlation was observed between iNOS-positive and iNOS-negative patients. CONCLUSION: PD-ECGF positivity in cancer cells is a predictive factor for a good prognosis in cervical squamous cell carcinoma treated with radiotherapy alone.     

宫颈鳞癌组织中PD-ECGF C-erbB-2 Cath-D的表达及临床意义

目的:探讨PD-ECGF、C-erbB-2和Cath-D三种蛋白在宫颈鳞癌中表达的临床意义及其相关性.方法:应用免疫组化法检测87例宫颈鳞癌组织及23例对照组中PD-ECGF、C-erbB-2和Cath-D的表达,采用x2检验,分析三种蛋白表达与临床病理参数的关系.结果:1)PD-ECGF在宫颈鳞癌中表达率高且与临床分期及组织分化程度有关;2)C-erbB-2在宫颈鳞癌中的表达率和对照组比较,差异有显著性(P＜0.01);表达率与临床分期、组织分化程度有显著相关性(P＜0.05);3)Cath-D的阳性表达率高于对照组,其表达与宫颈鳞癌组织分化程度、淋巴结转移相关;4)C-erbB-2与Cath-D之间阳性表达有显著相关性(P＜0.05).结论:PD-ECGF、C-erbB-2及Cath-D可作为判断宫颈鳞癌生物学行为的重要指标.     

Expression of IP-10 related to angiogenesis in uterine cervical cancers

Angiogenesis is essential for development, growth and advancement of solid tumours. Interferon-gamma-inducible protein 10 (IP-10) regulates lymphocyte chemotaxis, mediates vascular pericyte proliferation and acts as an angiostatic agent, thus inhibiting tumour growth. This prompted us to study the clinical implications of IP-10 expression related to angiogenesis in uterine cervical cancers. The levels of IP-10 decreased with advancement, and the prognosis of the 30 patients with low IP-10 expression in uterine cervical cancers was poor (66%), whereas the 24-month survival rate of the other patients with high IP-10 expression was 90%. Furthermore, IP-10 levels significantly reverse-correlated with vascular endothelial growth factor (VEGF) levels in uterine cervical cancers. Interferon-gamma-inducible protein 10 might work on suppression of angiogenesis associated with VEGF in advancement, and can be recognised as a prognostic indicator. Furthermore, IP-10 activation might be effective on the suppression of regrowth or recurrence after intensive treatment for advanced cervical cancers.     

Clinical implications of expression of interleukin-8 related to myometrial invasion with angiogenesis in uterine endometrial cancers.

BACKGROUND: Angiogenesis is essential for development, growth and advancement of solid tumors. The tumor-associated macrophage has been recognized among inflammatory cells as a candidate for supplying tumor angiogenic factors. Interleukin (IL)-8 is assumed to be a macrophage-derived mediator of angiogenesis. This prompted us to study the clinical implications of macrophage-derived angiogenesis in uterine endometrial cancers. PATIENTS AND METHODS: Sixty patients underwent curative resection for uterine endometrial cancers. The patient prognosis was analyzed with a 48 month survival rate after curative resection. In tissue of uterine endometrial cancers, the levels of IL-1alpha, IL-1beta, tumor necrosis factor-alpha, IL-8, basic fibroblast growth factor, vascular endothelial growth factor and platelet-derived endothelial cell growth factor were determined by enzyme immunoassay, and the localization and counts of microvessels and macrophages were determined by immunohistochemistry. RESULTS: There was a significant correlation between microvessel counts and IL-8 levels and between infiltrated macrophage counts and IL-8 levels in uterine endometrial cancers. Immunohistochemical staining revealed that the localization of IL-8 was similar to that of CD68 for macrophages. IL-8 levels were significantly increased during myometrial invasion from stage Ia to stages Ib through IV. CONCLUSIONS: IL-8 might act as an angiogenic switch in myometrial invasion in stage I uterine endometrial cancers. Furthermore, IL-8 supplied from infiltrated macrophages within and around the tumor might not be a prognostic indicator of advancement, but may be associated with myometrial invasion in uterine endometrial cancers.     

The expression of platelet-derived endothelial cell growth factor/thymidine phosphorylase associates with angiogenesis in epithelial ovarian cancer

Background The object of this study is to clarify the association of an angiogenic factor, PD-ECGF (platelet-derived endothelial cell growth factor/thymidine phosphorylase), with clinicopathologic factors, in this case tumor angiogenesis, in epithelial ovarian cancers. Methods Tumor specimens were obtained at the time of surgery from the primary lesion in 60 patients with epithelial ovarian cancer. Histologic cell types were assigned to tumors according to the World Health Organization classification: 26 were classified as serous adenocarcinoma, 15 as endometrioid adenocarcinoma, 9 as mucinous adenocarcinoma, 9 as clear cell carcinoma, and 1 as undifferentiated carcinoma. Surgical staging was based on the international Federation of Gynecology and Obstetrics (FIGO) staging system: 16 were stage I, 6 were stage II, 34 were stage III, and 4 were stage IV. Expression of PD-ECGF was evaluated by immunohistochemical staining. Microvessel density was assessed by immunostaining for factor VIII-related antigen in the most neovascularized area. Results Stroma cells stained more strongly than cancer cells (80% vs. 33%). The immunopositivity of PD-ECGF in stroma cells was higher in cases of advanced cancer. Expression of PD-ECGF in mucinous adenocarcinomas was significantly higher than that in serous adenocarcinomas, while PD-ECGF expression in clear cell carcinomas was significantly lower. The microvessel density in the cases with marked PD-ECGF-positive stroma cells was significantly higher than that in the cases with absent/minimal PD-ECGF-positive stroma cells (P<0.05). Conclusion The expression of PD-ECGF may play a crucial role in the promotion of angiogenesis in epithelial ovarian cancers.     

Influence of platelet-derived endothelial cell growth factor/thymidine phosphorylase on the cell cycle in head and neck squamous cell carcinoma in vitro

Platelet-derived endothelial cell growth factor (PD-ECGF) was isolated as an endothelial mitogen from platelets and demonstrated to have angiogenic activity and thymidine phosphorylase (TP) activity. It was reported that the overexpression of PD-ECGF occurred with the rapid tumor growth in vivo. In this study, we transfected PD-ECGF into the head and neck squamous cell carcinoma cell line IMC-3 and investigated the property of transfectants in vitro. Highly overexpressed PD-ECGF transfectants rapidly grew compared with parental cells and control vector (CV) transfectants (p<0.05). The expression of cyclin D1 and cyclin E were more enhanced in PD-ECGF transfectants than parental cells and CV transfectants, while the p27kip1 was inhibited in PD-ECGF transfectants. In PD-ECGF transfectants, S and G2/M-phase cells rapidly increased compared with parental cells and CV transfectants. These results showed that the cancer cell line with high expression of PD-ECGF had a rapid cell cycle and consequently facilitated rapid cell growth not only in vivo but also in vitro. Furthermore, the inhibitor of thymidine phosphorylase (TPI) suppressed the cell cycle and rapid cell growth that were acquired by PD-ECGF transfection. Since PD-ECGF was reported to be an independent, poor prognosis factor for head and neck cancer, TPI might be useful for the inhibition of cancer cell growth.     

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.