The angiogenic switch for vascular endothelial growth factor (VEGF)-A, VEGF-B, VEGF-C, and VEGF-D in the adenoma-carcinoma sequence during colorectal cancer progression.
ABSTRACT Angiogenesis is essential for tumour growth and metastasis. It is controlled by angiogenic factors, one of the most important being vascular endothelial growth factor (VEGF)-A. Although its role has been demonstrated in many tumour types including colorectal carcinoma (CRC), the importance of the newer family members in adenoma, invasive tumour growth, and progression to a metastatic phenotype has been poorly characterized in CRC. The aim of this study was to determine the role and timing of the VEGF angiogenic switch during CRC progression. We measured the gene expression of VEGF ligands (VEGF-A, VEGF-B, VEGF-C, and VEGF-D) and their receptors (VEGFR-1, VEGFR-2, and VEGFR-3), in normal colorectal tissues (n = 20), adenomas (n = 10), and in CRC (n = 71) representing different Duke's stages using ribonuclease protection assay, semi-quantitative relative reverse transcriptase polymerase chain reaction, together with the pattern of their expression by immunohistochemistry. VEGF-A mRNA was the most abundant in colorectal tissue, followed by VEGF-B, VEGF-C, and VEGF-D. VEGF-A and VEGF-B mRNAs were significantly more abundant in adenomas (p = 0.0003 and p = 0.04 respectively) compared with normal tissues, while VEGF-A and VEGF-C were significantly increased in carcinomas compared with normal tissues (p = 0.0006 and p = 0.0009 respectively). A significantly greater amount of VEGF-C mRNA was present in carcinomas compared with adenomas (p = 0.03), whereas there was a significant reduction of VEGF-B in carcinomas compared with adenomas (p = 0.0002). VEGF-D mRNA was significantly more abundant in normal tissues than in adenomas (p = 0.0001) and carcinomas (p < 0.0001). In normal tissues distant from the primary tumour, there was a significantly greater amount of VEGF-A and VEGF-D mRNA in patients with Duke's B and Duke's C respectively, compared with Duke's A stage tumours (p = 0.04 and p = 0.01 respectively). Immunohistochemistry showed low basal levels of all ligands in histologically normal tissues and their expression in the epithelium of tumours reflected the levels of mRNA expression identified. VEGF-A and VEGF-C mRNA levels correlated significantly with tumour grade (p = 0.01 and p = 0.01 respectively) and tumour size (p = 0.001 and p = 0.01 respectively), but not with patient age, sex, presence of infiltrative margin, lymphocytic response, vascular invasion, Duke's stage, or lymph node involvement (p > 0.05). VEGF-B mRNA correlated with an infiltrative margin (p = 0.04) but no other clinicopathological variable, and expression of VEGF-D demonstrated no association with any parameter examined. VEGFR-1 was significantly correlated with tumour grade (p = 0.02), Duke's stage (p < 0.001), and lymph node involvement (p = 0.004), VEGFR-2 with lymph node involvement (p = 0.02), and VEGFR-3 did not correlate with any of the clinicopathological variables tested. These results suggest that VEGF-A and VEGF-B play a role early in tumour development at the stage of adenoma formation and that VEGF-C plays a role in advanced disease when there is more likelihood of metastatic spread. The finding of increased levels of VEGF-A and VEGF-D expression in normal tissues collected from a site distant from the primary tumour indicates changes in the surrounding tumour environment that may enhance the subsequent spread of tumour cells.
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ABSTRACT: Recent studies have suggested that vascular endothelial growth factor (VEGF), in addition to its proangiogenic properties, also functions as a survival factor for endothelial cells. The authors hypothesized that inhibition of VEGF activity by blockade of VEGF receptor-2 (R-2) function prevents angiogenesis and decreases tumor growth in colon carcinoma liver metastases. Spleens of mice were injected with human colon carcinoma cells producing liver metastases. After 7 days of tumor growth, groups of mice received either antibody to VEGFR-2 (DC101) or phosphate-buffered saline (control). In a follow-up experiment, a similar treatment regimen was followed except that mice were sacrificed at 1-week intervals to assess the time course of endothelial cell and tumor cell apoptosis. After 21 days of therapy, the authors observed a significant decrease in vessel counts in liver metastases from human colon carcinoma in nude mice after therapy with VEGFR-2 antibody. Tumor cell apoptosis was increased significantly in the tumors of mice receiving DC101. Temporal studies with immunofluorescent double staining for the microvasculature and apoptotic cells revealed an increase in endothelial cell apoptosis that preceded an increase in tumor cell apoptosis. In vitro, treatment of human umbilical vein endothelial cells with antibody to VEGFR-2 produced a > 2.5-fold increase in endothelial cell apoptosis. Therapy targeting the VEGFR-2 inhibited tumor growth in a murine model of colon carcinoma liver metastasis. Surprisingly, this therapy did not only inhibit angiogenesis but also led to endothelial cell death. These findings suggest that VEGF, via VEGFR-2 signaling, functions as a survival factor for tumor endothelial cells in liver metastases from colon carcinoma.Cancer 08/2000; 89(3):488-99. · 5.20 Impact Factor
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ABSTRACT: A growth factor for vascular endothelial cells was identified in the media conditioned by bovine pituitary follicular cells and purified to homogeneity by a combination of ammonium sulfate precipitation, heparin-sepharose affinity chromatography and two reversed phase HPLC steps. The growth factor was a cationic, heat stable and relatively acid stable protein and had a molecular weight, as assessed by silver-stained SDS-PAGE gel, of approximately 45,000 under non reducing conditions and approximately 23,000 under reducing conditions. The purified growth factor had a maximal mitogenic effect on adrenal cortex-derived capillary endothelial cells at the concentration of 1-1.2 ng/ml (22-26 pM). Further characterization of the bioactivity of the growth factor reveals that it exerts mitogenic effects also on vascular endothelial cells isolated from several districts but not on adrenal cortex cells, lens epithelial cells, corneal endothelial cells, keratynocytes or BHK-21 fibroblasts, indicating that its target cells specificity is unlike that of any previously characterized growth factor. Microsequencing reveals a unique N-terminal amino acid sequence. On the basis of its apparent target cell selectivity, we propose to name this factor vascular endothelial growth factor (VEGF).Biochemical and Biophysical Research Communications 07/1989; 161(2):851-8. · 2.41 Impact Factor
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ABSTRACT: We aim to determine the clinical usefulness of pre-operative serum vascular endothelial growth factor (VEGF) as a predictor of outcome in patients undergoing curative resection for colorectal cancer. Serum VEGF was assayed by quantitative ELISA in 81 patients prior to curative resection for node-negative (n = 53) and node-positive (n = 28) disease. Median follow-up for patients without cancer death was 27 months (range 21-37). Pre-operative serum VEGF was significantly higher in patients who went on to develop metastases than those who did not (median, 713 pg ml-1 vs. 314 pg ml-1, P < 0.0001). Using multivariate Cox regression analysis, pre-operative serum VEGF was the most important prognostic factor independent of nodal status and adjuvant chemotherapy, and was superior to nodal status in predicting outcome (P < 0.00001). At 575 pg ml-1, pre-operative serum VEGF was 64% sensitive and 89% specific in predicting the development of metastases in curative resections, with a positive predictive value of 73% and a negative predictive value of 85%. Pre-operative serum VEGF is a powerful predictor of outcome following curative surgery for colorectal cancer. These data support the measurement of pre-operative serum VEGF as a method for selecting patients who require adjuvant therapy.British Journal of Cancer 12/2000; 83(11):1425-31. · 5.08 Impact Factor