Immunohistochemical expression of vascular endothelial growth factor and microvessel counting as prognostic indicators in node-negative colorectal cancer

Abstract

This manuscript reports a carefully controlled study of patients with Dukes B colorectal cancer (Dukes stage A, n=12 and Dukes stage B, n=44). Immunohistochemistry has been used to demonstrate reactivity for vascular endothelial growth factor (VEGF), and to measure levels of microvessel density (MVD) in order to assess the relationship of tumor angiogenesis with clinical outcome. Immunohistochemistry was performed using antibodies to VEGF and CD34 (for intratumoral vessel identification) and counting was performed at the invasive margin of the tumor. Results showed that for Dukes stage A patients 4/12 died of their disease, none of whose tumor was VEGF positive. In contrast, 2 patients who survived were positive for VEGF cytoplasmically, but neither showed increased tumor MVD. In Dukes B patients 10/44 died, 5 of whose tumor demonstrated VEGF reactivity, both in malignant cells and in tumor vascular endothelium. MVD ranged from 11 to 53 (median 28) for Dukes A cases and from 9 to 69 (median 32.5) for the Dukes B group. Kaplan-Meier plots and log rank test statistics for Dukes B patients demonstrated that VEGF reactivity in cells, and in tumor vascular endothelium was correlated with survival (p=0.047 and p < or = 0.06, respectively). There was a significant relationship between the presence of VEGF reactivity on vascular endothelium and outcome by Fisher's exact test (p=0.018). Similarly, by the same test VEGF positivity was significantly correlated with patient mortality (p=0.032). The presence of endothelial VEGF reactivity correlated with VEGF in malignant cells (p=0.0001) by Mann-Whitney U test and a significant inverse relationship between vessel density and patient survival was demonstrated (p = 0.019). The finding that in Dukes B patients MVD was inversely correlated with mortality supports the hypothesis that a low microvascular count is predicted close to the invasive margin, where VEGF expression is upregulated in response to hypoxia, induced by a lack of a functional vasculature. These data will be used to identify cohorts of patients who have a high risk of relapse and can be selected for adjuvant therapies such as VEGF antibody or antitumor antibody-directed therapy.