Line II

Line III

Line IV

PhD theses

Movies

Funding

Angiogenesis and immunity

The main aim of research line III is to investigate the relationship between angiogenesis and the immune system. The ultimate goal is to investigate whether angiogenesis inhibition can be used to improve immunotherapy.

Angiogenic factors induce endothelial cell anergy

Endothelial cell adhesion molecules like intercellular adhesion molecule-1 (ICAM-1), vascular cell adhesion molecule (VCAM-1), P(latelet)- and E(ndotelial)-selectin are involved in the lymphocyte adhesion cascade. Selectins mediate rolling of leukocytes, which is followed by strong adhesion and transmigration mediated by the immunoglobulin-related endothelial adhesion molecules, ICAM-1 and VCAM-1. The expression of these molecules is increased by inflammatory cytokines like TNF , IL-1 and INF at the site of inflammation.
Tumor cells produce high concentrations of angiogenic factors like bFGF and VEGF that activate endothelial cells (ECs) to proliferate, migrate and form new blood vessels. Interestingly, these angiogenic factors also induce down-regulation of adhesion molecule expression on EC (see figure 1, Griffioen et al. Cancer Res. 56:1111, 1996). This leads to fewer leukocyte-EC interactions and ultimately to tumor escape from immunity (Dirkx et al. Cancer Res. 63:2322, 2003).

Figure 1: A. FACS analysis of whole-tumor cell suspensions. Endothelial cells are identified with EN4 anti-CD31 antibody. Upper panels are from normal renal tissue, lower panels are from renal cell carcinoma tissue. B. Quantification of the expression differences. (from: Griffioen et al. Cancer Research 56:1111, 1996)

Angiogenic potential determines leukocyte tumor infiltration and patient survival

Human studies in ductal and medullary carcinoma revealed a relationship between angiogenic profile and leukocyte infiltration (Ter Steege et al. Clin.Cancer Res. 2004). Medullary carcinoma - with better clinical prognosis- showed a higher number of leukocyte infiltrations compared to ductal carcinoma. This was accompanied by an enhanced ICAM-1 expression in medullary carcinoma. The up-regulation can be explained by the angiogenic profile of these tumors, which was assessed by qPCR. The expression of angiogenic factors in medullary carcinoma was reduced compared to the expression in ductal carcinoma leading to an up-regulation of ICAM-1 expression and leukocyte tumor infiltration.
Studies in archival colorectal carcinoma (CRC) showed similar results. We found a correlation between the intrinsic tumor parameters of ongoing angiogenesis and leukocyte infiltration (amount and composition) with prognosis and survival in CRC (Baeten et al. Clin.Gastroenterol.Hepatol. In press 2006). These findings have a potential impact on therapeutic applications for both anti-angiogenesis as well as immunotherapy.

Angiogenesis inhibitors overcome endothelial cell anergy

In vitro studies on endothelial cells (EC) suggested that angiogenesis mediated downregulation of endothelial adhesion molecules could be prevented by treatment with angiogenesis inhibitors. It was hypothesized that anti-angiogenesis therapy in vivo could circumvent endothelial anergy. We found that the angiogenesis inhibitors anginex, endostatin and angiostatin, and also the chemotherapeutic agent paclitaxel were able to significantly stimulate leukocyte-tumor vessel wall interactions (click the links below to view video images of intravital microscopy) by up-regulation of endothelial adhesion molecule expression. Consequently the infiltration of leukocytes into the tumor was augmented (see figure 2). The current results suggest that immunotherapy strategies can be improved by combination with anti-angiogenesis.

Please click the links below to view video images of leukocyte vessel wall interactions in

normal control tissue

tumor tissue

tumor tissue of mice treated with anginex

Figure 2: Relationship between angiogenesis inhibition and leukocyte infiltration.
Tumor size (A), microvessel density assessment by CD31 staining (B) and infiltration by CD45+ leukocytes (C) of LS174T human colon carcinoma with and without anginex treatment. D. Immunohistochemical images of microvessel density (CD31), and infiltration by CD45+ leukocytes and CD8+ T lymphocytes in B16F10 melanoma of control and anginex treated mice. Bar in upper left panel represents 50 µm. E-H: Effect of increasing doses of anginex (2, 6, 12 mg/kg/day) on tumor size, microvessel density, total leukocyte (CD45)-, and cytotoxic T lymphocyte (CD8) infiltration in B16F10 melanoma.*p<0.01, **p<0.001 and ***p<0.0001 as compared to control.