Combined inhibition of VEGF‐ and PDGF‐signaling enforces tumor vessel regression by interfering with pericyte‐mediated endothelial cell survival mechanisms

Abstract

Destruction of existing tumor blood vessels may be achieved by targeting vascular endothelial growth factor (VEGF) signaling, which mediates not only endothelial cell proliferation but also endothelial cell survival. In this study, however, intravital microscopy failed to demonstrate that targeting of VEGFR-2 (by the tyrosine kinase inhibitor SU5416) induces significant regression of experimental tumor blood vessels. Immunohistochemistry, electron microscopy, expression analyses, and in situ hybridization provide evidence that this resistance of tumor blood vessels to VEGFR-2 targeting is conferred by pericytes that stabilize blood vessels and provide endothelial cell survival signals via the Ang-1/Tie2 pathway. In contrast, targeting VEGFR-2 plus the platelet-derived growth factor receptor (PDGFR)-beta system (PDGFR-beta) signaling (by SU6668) rapidly forced 40% of tumor blood vessels into regression, rendering these tumors hypoxic as shown by phosphorescence quenching. TUNEL staining, electron microscopy, and apoptosis blocking experiments suggest that VEGFR-2 plus PDGFR-beta targeting enforced tumor blood vessel regression by inducing endothelial cell apoptosis. We further show that this is achieved by an interference with pericyte-endothelial cell interaction. This study provides novel insights into the mechanisms of how 1) pericytes may provide escape strategies to anti-angiogenic therapies and 2) novel concepts that target not only endothelial cells but also pericyte-associated pathways involved in vascular stabilization and maturation exert potent anti-vascular effects.

Inhibition of Vascular Endothelial Growth Factor (VEGF) Signaling in Cancer Causes Loss of Endothelial Fenestrations, Regression of Tumor Vessels, and Appearance of Basement Membrane Ghosts

“…This change is consistent with previous reports of the normalization of tumor vessels by VEGF/VEGFR inhibition. 32,33 Because this change in pericyte phenotype was found not only with AG013736 but also with ligand-specific VEGF-Trap, it is likely to result from inhibition of VEGF signaling. Receptor tyrosine kinase inhibitors that target PDGFRs reportedly produce pericyte loosening 33 or detachment 36,49 from endothelial cells of tumor vessels, arguing against a direct role of PDGFR inhibition in the changes we observed.…”

“…The complementary effects of inhibiting VEGFR and PDGFR signaling are consistent with this idea. 33,36 …”

“…28 -31 VEGF/VEGFR inhibition can decrease the diameter, tortuosity, and permeability of tumor vessels 28 and even transform surviving tumor vessels into a more normal phenotype. 32,33 Considering the important role of VEGF and its receptors in regulating vascular function, we sought to characterize the changes in tumor vessels produced by agents that block the action of this growth factor.…”

“…27,35 We focused on changes occurring during the first week of treatment of established tumors to identify primary vascular effects of the inhibitors that precede or accompany the reduction in tumor growth documented by others. 21,27,31,33,35,36 Fluorescence, confocal, and electron microscopic approaches applied to tissues fixed in situ by vascular perfusion provided cellular and molecular readouts for assessing patency and blood flow of individual vessels, endothelial sprouts and fenestrations, VEGFR-2 and VEGFR-3 immunoreactivity, pericyte morphology, and changes in the vascular basement membrane. The orderly microvasculature of the mouse trachea was used to validate some of the methods in a simpler system.…”

Incorporation of anti-angiogenesis therapy in the management of advanced ovarian carcinoma—Mechanistics, review of phase III randomized clinical trials, and regulatory implications

“…The FGF family activates angiogenesis via interaction with FGF receptors 1 and 2 [33]. It is thought that signaling via these alternate pathways (PDGF, FGF) may mediate resistance to VEGF inhibition, supporting a multi-targeted approach [34][35][36][37][38]. In response to receptor-ligand binding, downstream signaling pathways, PI3K-Akt-mTOR and Ras-MEK-Erk, are activated and have been identified as potential targets for drug development [39,40].…”