Abstract 2307: Small GTPase R-Ras promotes normalization of tumor vasculature: a potential oxygen-sensing independent mechanism.

Abstract Newly formed blood vessels in tumors fail to mature into fully functional vessels due to the chronically angiogenic microenvironment. The functional impairment of these vessels hampers drug delivery, thereby diminishing the efficacy of anti-tumor therapies. Excessive vessel permeability associated with tumors also causes clinical complications such as cerebral edema in brain cancer patients. The ability to control vessel maturity in tumors, therefore, provides a potential therapeutic opportunity. We show that R-Ras is essential for the establishment of mature, functional blood vessels in tumors. R-Ras is a small GTPase of the Ras family with antiangiogenic activity. We found that malformation and malfunction of tumor vessels are exacerbated by genetic disruption of R-Ras. R-Ras-deficient vessels exhibited severely impaired pericyte and basement membrane supports, disrupted adherens junctions, and excessive blood leakage in tumors. Tumor cell penetration into the blood circulation increased in R-Ras-deficient mice, supporting a role for R-Ras in controlling tumor vessel permeability. Decreased blood perfusion of tumor vessels in R-Ras-deficient mice elevated tumor hypoxia, which in turn led to decreased efficacy of radiotherapy in these mice. In contrast to R-Ras disruption, R-Ras gain-of-function improved vessel structure and blood perfusion and blocked plasma leakage by enhancing endothelial barrier function and pericyte association with nascent blood vessels. The studies of cell type-specific expression of R-Ras revealed that R-Ras in endothelial cells and in pericytes both contribute individually to the vessel regulation. For instance, endothelial R-Ras limits vascular permeability by inhibiting VE-cadherin internalization induced by VEGF without affecting VE-cadherin gene expression level. These findings indicate that R-Ras is critical for controlling blood circulation and vascular permeability in solid tumors. It has been shown that haplodeficiency of the oxygen-sensor PHD2 leads to normalization of tumor vasculature through stabilization of endothelial HIF-2 and consequent upregulation of soluble VEGFR1 and VE-cadherin gene expression in endothelial cells. Our studies of endothelial cells in hypoxia and normoxia indicate that R-Ras is not involved in oxygen-sensing mechanisms of vessel regulations mediated by the PHD2/HIF-2 pathway. There are important differences between the vessel regulations by PHD2/HIF-2 and by R-Ras. PHD2 haplodeficiency does not affect vessel density, area, or vessel dilation while it normalizes the endothelial barrier and stability. In comparison, R-Ras does normalize all of these vessel parameters while halting angiogenic sprouting. Our observations suggest that an alternative, oxygen sensing-independent vascular normalization phenomenon. Citation Format: Masanobu Komatsu, Junko Sawada, Takeo Urakami, Fangfei Li, Akane Urakami. Small GTPase R-Ras promotes normalization of tumor vasculature: a potential oxygen-sensing independent mechanism. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 2307. doi:10.1158/1538-7445.AM2013-2307