4779

For years, the chorioallantoic membrane (CAM) of an avian embryo has been exploited in the study of angiogenesis and tumor cell metastasis. We propose a novel application of this versatile system in the study of vascular permeability and vascular leakage. Currently the investigation of molecular and biochemical processes involved in both of these phenomena is limited to the use of either simplified in-vitro assays or expensive in vivo analysis of Evans Blue dye extravasation in the tissues of mice. The CAM system we describe is an in vivo system that combines the versatility of in vitro assays with the tissue complexity of higher order in vivo systems. The CAM is a highly vascularized extra-embryonic membrane connected to the embryo through a continuous circulatory system. Because the membrane is external, the CAM is readily accessible for experimental manipulations, including the topical application or i.v. injection of treatments and the optical visualization of local responses. We have developed an in vivo assay for the visualization of vascular leak in response to vascular permeabilizing factors. Here we describe the use of this assay in screening a new class of drugs known as Vasopermeation Enhancement Agents (VEAs), which are designed to increase the uptake of cancer therapeutics at the tumor site. While the ability of VEAs to improve vasopermeability has been documented in the literature, the mechanism of VEA action is still unknown. The CAM is an ideal vascular model in which to clarify the process. Here we present data using lipopolysaccharides (LPS), which are known to induce vascular leakage, as a control. We also present data from Interleukin-2, whose vascular leak side effects have limited its usage as a cancer therapeutic. Most importantly, we present data from a fragment of Interleukin-2 referred to as the Permeability Enhancing Peptide (PEP). PEP is in the forefront of development as a VEA for improving the therapeutic index of several standard chemotherapies. In each of these cases, we visually illustrate vascular leakage using fluorescent dextran as a marker and demonstrate how the phenomenon can be measured over time. Furthermore, we discuss the potential of this system in drug screening for future compounds. With respect to cost and experimental duration, CAMs provide an attractive alternative to murine models in early stage studies for the evaluation of vasopermeability or vascular leakage, be it the intentional purpose of a drug or an unwanted side effect that must be removed through genetic manipulation.

[Proc Amer Assoc Cancer Res, Volume 47, 2006]