Identifying novel modulators of vasculogenesis
Dr. Lamis Hammoud
Former Research Associate
Mammalian vascular development begins shortly after the initiation of gastrulation. Blood vessels develop by two processes: vasculogenesis and angiogenesis. Vasculogenesis occurs in both the embryo and yolk sac by aggregation of de-novo-forming angioblasts (endothelial precursors) into a primitive network of endothelial tubes known as the primitive vascular plexus. This plexus then undergoes angiogenesis: remodeling involving growth, proliferation, regression, migration and sprouting of new vessels from existing ones, resulting in the formation of the mature circulatory network. VEGF, acting through the Flk1/VEGFR2 receptor, is crucial for blood vessel formation and development. Many events that occur during embryonic vascular development are recapitulated during adult neoangiogenesis, which is critical to tumour growth and metastasis. While the latest antiangiogenic drugs, such as Avastin (anti-VEGF), have been shown to prolong life expectancy in cancer patients, they have serious side effects. Furthermore, relapses often occur necessitating the need for novel therapeutic targets.
The goals of our research are two-fold; to gain a better understanding of the downstream effectors of the VEGF/Flk1 pathway; and to develop a robust in vitro vascular differentiation assay using mouse embryonic stem (mES) cells suitable for small molecule screens to identify novel modulators of angiogenesis. Embryonic stem cells have the ability to differentiate into many tissues, such as blood vessels, in a manner that resembles embryonic development. To that end, we have optimized an in vitro mES cell-based differentiation assay using Flk-eGFP ES cells. This assay has been validated using a variety of vascular modulators. Furthermore, we developed an algorithm using the Cellomics ArrayScan platform, based on the neuronal profiling software, to quantify total expression of Flk1 as well as the number of fluorescent sprouts. Currently, we are using this vascular differentiation assay to screen a kinome small molecule inhibitor library provided by the Medicinal Chemistry Platform of the Ontario Institute for Cancer Research (OICR). Promising candidates showing quantitative deviation from control cultures have been observed, will be validated in secondary assays, and their efficacy will be tested in preclinical tumour models in mice.