Identification of vascular cues contributing to cancer cell stemness and function.

Glioblastoma stem cells (GSCs) reside close to blood vessels (BVs) but vascular cues contributing to GSC stemness and the nature of GSC-BVs cross talk are not fully understood. Here, we dissected vascular cues influencing GSC gene expression and function to perfusion-based vascular cues, as well as to those requiring direct GSC-endothelial cell (EC) contacts. In light of our previous finding that perivascular tumor cells are metabolically different from tumor cells residing further downstream, cancer cells residing within a narrow, < 60 µm wide perivascular niche were isolated and confirmed to possess a superior tumor-initiation potential compared with those residing further downstream. To circumvent reliance on marker expression, perivascular GSCs were isolated from the respective locales based on their relative state of quiescence. Combined use of these procedures uncovered a large number of previously unrecognized differentially expressed GSC genes. We show that the unique metabolic milieu of the perivascular niche dominated by the highly restricted zone of mTOR activity is conducive for acquisition of GSC properties, primarily in the regulation of genes implicated in cell cycle control. A complementary role of vascular cues including those requiring direct glioma/EC contacts was revealed using glioma/EC co-cultures. Outstanding in the group of glioma cells impacted by nearby ECs were multiple genes responsible for maintaining GSCs in an undifferentiated state, a large fraction of which also relied on Notch-mediated signaling. Glioma-EC communication was found to be bidirectional, evidenced by extensive Notch-mediated EC reprogramming by contacting tumor cells, primarily metabolic EC reprogramming.