Introduction: Vascular endothelial growth factor (VEGF) is a key regulator of angiogenesis and vascular function. The mechanisms of VEGF signaling in angiogenesis have been extensively studied. However, its effects on tumor cell function remain to be elucidated. Our studies on depletion of VEGF by somatic knockout of the VEGF gene or by siRNA in human colorectal cancer (CRC) cell lines have demonstrated that loss of VEGF expression led to significantly decreased cell growth, increased apoptosis, and enhanced chemo-sensitivity of CRC cells through an intracrine signaling mechanism. These intracrine effects were mediated through VEGF by regulation of the activity of multiple receptor tyrosine kinases (e.g. EGFR, cMET) and downstream AKT signaling. Similar effects were observed by depletion of VEGF receptor 1 (VEGFR1), supporting the role of a novel intracellular VEGF-VEGFR1 complex in CRC cell survival. Since VEGF signaling has been shown to effect endothelial and tumor cell migration, we attempted to determine if inhibition of intracrine VEGF signaling affected CRC cell motility.
Methods: Migration and invasion of CRC cells, with and without VEGF depletion, were assayed using Boyden chambers and a serum gradient. Changes in cell morphology and epithelial to mesenchymal transition (EMT) markers and markers of cell motility were assessed by microscopy, immunostaining, and western blot analyses.
Results: Depletion of VEGF by siRNAs in multiple CRC cell lines led to a very strong inhibition of migration and invasion of CRC cells. Such inhibition was not observed when CRC cells were treated with bevacizumab to inhibit paracrine or autocrine signaling. Further examination revealed no significant changes in cell morphology. Other than Twist, there were no significant differences between specific markers of EMT indicating that differences in migration and invasion were not due to classic alterations in EMT. However, significant changes in activation of mediators of cell motility were observed. VEGF depleted CRC cells demonstrated significantly lower levels of phosphorylated focal adhesion kinase (p-FAK) compared to cells with normal VEGF expression. Analyses of upstream regulators indicated strong reduction in either p-cMET or p-SRC depending on cell type.
Conclusions: Inhibition of intracrine VEGF signaling strongly inhibits CRC cell migration and invasion by regulating proteins involved in cell motility.
Citation Format: Rajat Bhattacharya, Ling Xia, Fan Fan, Rui Wang, Delphine Boulbes, Xiang-Cang Ye, Lee Ellis. Inhibition of intracrine VEGF signaling prevents colorectal cancer cell migration and invasion. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 3255.