Abstract 4170: The role of VEGF-C for cell viability, tumor growth and bevacizumab resistance in glioblastoma multiforme
Authors: Michaelsen Signe R. , Nedergaard Mette K. , Urup Thomas , Villingshoej Mette , Kjaer Andreas , Perryman Lara , Erler Janine T. , Lassen Ulrik , Poulsen Hans S. 
Published: August 2015

Abstract
Although treatment with the VEGF-A targeting antibody Bevacizumab has resulted in substantial response rates in patients with the brain tumor Glioblastoma Multiforme, patients invariable progress. The VEGF-A receptor VEGFR2, has in addition to be expressed on endothelial cells, recently been shown also to be expressed by GBM tumor cells. Autocrine VEGF-A/VEGFR2 signaling has been identified in GBM cells, but contradicting results exist for the effects of inhibiting VEGF-A and VEGFR2 respectively; indicating that VEGFR2 in GBM at least partly is activated by other factors than VEGF-A. Study aim was to search for other factors responsible for VEGFR2 activation in GBM cells and to examine their role for GBM growth and response to Bevacizumab.
To study VEGFR2 regulation we examined a VEGFR2-positive and a VEGFR2-negative GBM cell culture, which both expressed VEGF-A. In line with others, we found that VEGFR2 phosphorylation could be stimulated by recombinant VEGF-A, and found that inhibition of receptor phosphorylation by SU1498 resulted in significantly reduced proliferation of the VEGFR2-positive cells, while inhibition of the VEGF-A expressed by the cells using Bevacizumab only had minimal effect on proliferation. Examinations revealed that the VEGFR2-positive cells also were positive for the VEGF variant VEGF-C. Addition of recombinant VEGF-C protein to the VEGFR2-positive cells could stimulate VEGFR2 phosphorylation, while inhibition of VEGF-C using siRNA constructs resulted in reduced in vitro growth of VEGFR2-positive cells. Further, when injected into the brains of mice, VEGF-C-siRNA transfected cells resulted in reduced tumor growth and increased survival compared to control cells. Moreover, measurement of the VEGF-C mRNA level in 19 GBM patient tumors showed that all tumors were positive for VEGF-C expression, although the level was varying. To examine if VEGF-C expression represents a possible mechanism behind insensitivity towards Bevacizumab therapy, we are currently investigating the expression of VEGF-C in paired samples from GBM patients taken before and after Bevacizumab therapy and in xenograft tumors from mice receiving this treatment.
In conclusion, our current results show that VEGF-C is of importance for GBM cell viability and tumor growth presumable due to its ability to stimulate autocrine activation of VEGFR2. VEGF-C expression therefore could respresent a possible mechanism behind Bevacizumab resistance. An update on this will be presented.
Citation Format: Signe R. Michaelsen, Mette K. Nedergaard, Thomas Urup, Mette Villingshoej, Andreas Kjaer, Lara Perryman, Janine T. Erler, Ulrik Lassen, Hans S. Poulsen. The role of VEGF-C for cell viability, tumor growth and bevacizumab resistance in glioblastoma multiforme. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 4170. doi:10.1158/1538-7445.AM2015-4170