Abstract LB-65: Bevacizumab fate in glioblastoma: Mechanism of internalization by endothelial cells and glioma stem cells

Abstract

Abstract Antiangiogenic therapy shows great promise for treatment of cancer. Bevacizumab is a humanized mAb that blocks VEGF-A, thereby inhibiting angiogenesis. It has received FDA approval for patients with recurrent glioblastoma (GBM); however, greater than 30% of patients are non-responsive and the underlying mechanism for the lack of response is not known. It has been assumed that Bevacizumab solely targets circulating VEGF-A; however, VEGF-A found in the tumor could also be an important target for Bevacizumab that impacts the response of patients. We hypothesized that Bevacizumab is transcytosed across brain endothelial cells (ECs), gains access to the perivascular niche containing glioma stem cells (GSCs), and that differences in GSC endocytosis of Bevacizumab and either recycling or degradation determine a patients response. We found that Bevacizumab is internalized by normal and tumor-isolated brain ECs in a time-dependent manner that is partially inhibited by amiloride and stimulated by EGF, suggesting Bevacizumab enters the cells by an endocytic pathway such as macropinocytosis. In a standard transcytosis assay, we showed that brain ECs transcytose 62% of Bevacizumab from the upper chamber to the lower chamber over two-hours. We also found that GSCs internalize Bevacizumab in a manner resembling macropinocytosis and that it is partially co-localized with LAMP1, suggesting Bevacizumab is partly targeted for degradation. Furthermore, in an orthotopic xenograft model of GBM administered Bevacizumab, we found that Bevacizumab was localized to the perivascular space, found within perivascular GBM tumor cells, and that a gradient of Bevacizumab was detected with less observed at further distances from vessels. Ongoing experiments will determine the endocytic compartment(s) containing Bevacizumab in GSCs and its trafficking. Understanding the mechanism of internalization and trafficking of Bevacizumab in GBM tumors and in isolated GSCs from different molecular subtypes will likely help us understand who will be responsive to Bevacizumab, and perhaps who will be responsive to therapy with other mAbs. Citation Format: Gaelle M. Muller-Greven, Cathleen Carlin, Jeremy Rich, Petra Hamerlik, Candece L. Gladson. Bevacizumab fate in glioblastoma: Mechanism of internalization by endothelial cells and glioma stem cells. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr LB-65. doi:10.1158/1538-7445.AM2014-LB-65