Macropinocytosis of Bevacizumab by Glioblastoma Cells in the Perivascular Niche Affects their Survival

• Type of Material: Article
• Authors: Gaelle Mueller-Greven, Cleveland Clinic; Cathleen R. Carlin, Case Western Reserve University; Monica E. Burgett, Cleveland Clinic; Manmeet S. Ahluwalia, Cleveland Clinic; Adam Lauko, Cleveland Clinic; Amy S. Nowacki, Cleveland Clinic; Cameron J. Herting, Emory University; Maha A. Qadan, Cleveland Clinic; Markus Bredel, University of Alabama Birmingham; Steven A. Toms, Geisinger Medical Center; Justin D. Lathia, Cleveland Clinic; Dolares Hambardzumyan, Emory University; Jann N. Sarkaria, Mayo Clinic; Petra Hamerlik, Danish Cancer Society; Candece L. Gladson, Cleveland Clinic
• Language: English
• Date: 2017-11-15
• Publisher: American Association for Cancer Research
• Publication Version: Author Accepted Manuscript (After Peer Review)
• Copyright Statement: ©2017 AACR.
• Final Published Version (URL): http://dx.doi.org/10.1158/1078-0432.CCR-17-0249
• Title of Journal or Parent Work: Clinical Cancer Research
• ISSN: 1078-0432
• Volume: 23
• Issue: 22
• Start Page: 7059
• End Page: 7071
• Grant/Funding Information: The Leica-SP5-confocal/multi-photon microscope utilized was purchased with partial funding from NIH SIG-grant 1S10RR026820-01.; This research was supported by NIH grants R01CA175120 (C.L. Gladson), U01 CA160882 (D. Hambardzumyan), R01CA176830 and Mayo Brain Tumor SPORE P50 CA108961 (J.N. Sarkaria), and Velosano Foundation funding (C.L. Gladson).
• Keywords: Science & Technology; ENDOTHELIAL GROWTH-FACTOR; IN-VITRO; CANCER; INDUCED AUTOPHAGY; TUMOR-GROWTH; ENDOCYTIC PATHWAY; STEM-CELLS; RECURRENT GLIOBLASTOMA; Life Sciences & Biomedicine; FC-RECEPTOR; Oncology; PROTEIN
• Research Categories: Health Sciences, Oncology

Abstract

Purpose: Bevacizumab, a humanized monoclonal antibody to VEGF, is used routinely in the treatment of patients with recurrent glioblastoma (GBM). However, very little is known regarding the effects of bevacizumab on the cells in the perivascular space in tumors. Experimental Design: Established orthotopic xenograft and syngeneic models of GBM were used to determine entry of monoclonal anti-VEGF-A into, and uptake by cells in, the perivascular space. Based on the results, we examined CD133+ cells derived from GBM tumors in vitro. Bevacizumab internalization, trafficking, and effects on cell survival were analyzed using multi-label confocal microscopy, immunoblotting, and cytotoxicity assays in the presence/absence of inhibitors. Results: In the GBM mouse models, administered anti-mouse-VEGF-A entered the perivascular tumor niche and was internalized by Sox2+/CD44+ tumor cells. In the perivascular tumor cells, bevacizumab was detected in the recycling compartment or the lysosomes, and increased autophagy was found. Bevacizumab was internalized rapidly by CD133+/Sox2+-GBM cells in vitro through macropinocytosis with a fraction being trafficked to a recycling compartment, independent of FcRn, and a fraction to lysosomes. Bevacizumab treatment of CD133+ GBM cells depleted VEGF-A and induced autophagy thereby improving cell survival. An inhibitor of lysosomal acidification decreased bevacizumab-induced autophagy and increased cell death. Inhibition of macropinocytosis increased cell death, suggesting macropinocytosis of bevacizumab promotes CD133+ cell survival. Conclusions: We demonstrate that bevacizumab is internalized by Sox2+/CD44+-GBM tumor cells residing in the perivascular tumor niche. Macropinocytosis of bevacizumab and trafficking to the lysosomes promotes CD133+ cell survival, as does the autophagy induced by bevacizumab depletion of VEGF-A.