Macropinocytosis of bevacizumab by glioblastoma cells in the perivascular niche affects their survival

Gaëlle Müller-Greven, Cathleen R. Carlin, Monica E. Burgett, Manmeet S. Ahluwalia, Adam Lauko, Amy S. Nowacki, Cameron J. Herting, Maha A. Qadan, Markus Bredel, Steven A. Toms, Justin D. Lathia, Dolores Hambardzumyan, Jann N Sarkaria, Petra Hamerlik, Candece L. Gladson

Research output: Contribution to journalArticle

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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.

Original languageEnglish (US)
Pages (from-to)7059-7071
Number of pages13
JournalClinical Cancer Research
Volume23
Issue number22
DOIs
StatePublished - Nov 15 2017

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Glioblastoma
Survival
Vascular Endothelial Growth Factor A
Autophagy
Cell Survival
Lysosomes
Neoplasms
Recycling
Bevacizumab
Cell Death
Antibodies, Monoclonal, Humanized
Immunoblotting
Heterografts
Confocal Microscopy
Research Design

ASJC Scopus subject areas

  • Oncology
  • Cancer Research

Cite this

Müller-Greven, G., Carlin, C. R., Burgett, M. E., Ahluwalia, M. S., Lauko, A., Nowacki, A. S., ... Gladson, C. L. (2017). Macropinocytosis of bevacizumab by glioblastoma cells in the perivascular niche affects their survival. Clinical Cancer Research, 23(22), 7059-7071. https://doi.org/10.1158/1078-0432.CCR-17-0249

Macropinocytosis of bevacizumab by glioblastoma cells in the perivascular niche affects their survival. / Müller-Greven, Gaëlle; Carlin, Cathleen R.; Burgett, Monica E.; Ahluwalia, Manmeet S.; Lauko, Adam; Nowacki, Amy S.; Herting, Cameron J.; Qadan, Maha A.; Bredel, Markus; Toms, Steven A.; Lathia, Justin D.; Hambardzumyan, Dolores; Sarkaria, Jann N; Hamerlik, Petra; Gladson, Candece L.

In: Clinical Cancer Research, Vol. 23, No. 22, 15.11.2017, p. 7059-7071.

Research output: Contribution to journalArticle

Müller-Greven, G, Carlin, CR, Burgett, ME, Ahluwalia, MS, Lauko, A, Nowacki, AS, Herting, CJ, Qadan, MA, Bredel, M, Toms, SA, Lathia, JD, Hambardzumyan, D, Sarkaria, JN, Hamerlik, P & Gladson, CL 2017, 'Macropinocytosis of bevacizumab by glioblastoma cells in the perivascular niche affects their survival', Clinical Cancer Research, vol. 23, no. 22, pp. 7059-7071. https://doi.org/10.1158/1078-0432.CCR-17-0249
Müller-Greven G, Carlin CR, Burgett ME, Ahluwalia MS, Lauko A, Nowacki AS et al. Macropinocytosis of bevacizumab by glioblastoma cells in the perivascular niche affects their survival. Clinical Cancer Research. 2017 Nov 15;23(22):7059-7071. https://doi.org/10.1158/1078-0432.CCR-17-0249
Müller-Greven, Gaëlle ; Carlin, Cathleen R. ; Burgett, Monica E. ; Ahluwalia, Manmeet S. ; Lauko, Adam ; Nowacki, Amy S. ; Herting, Cameron J. ; Qadan, Maha A. ; Bredel, Markus ; Toms, Steven A. ; Lathia, Justin D. ; Hambardzumyan, Dolores ; Sarkaria, Jann N ; Hamerlik, Petra ; Gladson, Candece L. / Macropinocytosis of bevacizumab by glioblastoma cells in the perivascular niche affects their survival. In: Clinical Cancer Research. 2017 ; Vol. 23, No. 22. pp. 7059-7071.
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T1 - Macropinocytosis of bevacizumab by glioblastoma cells in the perivascular niche affects their survival

AU - Müller-Greven, Gaëlle

AU - Carlin, Cathleen R.

AU - Burgett, Monica E.

AU - Ahluwalia, Manmeet S.

AU - Lauko, Adam

AU - Nowacki, Amy S.

AU - Herting, Cameron J.

AU - Qadan, Maha A.

AU - Bredel, Markus

AU - Toms, Steven A.

AU - Lathia, Justin D.

AU - Hambardzumyan, Dolores

AU - Sarkaria, Jann N

AU - Hamerlik, Petra

AU - Gladson, Candece L.

PY - 2017/11/15

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N2 - 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.

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