Osteopontin mediates glioblastoma-associated macrophage infiltration and is a potential therapeutic target
Jun Wei, … , Shulin Li, Amy B. Heimberger
Jun Wei, … , Shulin Li, Amy B. Heimberger
Published January 2, 2019; First published October 11, 2018
Citation Information: J Clin Invest. 2019;129(1):137-149. https://doi.org/10.1172/JCI121266.
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Categories: Research Article Immunology

Osteopontin mediates glioblastoma-associated macrophage infiltration and is a potential therapeutic target

Abstract

Glioblastoma is highly enriched with macrophages, and osteopontin (OPN) expression levels correlate with glioma grade and the degree of macrophage infiltration; thus, we studied whether OPN plays a crucial role in immune modulation. Quantitative PCR, immunoblotting, and ELISA were used to determine OPN expression. Knockdown of OPN was achieved using complementary siRNA, shRNA, and CRISPR/Cas9 techniques, followed by a series of in vitro functional migration and immunological assays. OPN gene–deficient mice were used to examine the roles of non-tumor-derived OPN on survival of mice harboring intracranial gliomas. Patients with mesenchymal glioblastoma multiforme (GBM) show high OPN expression, a negative survival prognosticator. OPN is a potent chemokine for macrophages, and its blockade significantly impaired the ability of glioma cells to recruit macrophages. Integrin αvβ5 (ITGαvβ5) is highly expressed on glioblastoma-infiltrating macrophages and constitutes a major OPN receptor. OPN maintains the M2 macrophage gene signature and phenotype. Both tumor-derived and host-derived OPN were critical for glioma development. OPN deficiency in either innate immune or glioma cells resulted in a marked reduction in M2 macrophages and elevated T cell effector activity infiltrating the glioma. Furthermore, OPN deficiency in the glioma cells sensitized them to direct CD8+ T cell cytotoxicity. Systemic administration in mice of 4-1BB–OPN bispecific aptamers was efficacious, increasing median survival time by 68% (P < 0.05). OPN is thus an important chemokine for recruiting macrophages to glioblastoma, mediates crosstalk between tumor cells and the innate immune system, and has the potential to be exploited as a therapeutic target.

Authors

Jun Wei, Anantha Marisetty, Brett Schrand, Konrad Gabrusiewicz, Yuuri Hashimoto, Martina Ott, Zacharia Grami, Ling-Yuan Kong, Xiaoyang Ling, Hillary Caruso, Shouhao Zhou, Y. Alan Wang, Gregory N. Fuller, Jason Huse, Eli Gilboa, Nannan Kang, Xingxu Huang, Roel Verhaak, Shulin Li, Amy B. Heimberger

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Figure 4

OPN is chemotactic for macrophages.

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OPN is chemotactic for macrophages. (A) Transwell migration assays of hu...
(A) Transwell migration assays of human M0- and M2-skewed macrophages exposed to serum-free medium (negative control), 10% FBS (positive control), recombinant OPN (rOPN, 50 ng/ml), and conditioned medium (CM) from GSCs at 48 hours. Original magnification, ×100. (B) Sequential dilution of GSC CM demonstrating a dose response of OPN for macrophage migration. Data indicate mean ± SD and are representative of 3 independent experiments. (C) Supernatants from GSCs were treated with 10 μg/ml of OPN-neutralizing antibody or the isotype IgG control and then applied to the lower chamber in the presence or absence of 10 ng/ml OPN for attracting M0 and M2 macrophages. Data are shown as mean ± SD and are representative of 3 independent experiments. (D) Supernatants from OPN sgRNA/CRISPR (OPN KO) and NT scramble GSC cell lines (NT ctrl) were used to induce migration of M0 macrophages. Original magnification, ×100. Data are shown as mean ± SD and are representative of 2 independent experiments. Similar results were obtained using M2 macrophages (data not shown). P values were calculated using the 2-tailed 2-sample t test. *P < 0.05.