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 October 11, 2018
Citation Information: J Clin Invest. 2019;129(1):137-149. https://doi.org/10.1172/JCI121266.
View: Text | PDF
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

×

Figure 3

Reduction of macrophage infiltration and enhanced T cell effector activity in the tumor microenvironment is due to OPN loss.

Options: View larger image (or click on image) Download as PowerPoint
Reduction of macrophage infiltration and enhanced T cell effector activi...
Either WT or Opn–/– background mice were intracranially implanted with GL261 cells that were transfected with an NT control shRNA or shRNA specific to OPN (OPN shRNA) (50,000 cells per mouse, 8 mice/group, 4 groups). All mice were euthanized on day 14, and the tumor brain tissue, spleen, and blood were collected for ex vivo flow cytometry and immunohistochemical staining. (A) Percentages of CD206+CD11b+ macrophages in the spleen and F4/80+CD11b+ macrophages in the brain tumor were diminished when either OPN in the tumor or in the host background was knocked out. (B) Representative immunohistochemistry showing the frequency of Iba-1+ macrophages in the tumor; the graph summarizes the quantitative results. Original magnification, ×100 (scale bars: 100 μm). (C) Percentage of IFN-γ+CD4+ T cells and IFN-γ+CD8+ effector T cells in the spleen (left), PBMCs (middle), and tumor (right). (D) Percentage of IL-2+CD4+ T cells (left) and IL-2+CD8+ effector T cells (right) in the tumor. (E) Percentage of CD4+FoxP3+ Tregs in PBMCs. Data represent mean ± SD of 4 different animals in 1 experiment. P values were calculated based on 2-tailed 2-sample t test with Bonferroni’s correction. *P < 0.05, **P < 0.01, ***P < 0.001.