Targeting CXCR4 by a selective peptide antagonist modulates tumor microenvironment and microglia reactivity in a human glioblastoma model

Abstract

BackgroundThe CXCL12/CXCR4 pathway regulates tumor cell proliferation, metastasis, angiogenesis and the tumor-microenvironment cross-talk in several solid tumors, including glioblastoma (GBM), the most common and fatal brain cancer. In the present study, we evaluated the effects of peptide R, a new specific CXCR4 antagonist that we recently developed by a ligand-based approach, in an in vitro and in vivo model of GBM. The well-characterized CXCR4 antagonist Plerixafor was also included in the study.MethodsThe effects of peptide R on CXCR4 expression, cell survival and migration were assessed on the human glioblastoma cell line U87MG exposed to CXCL12, by immunofluorescence and western blotting, MTT assay, flow cytometry and transwell chamber migration assay. Peptide R was then tested in vivo, by using U87MG intracranial xenografts in CD1 nude mice. Peptide R was administered for 23 days since cell implantation and tumor volume was assessed by magnetic resonance imaging (MRI) at 4.7 T. Glioma associated microglia/macrophage (GAMs) polarization (anti-tumor M1 versus pro-tumor M2 phenotypes) and expressions of vascular endothelial growth factor (VEGF) and CD31 were assessed by immunohistochemistry and immunofluorescence.ResultsWe found that peptide R impairs the metabolic activity and cell proliferation of human U87MG cells and stably reduces CXCR4 expression and cell migration in response to CXCL12 in vitro. In the orthotopic U87MG model, peptide R reduced tumor cellularity, promoted M1 features of GAMs and astrogliosis, and hindered intra-tumor vasculature.ConclusionsOur findings suggest that targeting CXCR4 by peptide R might represent a novel therapeutic approach against GBM, and contribute to the rationale to further explore in more complex pre-clinical settings the therapeutic potential of peptide R, alone or in combination with standard therapies of GBM.Electronic supplementary materialThe online version of this article (doi:10.1186/s13046-016-0326-y) contains supplementary material, which is available to authorized users.

Phosphatidylcholine-specific phospholipase C inhibition down- regulates CXCR4 expression and interferes with proliferation, invasion and glycolysis in glioma cells

“…Plerixafor strongly decreased the CXCR4 expression on cell membrane of U87MG cells treated for 24h compared to untreated (CTRL) cells (Fig 1C, green fluorescence) as already reported [11]. However, at 48h of treatment with this antagonist the CXCR4 receptor was already re-expressed on cell membrane and its level was further increased at 72h (Fig 1C).…”

“…Furthermore, we recently investigated the role of CXCR4 in glioma-microenvironment interactions and demonstrated that reducing CXCR4 expression in human U87MG cells promoted M1 features in microglia/macrophages recruited to the tumor area, creating an environment potentially less favorable for tumor growth [11]. …”

Glioma associated microglia/macrophages, a potential pharmacological target to promote antitumor inflammatory immune response in the treatment of glioblastoma

“…These beneficial effects relied on reduced GAMs' infiltration at the tumor site and increased antitumor activities [63] . Inhibition of the C-X-C chemokine receptor type 4 (CXCR4) by a newly synthetized receptor antagonist, peptide R, reduced tumor growth, glioma cell invasiveness, and intratumor vessel formation while directing GAMs' immune activation toward a pro-inflammatory/antitumor phenotype [64] . Notably, SDF-1 suppression in a murine glioma resulted in delayed tumor growth and invasiveness, lower microvascular density, and higher density of microglia/macrophages in non-hypoxic compared to hypoxic regions.…”

FTY720 Exerts Anti-Glioma Effects by Regulating the Glioma Microenvironment Through Increased CXCR4 Internalization by Glioma-Associated Microglia

“…CXCR4, an important player in supportive interactions between tumor cells and other cells, participates in tumor cell proliferation, metastasis, angiogenesis, and the tumor microenvironment cross-talk in GBM. Studies have revealed that targeting the CXCR4 pathway may provide a therapeutic approach against glioma (12,38). Our data suggested that FTY720 can block the cross-talk between glioma and GAMs by increasing microglial CXCR4 internalization rather than by affecting CXCL12 secretion by glioma.…”