CSF1 Overexpression Promotes High-Grade Glioma Formation without Impacting the Polarization Status of Glioma-Associated Microglia and Macrophages.

Current therapies for high-grade gliomas extend survival only modestly. The glioma microenvironment, including glioma-associated microglia/macrophages (GAM), is a potential therapeutic target. The microglia/macrophage cytokine CSF1 and its receptor CSF1R are overexpressed in human high-grade gliomas. To determine whether the other known CSF1R ligand IL34 is expressed in gliomas, we examined expression array data of human high-grade gliomas and performed RT-PCR on glioblastoma sphere-forming cell lines (GSC). Expression microarray analyses indicated that CSF1, but not IL34, is frequently overexpressed in human tumors. We found that while GSCs did express CSF1, most GSC lines did not express detectable levels of IL34 mRNA. We therefore studied the impact of modulating CSF1 levels on gliomagenesis in the context of the GFAP-V12Ha-ras-IRESLacZ (Ras*) model. Csf1 deficiency deterred glioma formation in the Ras* model, whereas CSF1 transgenic overexpression decreased the survival of Ras* mice and promoted the formation of high-grade gliomas. Conversely, CSF1 overexpression increased GAM density, but did not impact GAM polarization state. Regardless of CSF1 expression status, most GAMs were negative for the M2 polarization markers ARG1 and CD206; when present, ARG1(+) and CD206(+) cells were found in regions of peripheral immune cell invasion. Therefore, our findings indicate that CSF1 signaling is oncogenic during gliomagenesis through a mechanism distinct from modulating GAM polarization status. Cancer Res; 76(9); 2552-60. ©2016 AACR.

Sleeping Beauty mouse models identify candidate genes involved in gliomagenesis.

Genomic studies of human high-grade gliomas have discovered known and candidate tumor drivers. Studies in both cell culture and mouse models have complemented these approaches and have identified additional genes and processes important for gliomagenesis. Previously, we found that mobilization of Sleeping Beauty transposons in mice ubiquitously throughout the body from the Rosa26 locus led to gliomagenesis with low penetrance. Here we report the characterization of mice in which transposons are mobilized in the Glial Fibrillary Acidic Protein (GFAP) compartment. Glioma formation in these mice did not occur on an otherwise wild-type genetic background, but rare gliomas were observed when mobilization occurred in a p19Arf heterozygous background. Through cloning insertions from additional gliomas generated by transposon mobilization in the Rosa26 compartment, several candidate glioma genes were identified. Comparisons to genetic, epigenetic and mRNA expression data from human gliomas implicates several of these genes as tumor suppressor genes and oncogenes in human glioblastoma.

CSF1 overexpression has pleiotropic effects on microglia in vivo.

Macrophage colony stimulating factor (CSF1) is a cytokine that is upregulated in several diseases of the central nervous system (CNS). To examine the effects of CSF1 overexpression on microglia, transgenic mice that overexpress CSF1 in the glial fibrillary acidic protein (GFAP) compartment were generated. CSF1 overexpressing mice have increased microglial proliferation and increased microglial numbers compared with controls. Treatment with PLX3397, a small molecule inhibitor of the CSF1 receptor CSF1R and related kinases, decreases microglial numbers by promoting microglial apoptosis in both CSF1 overexpressing and control mice. Microglia in CSF1 overexpressing mice exhibit gene expression profiles indicating that they are not basally M1 or M2 polarized, but they do have defects in inducing expression of certain genes in response to the inflammatory stimulus lipopolysaccharide. These results indicate that the CSF1 overexpression observed in CNS pathologies likely has pleiotropic influences on microglia. Furthermore, small molecule inhibition of CSF1R has the potential to reverse CSF1-driven microglial accumulation that is frequently observed in CNS pathologies, but can also promote apoptosis of normal microglia.