Targeting Tumor-Associated Macrophages to Radiosensitize Glioblastoma in a Mouse Model

Abstract

Tumor microenvironments contain immune cells such as macrophages that can be polarized in either a pro- or anti-tumor fashion, including in glioblastoma. Targeting a receptor on macrophages, triggering receptor expressed on myeloid cells 1 (TREM-1), has demonstrated anti-tumor effects in preclinical mouse studies in pancreatic and non-small cell lung cancer. A TREM-1 inhibitory peptide, GF9, has been developed within a blood-brain barrier permeable vehicle that mimics high density lipoproteins (HDL). This peptide was designed to disrupt association of TREM-1 with its signaling adaptor protein DAP12, inhibiting downstream effects, including the recruitment/polarization of pro-tumor macrophages. In this study, we examined the potential of GF9 treatment to sensitize glioblastoma tumors to radiation using a mouse model. In a survival experiment, thirty C57BL/6 mice were implanted with 20,000 GL261 glioblastoma tumor cells intracranially into the right caudate putamen. 4 days after tumor implantation, 10 mice were injected intraperitoneally daily with Tris-buffered saline (TBS/control), 10 mice with HDL (vehicle), and 10 mice with GF9 (2.5 mg/kg) in HDL (GF9-HDL) for 10 days. 7 days after tumor implantation, 5 (half) of the mice of each group were administered 2 Gy of whole brain radiation, one-field PA, every other day to a total dose of 6 Gy. Macrophages were isolated from harvested tumors using anti-F4/80 microbeads and RNA was then isolated. qPCR was performed on a panel of macrophage markers. Median survival of the control mice was 20 days, HDL mice was 22 days, GF9-HDL alone mice was 21 days, TBS+RT and HDL+RT mice was 23 days, and the GF9-HDL+RT combination mice was 26 days. The only significant difference in survival was found between the GF9-HDL+RT combination group and control group with a p=0.04 on a Mantel-Cox log-rank test. qPCR analysis showed a 6-fold decrease of mRNA levels of TGF-β, previously noted to be regulated by TREM-1 signaling in vitro, in tumor-associated macrophages in GF9-treated mice as compared to controls. The results from this preliminary pre-clinical study suggest that TREM-1 blockade using GF9 has the potential to stimulate the therapeutic effects of radiation in a preclinical mouse model of glioblastoma. TGF-β levels in brain tumor-associated macrophages were lowered with GF9 administration consistent with data from in vitro studies of TREM-1 signaling. Additional animal survival studies, serum analysis, and immunohistochemical staining will be performed to further examine the role of tumor-associated macrophages in the radioresistance of glioblastoma tumors.