2574 - Impact of Tumor-Associated Macrophage/Microglia Markers on Survival Outcome in Lower Grade Glioma Patients

W. Meng¹, X. Meng¹, H. Yamaguchi¹, E. Sebastian², K. Singh³, J. L. Fleming³, J. McElroy⁴, A. Grosu⁵, J. Haque⁶, and A. Chakravarti¹; ¹Department of Radiation Oncology, The James Cancer Center, Ohio State University Wexner Medical Center, Columbus, OH, ²Department of Radiation Oncology, Corewell Health William Beaumont University Hospital, Royal Oak, MI, ³The Ohio State University Comprehensive Cancer Center, Columbus, OH, ⁴The Ohio State University, Center for Biostatistics, Department of Biomedical Informatics, Columbus, OH, ⁵Department of Radiation Oncology, Medical Center - University of Freiburg, Faculty of Medicine, Freiburg, Germany, ⁶Department of Radiation Oncology, The Ohio State University Wexner Medical Center, Columbus, OH

Purpose/Objective(s): Glioma often displays resistance to standard temozolomide plus radiation therapy. The classification of gliomas into high-grade and low-grade glioma (LGG) based on IDH1/2 mutation status emphasizes the urgency to untangle the molecular complexities driving LGG progression. Our preliminary investigation revealed a correlation between the promoter methylation of the DNAX-activating protein of 12 kDa (DAP12), an adaptor protein of triggering receptor expressed on myeloid cells 2 (TREM-2), and survival outcomes in LGG patients. This study aims to investigate the role of DAP12 in association with survival outcomes in LGG patients and its regulatory mechanisms in the tumor microenvironment. Materials/

Methods: The clinical correlative study evaluated the overall survival (OS) benefits associated with the methylation status of the DAP12 promoter in two Infinium Methylation 450K array datasets. Further investigation explored the Pearson correlation between DAP12 gene expression and promoter methylation in the TCGA LGG cohort. Additionally, the role of TREM2/DAP12 as markers on tumor-associated macrophages/microglia (TAM) and their association with TAM polarization was examined using publicly available single-cell sequencing dataset and in vitro assays. The liquid chromatography–mass spectrometry secretome analysis used THP1 cells co-cultured with U87 IDH-mutant cells to investigate the crosstalk between glioma cells and TAMs in the tumor microenvironment.
Results: The clinical datasets revealed that promoter methylation regulation significantly suppressed DAP12 RNA expression (Pearson Correlation r = -0.39, P<0.001) in the TCGA LGG cohort, correlating with the better overall survival of LGG patients. Single-cell sequencing data and western blot identified the TREM2/DAP12 complex as a specific marker on TAMs, with their activation linked to TAM M2 polarization. Our results indicated that 2-hydroxyglutarate adversely affects DAP12 expression in vitro. The secretome analysis from the co-culture in vitro experiment demonstrated that the glioma microenvironment dramatically induced the activation of TREM2/DAP12 complex as well as angiogenesis, inflammation, chemotaxis, and invasion pathways.
Conclusion: These findings highlight the significance of DAP12 in LGG progression and its potential as a prognostic marker. Understanding the regulatory mechanisms and functional implications of TREM2/DAP12 activation sheds light on the avenues for further research and therapeutic interventions. Abstract 2574 – Table 1: Demographic and clinical characteristics

Characteristic	TCGA LGG (N=530)	University of Freiburg (N=61)
Median age (range)	41 (14-87)	41 (21-75)
Female sex, no. (%)	238 (45)	18 (30)
Male sex, no. (%)	291 (55)	42 (70)
Histology, no. (%)
Astrocytoma	197 (37)	11 (18)
Oligoastrocytoma	134 (25)	13 (21)
Oligodendroglioma	198 (38)	35 (58)
NA	1 (0)	2 (3)
IDH mutation, no. (%)
IDH mutant	91 (17)	53 (87)
IDH wild-type	34 (6)	8 (13)
NA	405 (77)	0 (0)