A functional subpopulation of human glioma associated macrophages linked to malignant glioma progression -RNA-seq

Malignant gliomas are progressive brain cancers with poor prognosis. Pro-tumorigenic glioma associated macrophages (GAM) have been implicated in disease progression however identification of pathogenic functional subsets is lacking. Macrophage functional specification is driven by transcription factor-associated gene regulatory networks, yet the core regulatory transcription factors that govern GAM functions remain unclear. Here we apply single cell RNA/ATAC sequencing to derive the imprint of the glioma tumor microenvironment on GAM transcriptional program specification and identify cell surface markers to prospectively isolate a functionally distinct subpopulation of GAMs in human samples present high grade tumors irrespective of IDH mutation status. This subset of GAMs, termed malignancy associated GAMs (mGAMs) spatially localize to hypoxic metabolic niches and possess a multitude of pro-tumorigenic functions. mGAMs also share somatic mutations with monocytes, suggesting a common bone marrow origin. mGAMs therefore represent a functional subset of GAMs in humans and a potential therapeutic target. Overall design: Patient-derived PBMCs from non-glioma patients are sorted for CD14+ monocytes through magnetic-activated cell sorting (MACS) using CD14 MicroBeads (Miltenyi Biotech, Germany). These monocytes are then co-cultured alongside a previously established patient-derived GBM cell line, MSK-103. Following the initial three-day co-culture, hypoxia was induced by culturing the cells under 0.5% O2 for 4 hours. Cells from monocultures and co-cultures at normoxic and hypoxic conditions are then collected, stained for CD45 (BioLegend, USA) and sorted through FACS