TMIC-45. Hypoxia dysregulates the epigenetic landscape and impairs the functions of glioma-associated myeloid cells

Hypoxia rapidly alters gene expression to allow cellular adaptation to challenging conditions and support tumour growth. Hypoxia also affects the chromatin structure by modification of histones and DNA methylation. Glioblastoma (GBM) is an aggressive, deadly primary brain tumour for which there is no effective treatment. The tumour microenvironment of GBM is highly heterogeneous, with infiltration of glioma-associated microglia and macrophages (GAMs) and the presence of necrotic, hypoxic regions. The mechanisms through which hypoxia alters functions of GAMs remain poorly understood. Here, we show that hypoxia modulates the expression of myeloid markers in the opposite ways: upregulates the monocytic marker Lgals3 and downregulates the microglial markers P2ry12 and Tmem119 in GAMs in vitro and in vivo, as shown using human and mouse GBM single-cell transcriptomics datasets, in vitro co-culture models and CODEX multi-marker proteomic analysis in human adult and paediatric GBM tumour sections. Hypoxia dysregulates additional GAM subtype markers through changes in accessible chromatin, as determined with ATACseq. While hypoxia alone decreases the overall chromatin accessibility at gene promoters, exposure to glioma cells under hypoxic conditions leads to both increases and decreases of chromatin accessibility in promoters of microglial cells. Hypoxia downregulates the chromatin accessibility at the regions enriched in motifs of transcription factors regarded as master regulators of microglial cell identity and function, including SPI1 or IRF8. Moreover, changes in histone acetylation are responsible for Lgals3 upregulation and lipid accumulation in GAMs in hypoxic conditions. Overall, our data highlights the importance of hypoxic stress as a strong intratumoral regulator of myeloid cell functions, which adds complexity to the characterisation of particular GAMs subpopulations. Studies were supported by the National Science Centre (Poland) grant (2019/33/B/NZ1/01556) and the EU Cancer Mission Horizon Europe HIT-GLIO project (101136835) founded by the European Health and Digital Executive Agency.