Abstract
Paediatric high-grade gliomas (pHGGs) are aggressive brain tumours and the leading cause of deaths in children. The management of pHGGs is one of the greatest challenges in paediatric oncology as those tumours are considered “cold”, immunosuppressed. Preclinical research is struggling to find new targets with the potential to revolutionise the therapeutic approach. Tumour-host interactions in the brain tumour microenvironment (TME) are poorly understood. In the HIT-GLIO project we aim to perform a comprehensive analysis of pHGG TME, define new immunological targets and compounds/strategies that would change future of those paediatric patients. We combined single-cell (sc)RNseq with 40 immune cell markers, sc-T cell receptor (TCR) profiling, Visium 10X Genomics spatial transcriptomics, multimodal CODEX (Co-detection by indexing) staining and computational approaches to define diverse immune cell types, cellular states and spatial niches that contribute to a cold TME of pHGGs. Thus far, we profiled 180,000 immune cells from biopsies and corresponding cerebrospinal fluids (CSF) of eight pHGGs, 4 diffuse midline gliomas (DMGs). We found that CD45+ cells in pHGG tissue samples predominantly comprise of CD11b+/TMEM119 microglia with less frequent immunosuppressive CD68+Gal3+macrophages, with very few CD3+ T lymphocytes. pHGG-associated microglia express transcriptomic programs related to ECM remodeling and angiogenesis, with low expression of chemokines and cytokines. Tumour-associated monocytes show low expression of inflammatory factors, and macrophages display phagocytic and immunosuppressive phenotypes. Analysis of paired scRNA/TCR sequencing data revealed the combinatorial impact of TCR utilization on phenotypic diversity and clonality of T cells in CSF and tumours. Overall, our data demonstrate the abundance of microglia, low inflammatory and chemotactic signature of pHGGs which may contribute to the lack of lymphocytes and the immunosuppressive phenotype of pHGG-associated microglia and macrophages. Studies supported by the EU Cancer Mission Horizon Europe HIT-GLIO project (101136835) founded by the European Health and Digital Executive Agency.