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ABSTRACTNeural stem cells (NSCs) are multipotent cells of the central nervous system (CNS) capable of self‐renewal, differentiation, and responding to and shaping the surrounding microenvironment. Their continuous crosstalk with surrounding CNS cells is a key component of their therapeutic potential, particularly in tissue repair and regeneration. Communication in the CNS relies on complementary mechanisms, including connexins (Cxs)‐based intercellular communication, to maintain homeostasis and coordinate responses to physiological and pathological stimuli. Itaconate, an endogenous shunt product of the tricarboxylic acid cycle, functions as an immunometabolite involved in inflammation and oxidative stress and has recently been implicated in neuroimmune modulation. Although itaconate influences several signalling cascades and is exchanged between cells and/or released into the extracellular milieu, its effects on Cxs expression in NSCs and whether the modulation of Cxs expression profile represents a driving factor in shaping cell fate remain unclear. Here, we investigated the effect of dimethyl itaconate, a cell‐permeable esterified itaconate derivative, on the expression profile of Cxs in NSCs and its potential to modulate NSCs fate and differentiation. We found that dimethyl itaconate modulates Cxs expression in NSCs, increasing Cx36 levels, and promotes NSCs differentiation toward a neuronal phenotype, while inhibition of Cxs‐based channels with carbenoxolone or mefloquine abolishes these dimethyl itaconate‐induced effects. Collectively, these findings highlight a regulatory role for cell‐permeable itaconate and contribute to the understanding of intercellular communication in the CNS microenvironment, providing insights into potential therapeutic strategies for CNS repair and regeneration. The immunometabolite itaconate modulates the connexin profile of neural stem cells, and it upregulates Cx36‐based channels, promoting the differentiation toward a neuronal phenotype. These effects were abolished by connexin blockade, supporting a role for itaconate in the neuroimmune axis in inflammatory and neurodegenerative disorders.

Pediatric high-grade gliomas (pHGG) and pediatric diffuse midline gliomas (pDMG) are devastating diseases without durable and curative options. Although targeted immunotherapy has shown promise, the field lacks immunocompetent animal models to study these processes in detail. To achieve this, we developed a fully immunocompetent, genetically engineered mouse model (GEMM) for pDMG and pHGG that incorporates the glioma-associated antigen, interleukin 13 receptor alpha 2 (IL13RA2). Utilizing the RCAS-Tva delivery system in Nestin-Tva mice, we induced gliomagenesis by overexpressing PDGFB and deleting p53 (p53fl/fl) or both p53 and PTEN (p53fl/fl PTENfl/fl), with or without IL13RA2 in neonatal mice. De novo tumors developed in models with and without IL13RA2, showing no statistical difference in onset (n = 33, 38 days, p = 0.62). The p53fl/fl PTENfl/fl tumors displayed more aggressive characteristics (n = 12, 31 days). Tumors exhibited features typical of high-grade glioma, including infiltration, pseudopalisading necrosis, and microvascular proliferation. They also showed a high Ki-67 index, variable IL13RA2 expression, a high frequency of CD11b + macrophages, and a low proportion of CD3 + T cells. The model proved effective for evaluating IL13RA2-targeted immunotherapies, with a significant response to CAR T-cell treatment that extended survival (46 days vs. 28 days control; p < 0.0001) and achieved 25% long-term survival in mice. This model facilitates the preclinical assessment of IL13RA2-directed therapies and holds potential for clinical application.Supplementary InformationThe online version contains supplementary material available at 10.1186/s40478-025-01991-4.

Histone H3-mutant gliomas are deadly brain tumors characterized by a dysregulated epigenome and stalled differentiation. In contrast to the extensive datasets available on tumor cells, limited information exists on their tumor microenvironment (TME), particularly the immune infiltrate. Here, we characterize the immune TME of H3.3K27M and G34R/V-mutant gliomas, and multiple H3.3K27M mouse models, using transcriptomic, proteomic and spatial single-cell approaches. Resolution of immune lineages indicates high infiltration of H3-mutant gliomas with diverse myeloid populations, high-level expression of immune checkpoint markers, and scarce lymphoid cells, findings uniformly reproduced in all H3.3K27M mouse models tested. We show these myeloid populations communicate with H3-mutant cells, mediating immunosuppression and sustaining tumor formation and maintenance. Dual inhibition of myeloid cells and immune checkpoint pathways show significant therapeutic benefits in pre-clinical syngeneic mouse models. Our findings provide a valuable characterization of the TME of oncohistone-mutant gliomas, and insight into the means for modulating the myeloid infiltrate for the benefit of patients. Histone H3-mutant gliomas are deadly brain tumours and the tumour microenvironment is not fully understood. Here the authors profile the immune microenvironment from human samples and mouse models and implicate myeloid cells in immune suppression and show inhibition of myeloid cells and checkpoint blockade demonstrates therapeutic benefits in mice.