CHD2 Regulates Neuron-glioma Interactions in Pediatric Glioma.

High-grade gliomas (HGG) are deadly diseases for both adult and pediatric patients. Recently, it has been shown that neuronal activity promotes progression of multiple subgroups of HGG. However, epigenetic mechanisms that govern this process remain elusive. Here we report that the chromatin remodeler CHD2 regulates neuron-glioma interactions in diffuse midline glioma (DMG) characterized by onco-histone H3.1K27M. Depletion of CHD2 in H3.1K27M DMG cells compromises cell viability and neuron-to-glioma synaptic connections in vitro, neuron-induced proliferation of H3.1K27M DMG cells in vitro and in vivo, activity-dependent calcium transients in vivo, and extends the survival of H3.1K27M DMG-bearing mice. Mechanistically, CHD2 coordinates with the transcription factor FOSL1 to control the expression of axon-guidance and synaptic genes in H3.1K27M DMG cells. Together, our study reveals a mechanism whereby CHD2 controls the intrinsic gene program of the H3.1K27M DMG subtype, which in turn regulates the tumor growth-promoting interactions of glioma cells with neurons.

Focused ultrasound-mediated blood-brain barrier opening is safe and feasible with moderately hypofractionated radiotherapy for brainstem diffuse midline glioma.

BACKGROUND: Diffuse midline glioma (DMG) is a pediatric tumor with dismal prognosis. Systemic strategies have been unsuccessful and radiotherapy (RT) remains the standard-of-care. A central impediment to treatment is the blood-brain barrier (BBB), which precludes drug delivery to the central nervous system (CNS). Focused ultrasound (FUS) with microbubbles can transiently and non-invasively disrupt the BBB to enhance drug delivery. This study aimed to determine the feasibility of brainstem FUS in combination with clinical doses of RT. We hypothesized that FUS-mediated BBB-opening (BBBO) is safe and feasible with 39 Gy RT. METHODS: To establish a safety timeline, we administered FUS to the brainstem of non-tumor bearing mice concurrent with or adjuvant to RT; our findings were validated in a syngeneic brainstem murine model of DMG receiving repeated sonication concurrent with RT. The brainstems of male B6 (Cg)-Tyrc-2J/J albino mice were intracranially injected with mouse DMG cells (PDGFB+, H3.3K27M, p53-/-). A clinical RT dose of 39 Gy in 13 fractions (39 Gy/13fx) was delivered using the Small Animal Radiation Research Platform (SARRP) or XRAD-320 irradiator. FUS was administered via a 0.5 MHz transducer, with BBBO and tumor volume monitored by magnetic resonance imaging (MRI). RESULTS: FUS-mediated BBBO did not affect cardiorespiratory rate, motor function, or tissue integrity in non-tumor bearing mice receiving RT. Tumor-bearing mice tolerated repeated brainstem BBBO concurrent with RT. 39 Gy/13fx offered local control, though disease progression occurred 3-4 weeks post-RT. CONCLUSION: Repeated FUS-mediated BBBO is safe and feasible concurrent with RT. In our syngeneic DMG murine model, progression occurs, serving as an ideal model for future combination testing with RT and FUS-mediated drug delivery.

Immune Response following FLASH and Conventional Radiation in Diffuse Midline Glioma.

PURPOSE: Diffuse midline glioma (DMG) is a fatal tumor traditionally treated with radiation therapy (RT) and previously characterized as having a noninflammatory tumor immune microenvironment (TIME). FLASH is a novel RT technique using ultra-high dose rate that is associated with decreased toxicity and effective tumor control. However, the effect of FLASH and conventional (CONV) RT on the DMG TIME has not yet been explored. METHODS AND MATERIALS: Here, we performed single-cell RNA sequencing (scRNA-seq) and flow cytometry on immune cells isolated from an orthotopic syngeneic murine model of brainstem DMG after the use of FLASH (90 Gy/sec) or CONV (2 Gy/min) dose-rate RT and compared to unirradiated tumor (SHAM). RESULTS: At day 4 post-RT, FLASH exerted similar effects as CONV in the predominant microglial (MG) population, including the presence of two activated subtypes. However, at day 10 post-RT, we observed a significant increase in the type 1 interferon α/ß receptor (IFNAR+) in MG in CONV and SHAM compared to FLASH. In the non-resident myeloid clusters of macrophages (MACs) and dendritic cells (DCs), we found increased type 1 interferon (IFN1) pathway enrichment for CONV compared to FLASH and SHAM by scRNA-seq. We observed this trend by flow cytometry at day 4 post-RT in IFNAR+ MACs and DCs, which equalized by day 10 post-RT. DMG control and murine survival were equivalent between RT dose rates. CONCLUSIONS: Our work is the first to map CONV and FLASH immune alterations of the DMG TIME with single-cell resolution. Although DMG tumor control and survival were similar between CONV and FLASH, we found that changes in immune compartments differed over time. Importantly, although both RT modalities increased IFN1, we found that the timing of this response was cell-type and dose-rate dependent. These temporal differences, particularly in the context of tumor control, warrant further study.