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.

Analysis of somatic mutations in 131 human brains reveals aging-associated hypermutability.

We analyzed 131 human brains (44 neurotypical, 19 with Tourette syndrome, 9 with schizophrenia, and 59 with autism) for somatic mutations after whole genome sequencing to a depth of more than 200×. Typically, brains had 20 to 60 detectable single-nucleotide mutations, but ~6% of brains harbored hundreds of somatic mutations. Hypermutability was associated with age and damaging mutations in genes implicated in cancers and, in some brains, reflected in vivo clonal expansions. Somatic duplications, likely arising during development, were found in ~5% of normal and diseased brains, reflecting background mutagenesis. Brains with autism were associated with mutations creating putative transcription factor binding motifs in enhancer-like regions in the developing brain. The top-ranked affected motifs corresponded to MEIS (myeloid ectopic viral integration site) transcription factors, suggesting a potential link between their involvement in gene regulation and autism.