Immune Response following FLASH and Conventional Radiation in Diffuse Midline Glioma (DMG)

Abstract

Purpose Diffuse Intrinsic Pontine Glioma is a fatal tumor traditionally treated with radiotherapy (RT) and previously characterized as having a non-inflammatory tumor immune microenvironment (TIME). FLASH is a novel RT technique using an ultra-fast dose-rate which is associated with decreased toxicity, effective tumor control and potential immune-sparing properties. However, the effect of FLASH on the DIPG tumor immune microenvironment (TIME) has not yet been explored. Methods Here, we perform single-cell RNA sequencing on immune cells isolated from an orthotopic syngeneic murine model of DIPG following the use of FLASH (90Gy/sec) or conventional (2Gy/min) dose-rate RT (CONV-RT), and compare to unirradiated tumor and normal brainstem. Results Sequencing of immune cells reveals 17 unique populations, most abundant of which is microglia. In the most activated microglia subtypes, both CONV-RT and FLASH show upregulation of type 1 interferon (IFN1) genes and pathway scores compared to unirradiated tumor. In macrophages (MACs) and dendritic cells (DCs), CONV-RT is significantly enriched for IFN1 while this response is less seen with FLASH. Further, FLASH shows an increase in CNS border-associated MACs and upregulation of a myeloid-derived suppressor cell (MDSC) signature in MONOs, less seen with CONV-RT. In the lymphocytes, FLASH yields a higher mature B cell proportion and upregulation of T-cell activation and trafficking markers compared to CONV-RT. Finally, we correlate our data with myeloid cells from cerebrospinal fluid of human DIPG patients and find overlap with our murine tumor- and treatment-associated markers. Conclusion Our work is the first to map CONV-RT and FLASH immune alterations with single-cell resolution in the DIPG TIME. We find that CONV-RT and FLASH sculpt the microglial compartment similarly while recruiting distinct non-resident myeloid subsets and mature B-cell fractions, highlighting the potential to combine each modality with unique immunotherapy regimens in this fatal disease.

Smart Radiotherapy Biomaterials for Image-Guided In Situ Cancer Vaccination

“…This approach has garnered significant recent attention due to its normal tissue sparing effects and represents a possible avenue for RT dose-escalation or dose-painting that merits further investigation [ 64 ]. Moreover, recent work has suggested that FLASH RT may also have immunologic implications, with two recent reports suggesting that FLASH RT can upregulate T-cell activating and trafficking markers in both glioblastoma and diffuse pontine glioma models [ 65 , 66 ].…”