DDEL-30. Multi-Omic Profiling of Human Peritumoral Brain Following Ultrasound-Mediated BBB Disruption Reveals Microglial Reprogramming and Enhanced Endothelial Crosstalk

Abstract Low-intensity pulsed-ultrasound coupled with microbubbles can transiently permeabilize the blood–brain barrier (BBB) and is being explored as a drug-delivery approach for treatment of glioblastoma (GBM). We hypothesized that sonication would not only facilitate BBB permeability but also reprogram peritumoral microglia toward a pro-repair, chemoattractant state. In a cohort of 13 recurrent GBM patients, we implanted a SonoCloud-9 ultrasound device, sonicated immediately before resection, and harvested paired sonicated and non-sonicated peritumoral biopsies 45 minutes later. We previously demonstrated that the BBB reseals within one hour, coinciding with rapid microglial recruitment and upregulation of motility and wound-healing programs (P<0.05). In this study, we build on those observations by applying multi-omic profiling—single-cell RNA sequencing (n = 7 patients), spatial transcriptomics (n = 2 patients), and integrated single-cell ATAC+RNA sequencing (n = 2 patients)—to map how sonication reshapes intercellular communication and chromatin accessibility in the peritumoral niche. Using cell–cell interactome modeling, we discovered a marked increase in VEGFA, IL1, and IL-6 signaling between microglia and endothelial cells following sonication (P<0.05). Spatial transcriptomics validated an increased interaction between endothelium and pro-inflammatory microglia. Integrated single-cell ATAC + RNA sequencing showed increased chromatin accessibility of the immune checkpoints TNFRSF18 (GITR) and PDCD1 (PD-1) together with AP-1 transcription-factor motif enrichment (P<0.05), linking sonicated microglia to a phagocytic yet immunoregulatory phenotype. Gene-ontology analysis of genes with increased chromatin accessibility by sonication revealed up-regulation of “chemotaxis” and “cell migration” pathways (P<0.05). These findings define a therapeutically actionable window in which ultrasound not only permits drug entry but also reprograms the innate immune niche toward a chemoattractant and acute repair state in the human brain. This provides a framework for optimizing the timing and selection of vascular-targeted or immunomodulatory agents and informing the design of future ultrasound clinical trial for GBM patients.