聚焦超声开放血脑屏障产生独特的组织特异性细胞外囊泡图谱用于胶质母细胞瘤液体活检

Introduction: Extracellular vesicles (EVs) are highly information-dense liquid biopsy reservoirs carrying proteins, RNAs, lipids, and metabolites that reflect the real-time state of their parent cells. In glioblastoma (GBM), EVs offer a uniquely accessible window into an otherwise sequestered compartment, yet the blood brain barrier (BBB) restricts their enrichment and profile within the circulation. Focused ultrasound (FUS) with microbubbles is now a clinically burgeoning method for transient and precise local opening of the BBB, creating an opportunity to interrogate EV flux from tumor and surrounding brain. While FUS-mediated BBB opening (BBBO) has been shown to augment circulating nucleic acids via “sonobiopsy,” its effects on EV release and cargo remain poorly defined. Here, we evaluate how FUS-BBBO alters the abundance and composition of tumor- versus brain-derived EVs enriched from blood plasma in a high-fidelity GBM model. Methods: Orthotopic GBMs were established via intracranial SB28 implantation (n=24). Tumors were screened by contrast-enhanced MRI and randomized into volume-matched sham or FUS treatment groups. BBBO was performed using a neuronavigation-guided preclinical FUS system. Plasma was collected 30 min or 24 h post-treatment. EVs were isolated by ion-exchange chromatography and characterized by NTA and Western blot. Tumor- and brain-derived EVs were enriched using Tumor SPARCs™ and Neuro SPARCs™, respectively, and subjected to DIA-MS proteomic analysis. Results: FUS-BBBO did not significantly alter total plasma EV concentrations associated with either compartment; rather, it induced substantial remodeling of EV cargo. Across samples, >5,000 proteins were identified. Approximately 80% of differentially abundant proteins were unique to either the tumor- or brain-derived EV panel, indicating compartment-specific responses to FUS. Strikingly, FUS exposure yielded 222 and 290 unique proteins within brain-derived and tumor-derived circulating EV populations, respectively. Pathway analysis revealed enrichment in vesicle trafficking, neurovascular stress signaling, cytoskeletal remodeling, and immune regulatory pathways - potentially reflecting FUS-induced alterations in tumor microenvironmental stress, BBB dynamics, and neuro-immune cross-talk. Conclusions: FUS-BBBO drives robust, compartment-specific reprogramming of EV proteomes in GBM, revealing biomarker candidates absent under sham conditions. These findings position EV profiling as a sensitive approach for capturing FUS-induced tissue remodeling and may expand the biomarker repertoire available for spatially selective GBM liquid biopsy. Ongoing studies are assessing temporal dynamics and correspondence with parental tissue proteomes.