Brain-derived extracellular vesicle profiling in Down syndrome-associated Alzheimer’s disease

Abstract Background Brain-derived extracellular vesicles (BDEVs) offer a non-invasive window into central nervous system pathology, but their molecular characterization across the Down syndrome (DS)—associated Alzheimer’s disease continuum (DSAD) remains unexplored. Methods We enriched plasma neuron- and astrocyte-derived EVs (NDEVs and ADEVs) from 57 individuals: 14 controls and 43 individuals with DS (20 asymptomatic and 23 symptomatic for AD). The ultrasensitive 127-plex CNS-NULISAseq panel was applied to profile these BDEVs subpopulations. We benchmarked NULISAseq analytical sensitivity against the Olink platform in ADEVs, characterized BDEV differential proteomic profiles across clinical groups, and correlated the findings with established plasma AD biomarkers (pTau217, NfL, and GFAP). Results NULISAseq demonstrated superior analyte detectability in ADEVs (87.4%) compared to the 11.9% observed in a previous Olink-based study using the same ADEV isolation protocol in an independent cohort. BDEV characterization revealed signatures of neurodegeneration, inflammation, and vascular/metabolic stress. Notably, FLT1 in ADEVs, and CXCL10, NEFH, PGK1, and NPY in NDEVs, were elevated in DS and significantly correlated with established plasma AD biomarkers. Moreover, group comparisons revealed stage-associated increases in these NDEV markers across the DSAD continuum. Conclusions This study supports multiplex profiling of BDEVs as a non-invasive window into central nervous system biology, providing biological information that complements current plasma biomarkers and offering novel insights into the molecular mechanisms driving DSAD.