Glycosylated Hypoxic Tumor-Derived Extracellular Vesicles as a Targeted siRNA Delivery System to Remodel the Immunosuppressive Microenvironment in Pancreatic Cancer.

Abstract

GalNAc-modified extracellular vesicles (EVs) derived from hypoxic pancreatic cancer cells and loaded with ANXA2 siRNA were developed to modulate the immunosuppressive tumor microenvironment (TME). Characterization confirmed successful surface modification and efficient siRNA loading in EVs with a diameter of approximately 100-120 nm. In vitro, these engineered EVs inhibited pancreatic cancer cell proliferation, migration, and invasion and promoted apoptosis. Additional cargo-matched comparison showed that GalNAc-modified hypoxic EVs carrying ANXA2 siRNA produced stronger ANXA2 silencing and more pronounced anti-migratory, anti-invasive, and pro-apoptotic effects than unmodified hypoxic EVs carrying the same siRNA, whereas GalNAc modification alone did not show an obvious independent antitumor effect. In a xenograft model, treatment reduced tumor volume by approximately 50% at week 4 and prolonged median survival from 45 to 60 days. Multi-omics and immune analyses suggested suppression of the PI3K/AKT signaling pathway and remodeling of the immunosuppressive TME, as reflected by increased CD8+ T-cell infiltration, reduced tissue-associated immunoreactivity for IL-6, IL-10, and TNF-α, and increased soluble IFN-γ, IL-2, and GM-CSF levels. These findings indicate that GalNAc-modified hypoxic EVs carrying ANXA2 siRNA can inhibit tumor progression and modulate the TME in pancreatic cancer, supporting their potential as a therapeutic delivery platform.