Tumor cell extracellular vesicles derived lncRNA XIST regulates the blood-brain barrier and NSCLC brain metastasis via miR-19b-3p/EPN2.

This study aimed to investigate the function and molecular mechanism of long non-coding RNA XIST derived from tumor extracellular vesicles (EVs) in brain metastasis of non-small cell lung cancer (NSCLC). Serum samples were collected from NSCLC patients with brain metastasis and healthy controls to detect the expression levels of XIST, miR-19b-3p, and EPN2 mRNA. EVs were isolated from A549 cells, and their morphology and particle size were characterized using transmission electron microscopy (TEM), nanoparticle tracking analysis (NTA), and western blot analysis. An in vitro blood-brain barrier (BBB) model was constructed using human brain microvascular endothelial cells (HBMECs) and astrocytes. After a 24-h treatment with EVs, PKH67 staining was employed to observe EV uptake by HBMECs and astrocytes. The role of XIST in BBB integrity and NSCLC cell invasion was evaluated through RT-qPCR, Western blot, transendothelial electrical resistance (TEER) measurement, rhodamine-dextran permeability assay, immunofluorescence staining, Transwell migration assay, and interactive video imaging system. The binding interactions among XIST, miR-19b-3p, and EPN2 were confirmed by dual-luciferase reporter assay and RNA immunoprecipitation. Results revealed that EVs derived from A549 cells exhibited typical characteristics and were efficiently internalized by HBMECs, leading to reduced TEER values, decreased tight junction proteins expression, and increased rhodamine-dextran permeability. XIST was overexpressed in NSCLC patients with brain metastasis and in A549 and H1299 cells, and its knockdown in A549 cells alleviated BBB disruption and suppressed A549 and H1299 cell invasion. Knockdown of EPN2 in HBMECs produced effects similar to those observed with XIST knockdown. Notably, knockdown of miR-19b-3p partially reversed the effects of EPN2 knockdown on both BBB integrity and NSCLC cell invasion. In vivo experiments demonstrated that XIST inhibition attenuated NSCLC brain metastasis by improving BBB integrity. Mechanistically, XIST positively regulated EPN2 expression by sponging miR-19b-3p. In conclusion, inhibition of XIST in A549 cells mitigates BBB damage and suppresses NSCLC brain metastasis by modulating the miR-19b-3p/EPN2 axis.