Optimization of an Extracellular Vesicle Isolation and microRNA Profiling Pipeline for Early Detection of Hepatocellular Carcinoma

Early detection of hepatocellular carcinoma (HCC) remains a major clinical challenge, limiting curative treatment options. Extracellular vesicles (EVs) circulating in plasma carry enriched microRNA (miRNA) cargo that reflects the pathophysiology of HCC and hold promise as minimally invasive biomarkers. In this study, we developed and optimized a pipeline for EV isolation from human plasma, followed by miRNA profiling, aiming to maximize yield, purity, and diagnostic sensitivity for early‐stage HCC. We compared different EV isolation methods (ultracentrifugation, precipitation, immuno‐capture) and evaluated downstream miRNA extraction and qPCR/NGS profiling workflows. In a pilot cohort of early‐stage HCC patients (n = ۳۰) versus cirrhosis controls (n = ۳۰), we identified a panel of EV‐miRNAs (miR-۲۱-۵p, miR-۹۲a-۳p, miR-۱۹-۳p) whose expression discriminated HCC from controls (AUC ~۰.۸۵) when the optimized pipeline was used. Our results demonstrate that careful methodological optimization enhances the performance of EV-miRNA-based liquid biopsy for HCC, and support further large‐scale validation.

کلیدواژه ها:

extracellular vesicles ، microRNA ، hepatocellular carcinoma ، liquid biopsy ، early detection