Background and aims: Portal hypertension (PH) is a driving force for the progression of chronic liver disease. Complications from PH develop when hepatic venous pressure gradient (HVPG) exceeds 10 mmHg, defining the presence of clinically significant portal hypertension (CSPH). However, measuring HVPG is invasive method, with limited availability. Thus, reliable non-invasive tools would be welcome alternative. Circulating extracellular vesicles (ECV) originating from the cells are valuable source of information pertaining the ongoing pathophysiological process including the molecules which might serve as the biomarkers. In this study we aimed to identify potential new serum biomarkers for CSPH in patients with compensated advanced chronic liver disease (cACLD) by proteomic profiling of serum ECV. Method: Severity of PH was assessed by HVPG measurement that served as the reference standard. Serum samples were pooled based on HVPG measurement in two groups: with and without CSPH. ECV were isolated from the serum pools using ultracentrifugation and vesicle membranes were lysed by sonication. ECV protein cargo was analyzed by Liquid Chromatography-Mass spectrometry (LC-MS). Samples were analyzed in triplicates and proteins identified with at least one peptide were considered relevant for analysis. Functional enrichment analysis of the isolated proteins was conducted using FunRich 3.1.3 analysis tool. Results: A total of 48 patients were included (30 in the CSPH group and 18 in the non-CSPH group, 75% males ; median age: 59, 9 ± 9, 8 years ; majority with alcoholic (48%) and non-alcoholic fatty liver disease (23%)). LC-MS analysis of ECV content resulted in identification of 733 proteins (38 distinctive for CSPH, and 75 for non-CSPH group), that were furtherly classified based on their cellular origin and function. Proteins involved in platelet degranulation, integrinmediated signaling pathway, receptor mediated endocytosis and regulation of cholesterol efflux were more represented, whereas those involved in opsonization, phagocytosis, complement activation, immune and inflammatory response were less represented in the CSPH group. Among the individual proteins that showed the most significant difference between the studied groups phospholipid transfer protein and beta-2- glycoprotein 1 were more represented, whereas complement C1q, C1r and C1s subcomponents and annexin A2 were less represented in the CSPH group. Conclusion: Results of this study provide additional insights into pathophysiological processes taking place along the development of PH in patients with cACLD. Distinctive protein profiles are identified between the patients with respect to the presence of CSPH. Several individual proteins are identified that should be furtherly studied as the potential non-invasive biomarkers of CSPH.