Endocrine Abstracts

Hepatocellular carcinoma (HCC) is an aggressive tumour frequently associated to an underlying chronic liver disease. In this sense, metabolic dysfunction-associated steatotic liver disease (MASLD) is considered as a growing cause of HCC development. Considering the need of novel therapeutic approaches, and the promising data about targeting the tumour microenvironment in cancer, we performed quantitative proteomics on HCC samples to characterize the components and regulators of the extracellular matrix (the matrisome). The cytosolic and nuclear proteomes of hepatic tissues of HCC patients [n=42; HCC vs non-tumour adjacent tissue (NTAT)] were determined using SWATH-MS. The abundance of matrisome proteins was evaluated in the proteomic HCC cohort, two retrospective HCC mRNA cohorts (n=151), and different in silico cohorts (8 HCC, 10 MASLD and cirrhosis; mRNA and protein). Functional assays (proliferation, migration, colony/tumorspheres formation) were performed in 2 HCC cell lines (Hep3B, SNU-387) after silencing/overexpressing SERPINF2. Finally, in silico scRNAseq cohorts of MASLD and non-MASLD-derived HCC were analysed. Through quantitative proteomics we identified 34 (cytosol) and 61 (nucleus) differentially expressed matrisome proteins (HCC vs NTAT). A broad dysregulation of the serpins family of protease inhibitors was corroborated in most MASLD, cirrhosis and HCC in silico cohorts, confirming their dysregulation in advanced stages of chronic liver disease and in HCC, wherein they were associated with aggressiveness and altered metabolic status. SERPINF2 was consistently dysregulated in most cohorts and associated to metabolic alterations (BMI, leptin/adiponectin ratio) and aggressiveness (survival, microinvasion). SERPINF2 silencing in HCC cell lines reduced all the functional parameters evaluated, while SERPINF2 overexpression had the opposite effect. Interestingly, the supernatant of SERPINF2-overexpressing cell lines did not affect proliferation, suggesting different roles of intracellular and extracellular SERPINF2. Finally, the analysis of HCC single-cell-RNAseq in silico cohorts revealed a decreased expression of SERPINF2 in tumour-associated endothelial cells and an increased expression in tumour cells. The comparison of cell populations with high or low SERPINF2 expression also pointed to an alteration of cell-to-cell signalling including angiogenic (i.e. VEGF) and immune-associated (i.e. MIF) factors, consistently with the associations observed in in silico RNAseq and with the positive correlation between SERPINF2 expression and the infiltration score in HCC patients. Taken together, we propose a potential role of serpins in the clinical management of chronic liver disease. Specially, SERPINF2 could represent a relevant tool for targeting HCC tumour cells and their microenvironment.

Fundings: ISCIII (PI20/01301, PI23/00652; co-funded by the European Union), MINECO (FPU20/03957), JdA (PEMP-0036-2020, BIO-0139), FSEEN and CIBERobn/ehd.