Abstract
IntroductionLiquid biopsy using circulating microvesicles and exosomes is emerging as a new diagnostic tool that could improve hepatocellular carcinoma (HCC) early diagnosis and screening protocols. Our study aimed to investigate the utility of plasma exosomal miR-21-5p and miR-92-3p for HCC diagnosis during screening protocols.MethodsThe study group included 106 subjects: 48 patients diagnosed with HCC during screening, who underwent a potentially curative treatment (surgical resection or liver transplantation), 38 patients with liver cirrhosis (LC) on the waiting list for liver transplantation, and 20 healthy volunteers. The exosomes were isolated by precipitation with a reagent based on polyethylene glycol and were characterized based on morphological aspects (i.e., diameter); molecular weight; CD63, CD9, and CD81 protein markers; and exosomal miR-21-5p and miR-92a-3p expression levels.ResultsWe first demonstrate that the exosome population isolated with the commercially available Total Exosome Isolation kit respects the same size ranging, morphological, and protein expression aspects compared to the traditional ultracentrifugation technique. The analysis of the expression profile indicates that miR-21-5p was upregulated (p = 0.017), and miR-92a-3p was downregulated (p = 0.0005) in plasma-derived exosomes from HCC subjects, independently from the patient’s characteristics. AUROC for HCC diagnosis based on AFP (alpha-fetoprotein) was 0.72. By integrating AFP and the relative expression of exosomal miR-21-5p and miR-92a-3p in a logistic regression equation for HCC diagnosis, the combined AUROC of the new exosomal miR HCC score was 0.85—significantly better than serum AFP alone (p = 0.0007).ConclusionTogether with serum AFP, plasma exosomal miR-21-5p and miR-92a-3p could be used as potential biomarkers for HCC diagnosis in patients with LC subjected to screening and surveillance.
Highlights
Liquid biopsy using circulating microvesicles and exosomes is emerging as a new diagnostic tool that could improve hepatocellular carcinoma (HCC) early diagnosis and screening protocols
The patients were divided into three groups: 48 patients diagnosed with HCC during the screening program, who underwent a potentially curative treatment at the Digestive Diseases and Liver Transplantation Center, Fundeni Clinical Institute; 38 patients included on the waiting list for liver transplantation for liver cirrhosis (LC) and, during the screening program for HCC; and as a control group (C) 20 healthy volunteers
Peripheral blood samples from all the subjects were collected in 5-mL vacutainer (Becton Dickinson) tubes spray-coated with EDTA as anticoagulant and centrifuged at 4200 × rpm for 10 min at room temperature (RT) to separate the plasma fraction
Summary
Liquid biopsy using circulating microvesicles and exosomes is emerging as a new diagnostic tool that could improve hepatocellular carcinoma (HCC) early diagnosis and screening protocols. Our study aimed to investigate the utility of plasma exosomal miR-21-5p and miR-92-3p for HCC diagnosis during screening protocols. HCC detection at an early stage is challenging as the disease usually progresses asymptomatically. Screening programs are used for cancer detection in patients at risk for tumor development and usually include ultrasound (US) with or without serum biomarkers, such as alpha-fetoprotein (AFP) (Galle et al, 2018; Heimbach et al, 2018). Serum AFP levels exhibit a low sensitivity for early HCC diagnosis, ranging from 20 to 65% (El-Serag and Davila, 2011; Tian et al, 2017)
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