Accurate liquid biopsy for the diagnosis of non-Alcoholic steatohepatitis and liver fibrosis

Giulia Angelini, Simona Panunzi, Lidia Castagneto-Gissey, Francesca Pellicanò, Andrea De Gaetano, Maurizio Pompili, Laura Riccardi, Matteo Garcovich, Marco Raffaelli, Luigi Ciccoritti, Ornella Verrastro, Maria Francesca Russo, Fabio Maria Vecchio, Giovanni Casella, James Casella-Mariolo, Luigi Papa, Pier Luigi Marini, Francesco Rubino, Carel W. Le Roux, Stefan BornsteinGeltrude Mingrone*

*Corresponding author for this work

Research output: Contribution to journalArticlepeer-review

22 Citations (Scopus)
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Objective: Clinical diagnosis and approval of new medications for non-Alcoholic steatohepatitis (NASH) require invasive liver biopsies. The aim of our study was to identify non-invasive biomarkers of NASH and/or liver fibrosis. Design: This multicentre study includes 250 patients (discovery cohort, n=100 subjects (Bariatric Surgery Versus Non-Alcoholic Steato-hepatitis-BRAVES trial); validation cohort, n=150 (Liquid Biopsy for NASH and Liver Fibrosis-LIBRA trial)) with histologically proven non-Alcoholic fatty liver (NAFL) or NASH with or without fibrosis. Proteomics was performed in monocytes and hepatic stellate cells (HSCs) with iTRAQ-nano-Liquid Chromatography-Mass Spectrometry/Mass Spectrometry (LC-MS/MS), while flow cytometry measured perilipin-2 (PLIN2) and RAB14 in peripheral blood CD14+CD16- monocytes. Neural network classifiers were used to predict presence/absence of NASH and NASH stages. Logistic bootstrap-based regression was used to measure the accuracy of predicting liver fibrosis. Results: The algorithm for NASH using PLIN2 mean florescence intensity (MFI) combined with waist circumference, triglyceride, alanine aminotransferase (ALT) and presence/absence of diabetes as covariates had an accuracy of 93% in the discovery cohort and of 92% in the validation cohort. Sensitivity and specificity were 95% and 90% in the discovery cohort and 88% and 100% in the validation cohort, respectively. The area under the receiver operating characteristic (AUROC) for NAS level prediction ranged from 83.7% (CI 75.6% to 91.8%) in the discovery cohort to 97.8% (CI 95.8% to 99.8%) in the validation cohort. The algorithm including RAB14 MFI, age, waist circumference, high-density lipoprotein cholesterol, plasma glucose and ALT levels as covariates to predict the presence of liver fibrosis yielded an AUROC of 95.9% (CI 87.9% to 100%) in the discovery cohort and 99.3% (CI 98.1% to 100%) in the validation cohort, respectively. Accuracy was 99.25%, sensitivity 100% and specificity 95.8% in the discovery cohort and 97.6%, 99% and 89.6% in the validation cohort. This novel biomarker was superior to currently used FIB4, non-Alcoholic fatty liver disease fibrosis score and aspartate aminotransferase (AST)-To-platelet ratio and was comparable to ultrasound two-dimensional shear wave elastography. Conclusions: The proposed novel liquid biopsy is accurate, sensitive and specific in diagnosing the presence and severity of NASH or liver fibrosis and is more reliable than currently used biomarkers. Clinical trials: Discovery multicentre cohort: Bariatric Surgery versus Non-Alcoholic Steatohepatitis, BRAVES, identifier: NCT03524365. Validation multicentre cohort: Liquid Biopsy for NASH and Fibrosis, LIBRA, identifier: NCT04677101.

Original languageEnglish
Article number327498
Early online date12 Jul 2022
Publication statusE-pub ahead of print - 12 Jul 2022




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