Development of a novel non-invasive biomarker panel for hepatic fibrosis in individuals with MASLD

- Background & Aims: Considering the escalating prevalence of metabolic dysfunction-associated steatotic liver disease (MASLD) and MASLD-related fibrosis, accurate non-invasive biomarkers for diagnosis and staging of fibrosis are urgently needed. This study Aims to develop a blood-based biomarker panel for fibrosis detection in individuals with MASLD. - Approach & Results: Using a translational diet-induced LDLr-/-.Leiden MASLD mouse model, candidate biomarkers were identified focused on the mechanism of collagen deposition, by integrating hepatic gene expression and new extracellular matrix deposition, as detected by dynamic D2O-labeling. To translate these findings to humans, gene expression profiles and biomarkers were analyzed in liver biopsies and serum samples from 67 individuals with histologically characterized MASLD and variable degrees of fibrosis. This led to the selection of three biomarkers for a blood-based fibrosis biomarker panel: IGFBP7, SSc5D and Sema4D. The accuracy of the biomarker panel was tested in a separate cohort of 128 individuals with histologically characterized MASLD across different stages of fibrosis. A Light Gradient Boosting Machine (LGBM) model was applied to predict fibrosis stage in MASLD (F0/F1: AUC = 0.90; F2: AUC = 0.91; F3/F4: AUC = 0.87). - Conclusion & Discussion: Using a translational Approach to identify collagen turnover related proteins indicative of fibrosis, we developed an accurate blood-based biomarker panel to detect and stage hepatic fibrosis in individuals with MASLD. Overall design: - Selection of translation cohort To translate the murine findings to humans, 74 individuals with MASLD were selected of whom stored liver biopsy material was available at the pathology department of the Erasmus Medical Center and Utrecht Medical Center, the Netherlands. A patient list was generated with the following search criteria: “steatosis OR steatohepatitis OR NASH OR NAFLD” from 1997 onwards. Patient records were reviewed and patients diagnosed with HBV, HCV, alcoholic steatohepatitis and auto-immune hepatitis were excluded. In addition, patients with other comorbidities, treated with steroids, hormonal therapy including anticonception and with benign liver tumors were excluded. Then, biobanked stained liver tissue slides were collected and reviewed by an external pathologists (JV& MD) resulting in a Fibrosis score for each individual liver biopsy. All patients that passed this review and were diagnosed as true MASLD patients were then included. Fibrosis is assessed on a scale from stages 0-4. Stage 0 denotes no fibrosis, stage 1 indicates perisinusoidal or periportal fibrosis, stage 2 reflects perisinusoidal and (peri)portal fibrosis, stage 3 signifies bridging fibrosis, and stage 4 represents cirrhosis. Stored tissue blocks containing the biopsy sample was than requested from the tissue biobank and further processed for RNA isolation and next generation sequencing. Out of the 74 samples of human formalin-fixed paraffin-embedded (FFPE) liver, a subset of 67 samples passed Quality Control (QC) and was selected for RNA sequencing.