Candidate biomarkers for the diagnosis and prognosis of drug-induced liver injury: An international collaborative effort

Abstract

Current blood biomarkers are suboptimal in detecting drug-induced liver injury (DILI) and predicting its outcome. We sought to characterize the natural variabilty and performance characteristics of 14 promising DILI biomarker candidates. Serum or plasma from multiple cohorts of healthy volunteers (n = 192 and n = 81), subjects who safely took potentially hepatotoxic drugs without adverse effects (n = 55 and n = 92) and DILI patients (n = 98, n = 28, and n = 143) were assayed for microRNA-122 (miR-122), glutamate dehydrogenase (GLDH), total cytokeratin 18 (K18), caspase cleaved K18, glutathione S-transferase α, alpha-fetoprotein, arginase-1, osteopontin (OPN), sorbitol dehydrogenase, fatty acid binding protein, cadherin-5, macrophage colony-stimulating factor receptor (MCSFR), paraoxonase 1 (normalized to prothrombin protein), and leukocyte cell-derived chemotaxin-2. Most candidate biomarkers were significantly altered in DILI cases compared with healthy volunteers. GLDH correlated more closely with gold standard alanine aminotransferase than miR-122, and there was a surprisingly wide inter- and intra-individual variability of miR-122 levels among healthy volunteers. Serum K18, OPN, and MCSFR levels were most strongly associated with liver-related death or transplantation within 6 months of DILI onset. Prediction of prognosis among DILI patients using the Model for End-Stage Liver Disease was improved by incorporation of K18 and MCSFR levels. Conclusion: GLDH appears to be more useful than miR-122 in identifying DILI patients, and K18, OPN, and MCSFR are promising candidates for prediction of prognosis during an acute DILI event. Serial assessment of these biomarkers in large prospective studies will help further delineate their role in DILI diagnosis and management.

Figure 1. Intra-Individual Variabilty in PSTC Cohort Observed in microR-122 (miR-122) Quantifications

Three fasting blood samples were collected from volunteers over the course of 21 days. Greater intra-subject variability was observed in miR-122 levels amongst black subjects in this study compared to white subjects. Each bar represents an individual subject while circles represent data for miR-122 measurements 1 (white), 2 (black), and 3 (gray).

Figure 2. Correlation Between Levels of Alanine Aminotransferase (ALT) and Liver-Specific Biomarkers

Correlation of ALT with glutamate dehydrogenase (GLDH; A) and microRNA-122 (miR-122; B). Data points are individual PSTC (Predictive Safety Testing Consortium) and SAFE-T (Safer and Faster Evidence-based Translation) subject samples and represent individuals that did not have drug-induced liver injury (DILI; gray) and individuals that did have DILI (black). Values are log normalized. Pearson r is shown.

Figure 3. Assessment of Keratin 18 (K18) Measurements

Differences in serum K18 (A), caspase cleaved K18 (ccK18; B), and Apoptotic Index (AI; C) between Drug-Induced Liver Injury Network patients that did not die/require a liver transplant by 6 months post-DILI-onset and those that did. Differences in AI (D) between SAFE-T patients experiencing DILI associated with flupirtine utilization and patients experiencing DILI unrelated to flupirtine. Data points represent individual patients. A dotted line for AI is drawn at 0.5, representing a score that suggests an equal contribution of apoptosis and necrosis. Values for K18 and ccK18 are log normalized. Significance is **p<0.01 and ***p<0.001.

Figure 4. Incorporation of Candidate Biomarkers Into Model of End-Stage Liver Disease (MELD) Score Prognostic Model

Prognostic model optimized for prediction of death/transplant in Drug-Induced Liver Injury (DILIN) patients (n=141) using the MELD score, total keratin 18 (K18), and macrophage colony stimulating factor receptor (MCSFR) measurements (sensitivity=0.933, specificity=0.889). White boxes represent branching points, light grey boxes represent patients not predicted to have an adverse outcome, and dark grey boxes represent patients predicted to die/require transplant. Numbers in italics represent false results (i.e. 18 false positives and 1 false negative).