Identification of Serum Biomarkers to Distinguish Hazardous and Benign Aminotransferase Elevations

Abstract

The standard circulating biomarker of liver injury in both clinical settings and drug safety testing is alanine aminotransferase (ALT). However, ALT elevations sometimes lack specificity for tissue damage. To identify novel serum biomarkers with greater specificity for injury, we combined unique animal models with untargeted proteomics, followed by confirmation with immunoblotting. Using proteomics, we identified 109 proteins in serum from mice with acetaminophen (APAP)-induced liver injury that were not detectable in serum from mice with benign ALT elevations due to high-dose dexamethasone (Dex). We selected four (alcohol dehydrogenase 1A1 [Aldh1a1], aldehyde dehydrogenase 1 [Adh1], argininosuccinate synthetase 1 [Ass1], and adenosyhomocysteinase [Ahcy]) with high levels for further evaluation. Importantly, all four were specific for injury when using immunoblots to compare serum from Dex-treated mice and mice with similar lower ALT elevations due to milder models of APAP or bromobenzene-induced liver injury. Immunoblotting for ALDH1A1, ADH1, and ASS1 in serum from APAP overdose patients without liver injury and APAP overdose patients with mild liver injury revealed that these candidate biomarkers can be detected in humans with moderate liver injury as well. Interestingly, further experiments with serum from rats with bile duct ligation (BDL)-induced liver disease indicated that Aldh1a1 and Adh1 are not detectable in serum in cholestasis and may therefore be specific for hepatocellular injury and possibly even drug-induced liver injury (DILI), in particular. Overall, our results strongly indicate that ALDH1A1, ADH1, and ASS1 are promising specific biomarkers for liver injury. Adoption of these biomarkers could improve pre-approval drug safety assessment.

Registered report protocol: Quantitative analysis of septin Cdc10-associated proteome in Cryptococcus neoformans

“…MS data will be acquired using the FTMS analyzer in profile mode at a resolution of 240,000 over a range of 375 to 1500 m/z. Following HCD activation, MS/MS data will be acquired using the ion trap analyzer in centroid mode and normal mass range with precursor mass-dependent normalized collision energy between 28.0 and 31.0 [ 48 , 49 ].…”

Prospective study of ALDH1A1 gene polymorphisms associated with antituberculosis drug‐induced liver injury in western Chinese Han population

“…34 In another aspect, changes in ALDH1A1 expression appear to be part of the early acutephase hepatic inflammatory response 35 and ALDH1A1 is a promising biomarker for liver injury. 36 Moreover, the ALDH1A1*2 allele has an increased risk of liver toxicity, compared with patients with wild-type alleles when treated with a high-dose chemotherapy combination of cyclophosphamide, thiotepa, and carboplatin. 37 Given the above evidence, it would be of great value to investigate the relationship between ALDH1A1 and liver injury from anti-TB treatment.…”

“…Rebollido‐Rios et al found that nonsmall cell lung cancers with increased expression of ALDH1A1, ALDH1A3, or ALDH3A1 may be targeted by strategies involving inhibitors of these isoenzymes as monotherapy or in combination with chemotherapy to overcome patient‐specific drug resistance 34 . In another aspect, changes in ALDH1A1 expression appear to be part of the early acute‐phase hepatic inflammatory response 35 and ALDH1A1 is a promising biomarker for liver injury 36 . Moreover, the ALDH1A1*2 allele has an increased risk of liver toxicity, compared with patients with wild‐type alleles when treated with a high‐dose chemotherapy combination of cyclophosphamide, thiotepa, and carboplatin 37 .…”

The Challenge of Interpreting Alanine Aminotransferase Elevations in Clinical Trials of New Drug Candidates

“…The bardoxolone methyl experience highlights the urgent need for better DILI biomarkers and some promising new biomarkers have been proposed. 8,9 The key limiting factor in validation of new DILI biomarkers is the lack of serial serum samples prospectively collected and archived from clinical trials that have demonstrated varying degrees of liver safety. This will require standardized sample collection, storage, and retrieval of relevant phenotypic data.…”

“…However, the effects are modest and not consistent with the substantial ALT elevations observed in the clinical trials and seem unlikely to convince regulators of the safety of this drug candidate. The bardoxolone methyl experience highlights the urgent need for better DILI biomarkers and some promising new biomarkers have been proposed 8,9 . The key limiting factor in validation of new DILI biomarkers is the lack of serial serum samples prospectively collected and archived from clinical trials that have demonstrated varying degrees of liver safety.…”