Direct Detection of miR-122 in Hepatotoxicity Using Dynamic Chemical Labeling Overcomes Stability and isomiR...

Area of Science:

• Biochemistry
• Molecular Biology
• Biomarker Discovery

Background:

• Circulating microRNAs (miRNAs) are stable, cross-species biomarkers.
• MicroRNA-122 (miR-122) is a specific biomarker for drug-induced liver injury (DILI).
• Existing detection methods face challenges with sample stability and miRNA variants (isomiRs).

Purpose of the Study:

• To develop and validate a novel assay for direct detection of miR-122 in blood.
• To assess the assay's utility in diagnosing DILI in humans and other species.
• To address limitations of current methods regarding sample stability and isomiR quantification.

Main Methods:

• Development of a single-molecule, dynamic chemical labeling (DCL) assay for direct miR-122 detection.
• Assay validation using human serum samples (n=192) from acetaminophen overdose cases.
• Comparative analysis with traditional PCR and RNA sequencing for stability and isomiR detection.

Main Results:

• The DCL assay accurately identified DILI risk in humans (AUC 0.98, P < 0.0001) using 10 µL serum/plasma.
• The assay demonstrated cross-species applicability, detecting liver injury in rats and dogs.
• DCL showed superior stability, with no signal loss after 14 days at room temperature, unlike PCR.
• RNA sequencing revealed DILI-specific miR-122 isomiRs, accurately quantified by DCL but not PCR.

Conclusions:

• The DCL assay provides accurate, direct measurement of miR-122 for diagnosing liver injury in humans and animals.
• This novel method overcomes critical challenges related to miRNA stability and isomiR complexity.
• DCL offers a promising tool for reliable DILI diagnostics and research.