Development and application of advanced analytical strategies for understanding paracetamol metabolism and toxicity.

Abstract

Paracetamol (N-acetyl-p-aminophenol [APAP]) has been authorised in the United Kingdom as an analgesic and antipyretic therapy for over 60 years. When ingested at supratherapeutic doses, its hepatic metabolism is altered from a non-toxic route via conjugations, with sulfate or glucuronic acid, to an oxidative pathway of metabolism via cytochrome p450 (CYP) enzymes resulting in the formation of the reactive intermediate N-acetyl-p-benzoquinone imine (NAPQI). This reacts with glutathione (GSH), depleting this in the liver, causing APAP induced liver injury. APAP overdose is the largest cause of acute liver failure (ALF) in the UK and the second largest cause of liver transplantation world-wide. In England and Wales APAP overdose results in around 100 000 hospital admissions per year, of which 50% result in extended stays, and contributes to around 150 deaths. While improved treatments for APAP-induced liver injury are being developed, it is important to understand how to predict the severity of liver injury; to reduce unnecessary hospital stays and the number of deaths. The hypotheses for the work described below are that GSH depletion following APAP activation to NAPQI is a major route of APAP toxicity, and metabolites related to GSH biosynthesis, including 5-oxoproline (5-OP) and ophthalmic acid (OA), are circulating biomarkers for GSH depletion. In addition, APAP- glutathione conjugate (APAP-SG) and related metabolites, are predictive of liver injury. For the development of new therapies, animal models of APAP overdose are required. A detailed knowledge of the routes of APAP metabolism in these species is crucial for understanding their translation to human. Analytical methods have been developed and applied to samples from animal models and patients. Plasma/serum concentrations of 5-OP have been shown to be a promising biomarker across animal models and human patients, especially when used in combination with concentrations of alanine aminotransferase (ALT) and glutathione-derived APAP conjugates.