Hepatotoxic covalent binding study

Are Reactive Metabolite Trapping and Covalent Binding Studies Reliable Predictors of Hepatotoxic Potential of Drug Candidates ... 

In rodents, preanalytical factors such as food intake and restraint may alter plasma ALT (see Chapter 12 also). A 50% food restriction over 210 days in rats resulted in elevated ALT values compared to controls (Schwartz, Tornaben, and Boxhill 1973), while reduced levels of ALT were observed in a study of fasting effects on the oral toxicity of several xenobiotics (Kast and Nishikawa 1981). Changes of ALT related to diet may reflect perturbations of gluconeogenesis (Toropila et al. 1996). There are now several examples where plasma ALT falls after the administration of xenobiotics due to effects on pyridoxal phosphate, which is a cofactor necessary for action of the aminotransferases AST and ALT (Dhami et al. 1979 Rhodes et al. 1987 Waner et al. 1990 Waner and Nyska 1991). Such effects may confuse the interpretation of data when hepatotoxicity occurs and tends to increase ALT, but where there is an opposing effect due to reductions in pyridoxal phosphate. Further complications with ALT have been described by Wells and To (1986) in covalent binding studies with acetaminophen. 

Roberts, S.A., Veronica, F.P. and Jollow, D.J. (1990) Acetaminophen structure-toxicity studies In vitro covalent binding of a non-hepatotoxic analog, 3-hydroxy-acetanilide. Toxicology and Applied Pharmacology, 105, 195-208. 

Numerous studies have demonstrated an apparent relationship between metabolite formation and toxicity. The N-hydroxylation of phenacetin may play a role in drug-induced hepatic necrosis . Similarly, N-hydroxylation may mediate acetaminophen hepatotoxicity . Acetylhydrazine and isopropylhydrazine, metabolites of isoniazid and iproniazid, may initiate hepatotoxicity through covalent binding of an electrophilic intermediate (see Chapter 32) . 

The metabolites 2- and 4- bromophenol can also be metabolized by a further oxidation pathway to yield catechols and quinones, some of which are cytotoxic and potentially hepatotoxic. Thus, in vitro studies have indicated that bromoquinones and bromocatechols may be responsible for some of the covalent binding to protein and reaction with glutathione. However, administration of primary phenolic metabolites does not cause hepatotoxicity. At least seven glutathione conjugates have been identified as metabolites of bromobenzene and its primary phenolic metabolites. 

Immunologic techniques were used to study acetaminophen toxicity in mice to elucidate mechanisms of liver toxicity. The hepatotoxicity of acetaminophen is mediated by a reactive metabolite, N-acetyl-p-ben-zoquinone imine (NAPQI). The metabolite binds to protein as 3-(cystein-S-yl)acetaminophen (3-Cys-A) and the amount of binding correlates with toxicity. This covalent binding, and in particular the 3-Cys-A adduct, is the most reliable biomarker of acetaminophen toxicity (9-12). 

Acetaminophen (4.108) is hepatotoxic at higher doses, a toxicity explained by a cytochrome P450 dependent activation to A-acetyl-p-benzoquinonimine, which binds covalently to critical proteins (Chapt. 7 in [21]). However, the role of the hydrolytic step in acetaminophen-induced nephrotoxicity is not entirely clear. Early studies suggested a deacetylase-dependent activation of... 

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