Metabonomic evaluation of idiosyncrasy-like liver injury in rats cotreated with ranitidine and lipopolysaccharide.

Idiosyncratic liver injury occurs in a small fraction of people on certain drug regimens. The cause of idiosyncratic hepatotoxicity is not known; however, it has been proposed that environmental factors such as concurrent inflammation initiated by bacterial lipopolysaccharide (LPS) increase an individual's susceptibility to drug toxicity. Ranitidine (RAN), a histamine-2 receptor antagonist, causes idiosyncratic liver injury in humans. In a previous report, idiosyncrasy-like liver toxicity was created in rats by cotreating them with LPS and RAN. In the present study, the ability of metabonomic techniques to distinguish animals cotreated with LPS and RAN from those treated with each agent individually was investigated. Rats were treated with LPS or its vehicle and with RAN or its vehicle, and urine was collected for nuclear magnetic resonance (NMR)- and mass spectroscopy-based metabonomic analyses. Blood and liver samples were also collected to compare metabonomic results with clinical chemistry and histopathology. NMR metabonomic analysis indicated changes in the pattern of metabolites consistent with liver damage that occurred only in the LPS/RAN cotreated group. Principal component analysis of urine spectra by either NMR or mass spectroscopy produced a clear separation of the rats treated with LPS/RAN from the other three groups. Clinical chemistry (serum alanine aminotransferase and aspartate aminotransferase activities) and histopathology corroborated these results. These findings support the potential use of a noninvasive metabonomic approach to identify drug candidates with potential to cause idiosyncratic liver toxicity with inflammagen coexposure.

The value of DNA methylation analysis in basic, initial toxicity assessments.

DNA methylation is an epigenetic mechanism regulating patterns of gene expression. Our goal was to see if the assessment of DNA methylation might be a useful tool, when used in conjunction with initial, basic in vitro tests, to provide a more informative preliminary appraisal of the toxic potential of chemicals to prioritize them for further evaluation. We sought to give better indications of a compound's toxic potential and its possible mechanism of action at an earlier time and, thereby, contribute to a rational approach of an overall reduction in testing by making improved early decisions. Global and GC-rich patterns of DNA methylation were evaluated along with more traditional cytolethality measurements, e.g., cytolethality and genotoxicity assessments, on rat hepatoma (H4IIE) cells. The relative toxic potential of model compounds camptothecin, 5-fluorouracil, rotenone, and staurosporine was estimated by employing DNA methylation assessments combined with our cytolethality data plus genotoxicity information gleaned from the literature. The overall contribution of the methylation assessment was threefold; it (1) strengthened a ranking based on genotoxicity; (2) provided an indication that a compound might be more potentially problematic than what cytolethality and genotoxicity assessments alone would indicate; and (3) suggested that compounds, particularly nongenotoxins, that are more potent regarding their ability to alter methylation, especially at noncytolethal concentrations, may be more potentially toxic. Altered methylation per se is not proof of toxicity; this needs to be viewed as a component of an evaluation.

Ranitidine treatment during a modest inflammatory response precipitates idiosyncrasy-like liver injury in rats.

Drug idiosyncrasy is an adverse event of unknown etiology that occurs in a small fraction of people taking a drug. Some idiosyncratic drug reactions may occur from episodic decreases in the threshold for drug hepatotoxicity. Previous studies in rats have shown that modest underlying inflammation triggered by bacterial lipopolysaccharide (LPS) can decrease the threshold for xenobiotic hepatotoxicity. The histamine-2 (H2)-receptor antagonist ranitidine (RAN) causes idiosyncratic reactions in people, with liver as a usual target. We tested the hypothesis that RAN could be rendered hepatotoxic in animals undergoing a modest inflammatory response. Male rats were treated with a nonhepatotoxic dose of LPS (44 x 10(6) endotoxin units/kg i.v.) or its vehicle and then 2 h later with a nonhepatotoxic dose of RAN (30 mg/kg i.v.) or its vehicle. Liver injury was evident only in animals treated with both RAN and LPS as estimated by increases in serum alanine aminotransferase, aspartate aminotransferase, and gamma-glutamyl transferase activities within 6 h after RAN administration. LPS/RAN cotreatment resulted in midzonal liver lesions characterized by acute necrosuppurative hepatitis. Famotidine (FAM) is an H2-antagonist for which the propensity for idiosyncratic reactions is far less than RAN. Rats given LPS and FAM at a dose pharmacologically equipotent to that of RAN did not develop liver injury. In vitro, RAN sensitized hepatocytes to killing by cytotoxic products from activated neutrophils, whereas FAM lacked this ability. The results indicate that a response resembling human RAN idiosyncrasy can be reproduced in animals by RAN exposure during modest inflammation.

Strategic importance of research support through pathology.

The pace at which new drug candidates are being identified by Discovery Research demands that they be screened for preclinical attributes rapidly and efficiently. The early identification and elimination of compounds with toxic liabilities will produce safer drugs in a shorter time period, and with an increased rate of success. Most major pharmaceutical companies now recognize the strategic role of pathology support for research and have developed specific units to effect this outcome. The early interaction of these pathologists with drug discovery teams to identify compounds with toxic liabilities is critical. Approaches being used include high throughput in vitro screens to predict the relative toxicity of discovery compounds and to provide early indications of underlying mechanisms, target profiling to predict consequences of receptor-ligand interactions at other-than-indicated target sites, and acute in vivo studies to establish tolerability limits and target organs of toxicity. These approaches include the application of contemporary tools such as genomics, proteomics, metabonomics, and genetically engineered animal models. To maximize the benefit of discovery pathology, it is critical that pharmaceutical companies also actively participate in non-proprietary knowledge sharing and the education of pathologists and toxicologists to lead these efforts in the future.