Virtual experiments provide plausible explanations for IVIVE underpredictions of hepatic clearance

We demonstrate how results of virtual experiments can suggest mechanism-based explanations for IVIVE underpredictions of hepatic clearance. We use agent-oriented, discrete-event methods. We experiment on software analogies of rats and hepatocyte cultures dosed identically with objects that mimic two idealized compounds, vC1 (not cleared) and vC2 (removal maximized). Hepatocytes in cultures and rat livers interact identically with vC1 and vC2. The rules governing vC1 and vC2 movements and interactions with hepatocytes are the same in both systems. The two software systems are independent; to be useful in discovering plausible explanations of IVIVE inaccuracies, we require that, in the absence of hepatocyte exposure differences, the culture-to-rat scaling factor = 1.0 for all corresponding measures. The probability of cell entry (pEnter) maps directly to the unbound-fraction of drug used by IVIVE methods. For vC1 (vC2), we achieve that validation target for pEnter = 0.05-1.0 (1.0). However, for pEnter = 0.05-0.8, vC2 removal rates during culture experiments underpredicted corresponding removal rates in rats. The magnitude of the underpredictions increases with decreasing pEnter. For example, using pEnter = 0.1 (0.3), peak vC2 removal rates in culture experiments underpredict corresponding removal rates in rats by 3.2 (1.4) fold. Underpredictions are a consequence of the biomimetic periportal-to-pericentral organization of hepatocytes within virtual livers are absent in virtual cultures. In rats, pericentral hepatocytes do more of the vC2 removal work. The results suggest that using IVIVE methods that abstract away influential features of hepatocyte organization within livers may contribute to IVIVE underpredictions.Significance StatementFrom experiments on virtual (software) counterparts of rats and hepatocyte cultures, we learned that IVIVE underpredictions are a consequence of the biomimetic periportal-to-pericentral organization of hepatocytes within virtual livers being absent in virtual cultures. Using IVIVE methods that abstract away influential features of hepatocyte organization within livers may contribute in part to some IVIVE underpredictions.Visual Abstract

Metabolomic Profile of Primary Turkey and Rat Hepatocytes and Two Cell Lines after Chloramphenicol Exposure

The purpose of this study was to assess the formation of chloramphenicol metabolites in primary turkey and rat hepatocyte cultures and human hepatoma (HepG2) cells and nonhepatic, Balb/c 3T3 fibroblasts. Additionally, the cytotoxicity of the drug was assessed through three biochemical endpoints: mitochondrial and lysosomal activity and cellular membrane integrity after 24 and 48 h exposure. The two metabolites of the drug, chloramphenicol glucuronide and nitroso-chloramphenicol, were detected to the greatest extent in both primary hepatocyte cultures by liquid chromatography–tandem mass spectrometry. Toxic nitroso-chloramphenicol was the main metabolite in the primary turkey hepatocyte cultures, but it was not in the primary rat hepatocyte cultures. The most affected endpoint in turkey and rat hepatocyte cultures was the disintegration of the cellular membrane, but in the cell lines, mitochondrial and lysosomal activities underwent the greatest change. The primary hepatocyte cultures represent valuable tools with which to study the species differences in the biotransformation and toxicity of drugs. To the best of our knowledge, this is the first report of differences in chloramphenicol metabolism in primary turkey and rat hepatocyte cultures.