Detection of nanolevel drug metabolites in an organotypic culture of primary human hepatocytes and porcine hepatocytes with special reference to a two-compartment model

ection of nanolevel drug metabolites in an organotypic culture of primary human hepatocytes and porcine hepatocytes with special reference to a two-compartment model Original Research (922) Total Article Views Authors: Acikg z A, Giri S, Bader A Published Date November 2012 Volume 2012:7 Pages 5859 - 5872 DOI: http://dx.doi.org/10.2147/IJN.S29651 Received: 03 January 2012 Accepted: 11 February 2012 Published: 27 November 2012 Ali Acikg z,1,2 Shibashish Giri,1 Augustinus Bader1 1Center for Biotechnology and Biomedicine, Cell Techniques and Applied Stem Cell Biology, Universit t Leipzig, Leipzig, Germany; 2Klinikum St Georg, Leipzig, Germany Abstract: The quantification of drug metabolites produced during drug metabolism is a growing concern for the pharmaceutical industry, regulatory agencies such as the US Food and Drug Administration, the European Medicines Agency, and others. As 70% of drugs are known reactive metabolites and have black box warnings, they are a major cause of drug-induced injury and lead to drug attrition in early or late clinical stages. According to a 2006 survey report of pharmaceutical companies, drug-induced liver injury was ranked first in terms of adverse events, and it remains the most common reason for restriction or withdrawal of a drug from the market by the Food and Drug Administration. Although there are many reasons underlying drug-induced liver injury, one of the most important is liver failure induced by drug metabolites. Generally, a drug produces metabolites that may bind to cellular molecules and trigger a toxicological effect, cause serious adverse drug reactions, or alter cellular functions. Experimental cellular models that attempt to qualify drug metabolites from cell cultures rely on human plasma and urine obtained from clinical trials and supernatant during early in vitro experiments. However, there is a lack of information about the quantification of drug metabolites inside human hepatocytes, where the drug is extensively metabolized. To overcome this limitation, we used the highly accepted, gold standard organotypic cellular model of primary human hepatocytes to investigate and quantify the parent drug, as well as drug metabolites inside human hepatocytes and outside human hepatocytes to evaluate the quantity of drug metabolites, which are assumed to have remained inside the primary human hepatocytes. We refer to this as a two-compartment model, where one compartment is supernatant compared with in vivo hepatic blood circulation, and the other is inside the hepatocyte cell compared with the inside of in vivo human liver. We detected the nanoconcentrations of all major metabolites (desmethyldiazepam, temazepam, and oxazepam) of the diazepam drug, both inside the cells (matrix) and outside the hepatocyte cells (supernatant) at different time points (primary human hepatocytes: 0, 1, 2, 4, 8, and 24 hours; primary porcine hepatocytes: 0, 1, 2, 5, and 24 hours) during biotransformation in an organotypic sa