Pharmacokinetics explain in vivo/in vitro discrepancies of carcinogen-induced gene expression alterations in rat liver and cultivated hepatocytes

Abstract

Cultivated hepatocytes represent a well-established in vitro system. However, the applicability of hepatocytes in toxicogenomics is still controversially discussed. Recently, an in vivo/in vitro discrepancy has been described, whereby the non-genotoxic rat liver carcinogen methapyrilene alters the expression of the metabolizing genes SULT1A1 and ABAT, as well as the DNA damage response gene GADD34 in vitro, but not in vivo. If the collagen sandwich cultures of hepatocytes really produce false-positive data, this would compromise its application in toxicogenomics. To revisit the putative in vivo/in vitro discrepancy, we first analyzed and modeled methapyrilene concentrations in the portal vein of rats. The relatively short half-life of 2.8 h implies a rapid decrease in orally administered methapyrilene in vivo below concentrations that can cause gene expression alterations. This corresponded to the time-dependent alteration levels of GADD34, ABAT and SULT1A1 RNA in the liver: RNA levels are altered 1, 6 and 12 h after methapyrilene administration, but return to control levels after 24 and 72 h. In contrast, methapyrilene concentrations in the culture medium supernatant of primary rat hepatocyte cultures decreased slowly. This explains why GADD34, ABAT and SULT1A1 were still deregulated after 24 h exposure in vitro, but not in vivo. It should also be considered that the earliest analyzed time point in the previous in vivo studies was 24 h after methapyrilene administration. In conclusion, previously observed in vitro/in vivo discrepancy can be explained by different pharmacokinetics present in vitro and in vivo. When the in vivo half-life is short, levels of some initially altered genes may have returned to control levels already 24 h after administration.

Toxicogenomics directory of chemically exposed human hepatocytes

“…Gene array analysis performed on the livers of rats facilitated the differentiation between genotoxic and non-genotoxic carcinogens (Ellinger-Ziegelbauer et al 2008;. These successful toxicogenomics studies in vivo prompted several ambitious research programs, including the EU-funded SEURAT-1 network and the ESNATs project Krug et al 2013;Schug et al 2013;Weng et al 2014;Balmer et al 2014;Campos et al 2014), with the aim to identify biomarkers of toxicity in vitro. One long-term goal of these projects is to identify biomarkers in the in vitro systems that can predict certain mechanisms of toxicity in vivo.…”

Highlight report: acetaminophen hepatotoxicity

“…Currently, intensive research activities are ongoing to establish in vitro systems particularly for hepatotoxicity (Grinberg et al 2014;Ramaiahgari et al 2014;Rodrigues et al 2013;Schug et al 2013), nephrotoxicity (Sanchez-Niño et al 2014;Fujiki et al 2013;, neurotoxicity (Barbosa et al 2014;Sisnaiske et al 2014;Frimat et al 2010) and developmental toxicity (Krug et al 2013;Rempel et al 2015;Balmer et al 2014;Zimmer et al 2014). For validation of in vitro systems with hepatocytes APAP represents a popular reference compound (Jennings et al 2014;Hengstler et al 2014).…”

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“…Vertical integration of computational models across different levels of biological organization has been used, for example, to simulate the effect of acetaminophen at the cellular scale (Krauss et al 2012;Diaz Ochoa et al 2013) or to describe the impact of steatosis on drug pharmacokinetics (Schwen et al 2014). In future, integrated computational models are generally expected to play an increasingly important role for integration and analysis of experimental data in drug development and toxicology (Magdy et al 2013;Godoy et al 2013;Zanger and Schwab 2013;Thomas et al 2013;Mielke et al 2011;Schug et al 2013;Heise et al 2012). …”