Cultured hepatocytes as investigational models for hepatic toxicity: practical applications in drug discovery and development

Ulrich, Roger G.; Bacon, Jeffrey W.; Cramer, Clay T.; Peng, Geoffrey W.; Petrella, Diane K.; Stryd, R.P.; Sun, E

doi:10.1016/0378-4274(95)03547-8

Cited by 44 publications

(24 citation statements)

References 17 publications

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“…The primary cell culture system is well-established and consists of confluent cultures of freshly isolated hepatocytes from humans or preclinical species (Acosta et al, 1985;Butterworth et al, 1989;Ulrich et al, 1995;Guillouzo et al, 1997;Groneberg et al, 2002). Primary hepatocyte cultures have the full complement of metabolizing enzymes, albeit with inherent donor-to-donor variability, and thus represent an assay system that is more representative of hepatocytes in vivo as compared to immortalized cells (Guillouzo et al, 1993).…”

Section: In Vitro Cell Systemsmentioning

confidence: 99%

“…Use of in vitro systems is not new to investigative toxicology or lead optimization processes. Systems such as primary cell cultures, immortalized cell lines, liver slices and whole, perfused livers are well-established models used for investigative work (Acosta et al, 1985;Butterworth et al, 1989;Ulrich et al, 1995;Guillouzo et al, 1997;Groneberg et al, 2002). Likewise, the use of HepG2 cell lines and primary hepatocyte cell cultures to assess potential cytotoxicity are promoted by several commercial laboratories.…”

Section: Introductionmentioning

confidence: 99%

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New Technologies and Screening Strategies for Hepatotoxicity: Use of In Vitro Models

2005

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Hepatotoxicity remains a significant cause for drug failures during clinical trials. This is due, in part, to the idiosyncratic nature of toxicity in humans and inherent physiological differences between humans and preclinical species leading to limited correct prediction of adverse responses in humans. To address this issue, robust screening assays are being developed, which have heightened predictive capacity for human hepatotoxicity, and may be utilized throughout the discovery and development phases in conjunction with traditional in vivo methods, for decision making during drug selection and risk assessment. This manuscript describes an example application of in vitro-based strategies using human hepatocyte cultures in lead optimization screening in conjunction with ADME profiling, for evaluation of compound-associated CYP450 induction potential, and the identification of potentially useful biomarkers as predictors of hepatotoxicity for use in vitro, and in preclinical species and humans.

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Section: In Vitro Cell Systemsmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

New Technologies and Screening Strategies for Hepatotoxicity: Use of In Vitro Models

2005

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“…Therefore speci c strategies using the addition of matrix materials or agents have been developed to maintain metabolizing activity (8,9,87). Studies addressing the metabolizing capacity of human, rat, dog, and monkey cultured hepatocytes revealed that for the model drug adinazolam, 24 hours after isolation the cells still generated metabolites similar to those formed in vivo; therefore, it was concluded that for at the least the rst few days, cultured hepatocytes may be used to identify potential toxic metabolites (99).…”

Section: Isolated Liver Cell Modelsmentioning

confidence: 99%

“…The methodology of liver cell cultures has been improved over the last years and currently, a variety of isolation, culture, and cryopreservation models have been established and evaluated (99). Shortly after the rst description of hepatocyte isolation by collagenase-and hyaluronidase-dissociatio n (61,62), new improvements were made by perfusing the livers in situ with the dissociating agents (12).…”

Section: Isolated Liver Cell Modelsmentioning

confidence: 99%

In Vitro Models to Study Hepatotoxicity

Groneberg¹,

Große-Siestrup

Fischer³

2002

Toxicol Pathol

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Drug discovery and developmen t consists of a series of processes starting with the demonstration of pharmacologica l effects in experimental cell and animal models and ending with drug safety and ef cacy studies in patients. A main limitation is often the unacceptabl e level of toxicity with the liver as the primary target organ. Therefore, approache s to study hepatic toxicity in the early phase of drug discovery represent an important step towards rational drug development . A variety of in vitro liver models have been developed in the past years. Next to their use in drug development , they can also be applied to study environmenta l toxins and their hepatotoxicity. The 3 main approache s are ex vivo isolated and perfused organ models, precision-cut liver slices and cell culture models. Although the advantage of whole organ perfusions is based on the assessment of physiologic parameters such as bile production and morphologi c parameters such as tissue histology, cell culture models can be ef ciently used to assess cellular metabolism, cytotoxicity and genotoxicit y. The advantage of precision-cut liver slices is based on the juxtaposition of cellular assays and tissue morpholog y. None of these models can be compared as they all focus on different elds of hepatoxicolog y. For the future, the ideal setup for testing the hepatic toxicity of a new compoun d could of primary studies in cell or slice cultures to assess cellular effects and secondary studies using ex vivo perfused organs to examine gross organ function parameters and histology.

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“…Hepatocyte suspensions are applied for short-term studies (Bayliss et al, 1994;Steinberg et al, 1999), whereas cultured hepatocytes are a good tool for in vitro long-term examinations, to detect phase 1 and phase 2 metabolites and to show in vitro-in vivo correlations (Ulrich et al, 1995;Nussler et al, 2001;Pritchett et al, 2002;Brandon et al, 2003). Recently, the pig has become a more popular alternative in testing of new drug candidates (Zuber et al, 2002).…”

mentioning

confidence: 99%

METABOLISM OF N-HYDROXYGUANIDINES (N-HYDROXYDEBRISOQUINE) IN HUMAN AND PORCINE HEPATOCYTES: REDUCTION AND FORMATION OF GLUCURONIDES

Froehlich¹,

Girreser²,

Clement³

2005

Drug Metab Dispos

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ABSTRACT:The biotransformation of N-hydroxydebrisoquine, a model substrate for N-hydroxyguanidines, was studied in vitro with cultured and characterized porcine and human hepatocytes. The objective of the present work was to compare the N-oxidative and N-reductive metabolism of this compound using a monolayer culture system with previously described microsomal studies and to investigate the phase 2 metabolism, in particular, the glucuronidation of this class of compounds. At the same time, the suitability of pig hepatocytes as a model system for the human metabolism could be investigated. Two glucuronides of the parent compound Nhydroxydebrisoquine were analyzed. For the first time, one of these phase 2 metabolites could be identified as an O-glucuronide of an N-hydroxyguanidine by comparing it to a synthesized authentic compound. The involvement of certain human UDP-glucuronosyltransferases (UGTs) was evaluated by incubating the substrate with eight human hepatic recombinant UGT enzymes. Metabolites were determined by a newly developed LC-MS (liquid chromatography/mass spectrometry) analysis using electrospray ionization (ESI). The known microsomal reduction of the N-hydroxylated compound was also demonstrated with hepatocytes. The N-hydroxylation of the corresponding reduced compound (debrisoquine), which was previously described with microsomes, could not be detected in hepatocytes. There was no qualitative difference in the formation of the described derivatives by human and porcine hepatocytes. All phase 2 metabolites identified in hepatocyte culture were also formed by glucuronosyltransferases. In culture, the N-reduction of the N-hydroxylated substrate is the dominating reaction, indicating a predominance of N-reduction in vivo.

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Cultured hepatocytes as investigational models for hepatic toxicity: practical applications in drug discovery and development

Cited by 44 publications

References 17 publications

New Technologies and Screening Strategies for Hepatotoxicity: Use of In Vitro Models

New Technologies and Screening Strategies for Hepatotoxicity: Use of In Vitro Models

In Vitro Models to Study Hepatotoxicity

METABOLISM OF N-HYDROXYGUANIDINES (N-HYDROXYDEBRISOQUINE) IN HUMAN AND PORCINE HEPATOCYTES: REDUCTION AND FORMATION OF GLUCURONIDES

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