Comparative Effects of Fibrates on Drug Metabolizing Enzymes in Human Hepatocytes

ABSTRACT:Induction of drug-metabolizing enzymes (DMEs) is highly speciesspecific and can lead to drug-drug interaction and toxicities. In this series of studies we tested the species specificity of the antidiabetic drug development candidate and mixed peroxisome proliferator-activated receptor (PPAR) ␣/␥ agonist (S)-4-O-tolylsulfanyl-2-(4-trifluormethyl-phenoxy)-butyric acid (EMD 392949, EMD) with regard to the induction of gene expression and activities of DMEs, their regulators, and typical PPAR target genes. EMD clearly induced PPAR␣ target genes in rats in vivo and in rat hepatocytes but lacked significant induction of DMEs, except for cytochrome P450 (P450) 4A. CYP2C and CYP3A were consistently induced in livers of EMD-treated monkeys. Interestingly, classic rodent peroxisomal proliferation markers were induced in monkeys after 17 weeks but not after a 4-week treatment, a fact also observed in human hepatocytes after 72 h but not 24 h of EMD treatment. In human hepatocyte cultures, EMD showed similar gene expression profiles and induction of P450 activities as in monkeys, indicating that the monkey is predictive for human P450 induction by EMD. In addition, EMD induced a similar gene expression pattern as the PPAR␣ agonist fenofibrate in primary rat and human hepatocyte cultures. In conclusion, these data showed an excellent correlation of in vivo data on DME gene expression and activity levels with results generated in hepatocyte monolayer cultures, enabling a solid estimation of human P450 induction. This study also clearly highlighted major differences between primates and rodents in the regulation of major inducible P450s, with evidence of CYP3A and CYP2C inducibility by PPAR␣ agonists in monkeys and humans.

Section: Discussionmentioning

confidence: 72%

“…This is most probably related, at least in part, to P450 induction (Prueksaritanont et al, 2005). Therefore, the aim of the present study was to compare the effects of EMD administration on DME regulators and DME mRNA expression in conjunction with related monooxygenase activities in different animal species and humans.…”

mentioning

confidence: 97%

Species Differences in the Response of Liver Drug-Metabolizing Enzymes to (S)-4-O-Tolylsulfanyl-2-(4-trifluormethyl-phenoxy)-butyric Acid (EMD 392949) in Vivo and in Vitro

Richert¹,

Tuschl²,

Viollon-Abadie³

et al. 2008

“…At the end of the incubation, drug-containing medium was discarded, and the cells were washed with drug-free medium. Cells were then exposed to the probe substrates 250 M testosterone (CYP3A4) or 100 M phenacetin (CYP1A2) for 1 h. The media were collected, and the metabolites produced by testosterone 6␤-hydroxylation or phenacetin de-ethylation were quantitated using liquid chromatography/tandem mass spectrometry as described previously (Prueksaritanont et al, 2005). The remaining cells were used to isolate mRNA, which was subjected to quantitative PCR to assess changes in CY3A4, CYP2B6, and CYP1A2 mRNA expression.…”

Section: Methodsmentioning

confidence: 99%

Comparison of Immortalized Fa2N-4 Cells and Human Hepatocytes as in Vitro Models for Cytochrome P450 Induction

Hariparsad¹,

Carr²,

Evers³

et al. 2008

ABSTRACT:Fa2N-4 cells have been proposed as a tool to identify CYP3A4 inducers. To evaluate whether Fa2N-4 cells are a reliable surrogate for cryopreserved human hepatocytes, we assessed the basal mRNA expression of 64 drug disposition genes in Fa2N-4 cells. Significant differences were found in the expression of major drugmetabolizing enzymes, nuclear receptors, and transporters between both cell types. Importantly, the expression of constitutive androstane receptor (CAR) and several hepatic uptake transporters was significantly lower (>50-fold) in Fa2N-4 cells, whereas the expression of pregnane X-receptor (PXR) and aryl hydrocarbon receptor (

“…The study was conducted using rhesus monkey hepatocytes (n ϭ 3) as described previously (Prueksaritanont et al, 2005(Prueksaritanont et al, , 2006. In brief, hepatocyte cultures were treated, in triplicate for each treatment, for 2 days with culture media containing various concentrations of rifampin or vehicle control (dimethyl sulfoxide, 0.1% v/v).…”

Section: Methodsmentioning

confidence: 99%

Rifampin Induces the in Vitro Oxidative Metabolism, but Not the in Vivo Clearance of Diclofenac in Rhesus Monkeys

Prueksaritanont

Li²,

Tang³

et al. 2006

Self Cite

ABSTRACT:Effects of rifampin on in vitro oxidative metabolism and in vivo pharmacokinetics of diclofenac (DF), a prototypic CYP2C9 marker substrate, were investigated in rhesus monkeys. In monkey hepatocytes, rifampin markedly induced DF 4-hydroxylase activity, with values for EC 50 of 0.2 to 0.4 M and E max of 2-to 5-fold over control. However, pretreatment with rifampin did not alter the pharmacokinetics of DF obtained after either i.v. or intrahepatic portal vein (i.pv.) administration of DF to monkeys. At the dose studied, plasma concentrations of rifampin reached 10 M, far exceeding the in vitro EC 50 values. Under similar treatment conditions, rifampin was previously shown to induce midazolam (MDZ) 1-hydroxylation in rhesus monkey hepatocytes (EC 50 and E max values ϳ0.2 M and ϳ2-to 3-fold, respectively), and markedly affected the in vivo pharmacokinetics of MDZ (>10-fold decreases in the i.pv. MDZ systemic exposure and its hepatic availability, F h ) in this animal species. In monkey liver microsomes, DF underwent, predominantly, glucuronidation, and, modestly, oxidation; the intrinsic clearance (CL int ‫؍‬ V max /K m ) value for the glucuronidation pathway accounted for >95% (versus about 75% in human liver microsomes) of the total (glucuronidation ؉ hydroxylation) intrinsic clearance value. In monkey hepatocytes, the hydroxylation also was a minor component (<10%) relative to the glucuronidation, supporting the liver microsomal finding. Collectively, our results suggest that the oxidative metabolism is not the major in vivo clearance mechanism of DF in either untreated or rifampin-treated monkeys and, conceivably, also in humans, raising a question about the utility of DF as an in vivo CYP2C9 probe.Induction of drug-metabolizing enzymes, especially the cytochrome P450 (P450) superfamily, by some drug molecules is a well known phenomenon and generally is undesirable since it can cause profound clinical effects, either by reducing therapeutic efficacy of drugs or enhancing toxicity from toxic or reactive metabolites (Thummel and Wilkinson, 1998). Accordingly, the potential for new chemical entities to cause P450 induction is usually assessed during lead optimization and identification in early drug discovery processes (Weaver, 2001;Worboys and Carlile, 2001). Currently, measurement of enzyme activities in cultured hepatocytes is the accepted and recommended method for studying P450 induction (LeCluyse, 2001;Bjornsson et al., 2003). However, systematic and quantitative extrapolations of such in vitro enzyme induction data to in vivo situations have not been extensively studied, and studies to date, including our recent investigation on in vitro-in vivo drug interactions in rhesus monkeys (Prueksaritanont et al., 2006), have been limited to CYP3A, the most abundant of all the human isoforms.In a quest to expand the database, we subsequently evaluated a relationship between in vitro-in vivo induction of CYP2C9 activity by rifampin, using diclofenac (DF) as a functional probe and the rhesus monkey as an anima...