In vivo and mechanistic evidence of nuclear receptor CAR induction by artemisinin

Abstract

Artemisinin (a sesquiterpene lactone endoperoxide) has become important in multi-drug treatment of malaria. There is evidence that artemisinin induces drug metabolism which could result in drug-drug interactions. The objective of this study was to characterize the inductive properties of artemisinin on drug-metabolizing cytochrome P450 (CYP450) enzymes. The possibility of artemisinin to induce CYP450 was studied in artemisinin-treated (orally for four days) and vehicle-treated rats using reverse transcriptase polymerase chain reaction (RT-PCR). The effect on enzymatic activities in mouse microsomes from multiple artemisinin administration (intraperitonally) to mice were also studied as well as the effect on the expression in mouse primary hepatocytes and HEK293 cells. Increased CYP2B1 mRNA levels in rats could be seen after artemisinin treatment as well as a weak but reproducible increase in the intensity of CYP1A2. Administration of artemisinin to mice up-regulated hepatic CYP2B10-dependent, and to a lesser extent, CYP2A5-dependent enzyme activities. In primary hepatocyte culture, artemisinin significantly increased the CYP2B10 mRNA levels whereas the CYP2A5 mRNA levels were increased to a lesser extent. No significant changes were seen in the levels of other CYP enzymes. Artemisinin was an activator of constitutive androstane receptor (CAR) but not pregnane X receptor (PXR) in HEK293 cells. The results demonstrate that the drug exerts its effects on drug metabolism via the CAR receptor that results in up-regulation of genes such as the Cyp2b. The weaker up-regulation of CYP2A5 might also be CAR-dependent or alternatively, a consequence of artemisinin toxicity. The results of this study are of importance when predicting potential drug-drug interactions in multi-drug therapies with artemisinin.

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

“…2 and 3). Among the compounds tested, rifampicin, bosentan, moricizine, and ritonavir are reported as selective PXR-activators (LeCluyse, 2001;Luo et al, 2002;van Giersbergen et al, 2002), phenobarbital, phenytoin, and efavirenz are PXR/CAR dual activators (Hariparsad et al, 2004;Wang et al, 2004;Trubetskoy et al, 2005;Bell and Michalopoulos, 2006;Faucette et al, 2007), and CITCO and artemisinin are selective CAR activators (Maglich et al, 2003;Simonsson et al, 2006). Rifampicin-mediated CYP3A4 induction was conducted in Fa2N-4 cells and four batches of human hepatocytes (DMQ, 527, 455, and LHO).…”

“…In addition, several coactivators and corepressors of PXR (Moore et al, 2006) such as the small heterodimer partner (SHP/NCOB2), nuclear receptor corepressor 2 (NCOR2/SMRT), steroid receptor coactivators 1 (SRC1/NCOA1) and 2 (SRC2/GRIP1), nuclear receptor interacting protein 1 (NRIP1/RIP140), peroxisome proliferator-activated receptor-␥ coactivator, and Forkhead transcription factor FKHR (Maglich et al, 2003;Simonsson et al, 2006). b Gene induction occurs via cross-talk between PXR and CAR (Wang et al, 2004;Trubetskoy et al, 2005;Bell and Michalopoulos, 2006).…”

Methadone Induces the Expression of Hepatic Drug-Metabolizing Enzymes through the Activation of Pregnane X Receptor and Constitutive Androstane Receptor

“…Nevertheless, it is noteworthy that the majority of known hCAR activators are PB-like compounds that activate hCAR through indirect mechanisms without direct ligand binding. As a matter of fact, CITCO and artemisinin are the only two hCAR agonists identified thus far (Maglich et al, 2003;Simonsson et al, 2006). Compared with the cell-based hCAR reporter assays, chemical-mediated hCAR nuclear translocation in hepatocytes of primary culture or in vivo appears to correlate well with hCAR activation and target gene induction, regardless of the distinction between direct or indirect mechanisms (Wang et al, 2004;Faucette et al, 2007).…”

A consecutive three alanine residue insertion mutant of human CAR: a novel CAR ligand screening system in HepG2 cells

“…Secondly, the effects of Andro, CLT, artemisinin (Simonsson et al, 2006), and PK11195 (Li et al, 2008), all ligands of CAR, on the transactivation potential of GAL4/DBD-hCAR/LBD(+3a.a.) were evaluated.…”