Molecular Basis of Metabolism-Mediated Conversion of PK11195 from an Antagonist to an Agonist of the Constitutive Androstane Receptor

Mol Pharmacol. 2017 Jul;92(1):75-87. doi: 10.1124/mol.117.108621. Epub 2017 Apr 25.


The constitutive androstane receptor (CAR) plays an important role in xenobiotic metabolism, energy homeostasis, and cell proliferation. Antagonism of the CAR represents a key strategy for studying its function and may have potential clinical applications. However, specific human CAR (hCAR) antagonists are limited and conflicting data on the activity of these compounds have been reported. 1-(2-chlorophenyl)-N-methyl-N-(1-methylpropyl)-3-isoquinolinecarboxamide (PK11195), a typical peripheral benzodiazepine receptor ligand, has been established as a potent hCAR deactivator in immortalized cells; whether it inhibits hCAR activity under physiologically relevant conditions remains unclear. Here, we investigated the effects of PK11195 on hCAR in metabolically competent human primary hepatocytes (HPH) and HepaRG cells. We show that although PK11195 antagonizes hCAR in HepG2 cells, it induces the expression of CYP2B6 and CYP3A4, targets of hCAR and the pregnane X receptor (PXR), in HPH, HepaRG, and PXR-knockout HepaRG cells. Utilizing a HPH-HepG2 coculture model, we demonstrate that inclusion of HPH converts PK11195 from an antagonist to an agonist of hCAR, and such conversion was attenuated by potent CYP3A4 inhibitor ketoconazole. Metabolically, we show that the N-desmethyl metabolite is responsible for PK11195-mediated hCAR activation by facilitating hCAR interaction with coactivators and enhancing hCAR nuclear translocation in HPHs. Structure-activity analysis revealed that N-demethylation alters the interaction of PK11195 with the binding pocket of hCAR to favor activation. Together, these results indicate that removal of a methyl group switches PK11195 from a potent antagonist of hCAR to an agonist in HPH and highlights the importance of physiologically relevant metabolism when attempting to define the biologic action of small molecules.

MeSH terms

  • Adult
  • Aged
  • Child
  • Coculture Techniques
  • Constitutive Androstane Receptor
  • Dose-Response Relationship, Drug
  • Female
  • Hep G2 Cells
  • Hepatocytes / drug effects
  • Hepatocytes / metabolism
  • Humans
  • Isoquinolines / chemistry*
  • Isoquinolines / metabolism*
  • Isoquinolines / pharmacology
  • Male
  • Middle Aged
  • Protein Structure, Secondary
  • Receptors, Cytoplasmic and Nuclear / agonists
  • Receptors, Cytoplasmic and Nuclear / antagonists & inhibitors
  • Receptors, Cytoplasmic and Nuclear / chemistry*
  • Receptors, Cytoplasmic and Nuclear / metabolism*
  • Structure-Activity Relationship


  • Constitutive Androstane Receptor
  • Isoquinolines
  • Receptors, Cytoplasmic and Nuclear
  • PK 11195