Cytochrome P450 (CYP) enzyme induction is a biological process where a=exposure to an administered drug increases the activity and production of liver enzymes, causing other drugs to break down faster. This results in lower systemic exposure to the drug but can also increase the clearance of any other drugs administered. This process can also reduce the exposure in safety studies compromising safety margins.
Cytochrome P450 Induction
The primary mechanism of cytochrome P450 induction is via increased gene transcription which typically occurs through nuclear receptor activation, an inducing substance enters a liver cell and binds to nuclear receptors like PXR, CAR, or AhR. Whilst the first evidence of enzyme induction may occur after multiple doses in vivo (AUC and T1/2 reduction) there are a number of in vitro systems that can be used.
Whilst cytochrome P450 induction can be assayed in hepatocytes it is more usual to use nuclear receptor transactivation assays to assess the potential of test compounds to cause enzyme induction. PXR and AhR nuclear receptor activation can be assayed using a stably-transfected human hepatoma cell lines and a luciferase reporter gene assay in 96-well format.
Pregnane X receptor (PXR)
The pregnane X receptor (hPXR) is the major determinant of CYP3A gene regulation by drugs and other xenobiotics. In addition, PXR mediates induction of P450s 2B6, 2C8/9, and 3A4, as well as the drug transporters MDR1, organic anion transporting polypeptide C, bile salt export protein, and multidrug resistance-associated protein 2.
Known PXR activators include
Binding site is large and hydrophobic with several important hydrogen bonding interactions. May be multiple binding conformations. Similar to CYP3A pharmacophore, many (but not all) CYP3A substrates/inhibitors are also CYP3A inducers. There have been a number of in silico models built to predict PXR activation Jacobs et al used PLS and VolSurf descriptors for the development of a QSAR model for PXR/AhR interaction DOI. Xiao et al used docking into a modeled PXR ligand-binding domain that takes into consideration the structural variations of five published X-ray structures of PXR–ligand complexes DOI.
Low molecular weight (MW < 300) compounds were in general found to be non-binders, and those molecules that do not match the shape of the PXR ligand-binding site may also act as a non-binder. Secondly, the favorable hydrophobic interactions, mostly through aromatic π–π interactions, and the presence of suitable hydrogen bond(s) between the compounds and PXR are attributes of strong binders.
Machine learning techniques have also been applied (support vector machine (SVM), k-nearest neighbor (k-NN), and artificial neural networks (ANN)) DOI using 73 descriptors selected from a pool of over 500 descriptors, whilst this method has the advantage of speed and capacity, it is much more difficult for the medicinal chemist to interpret and decide what to make next.
In an in silico study looking at currently prescribed drugs a combination of machine learning and docking studies were able to identify previously unknown PXR activators, interestingly several compounds were poorly predicted by the docking studies. Perhaps because as the authors noted
In general, we observed that because of the relatively large size of the binding pocket, compounds may move around and interact with different sites within the binding pocket.
A recent study using PXR agonist data obtained from the Tox21 database provided by PubChem (https://pubchem.ncbi.nlm.nih.gov/source/824) 336 descriptors were calculated and a classification model was constructed using Light Gradient Boosting Machine (LightGBM). Molecular descriptors with particularly high contributions in the PXR agonist prediction model included vsurf_D and vsurf_Wp, which are related to hydrophobic and polar volumes, respectively. Other features with strong contributions included logS and h_logS representing solubility, h_logD representing the partition coefficient.
The PDB structure of PNU-142721 bound to PXR highlights a number of the issues with docking studies, PDB 3R8D
PXR employs one hydrogen bond and fourteen van der Waals contacts to interact with the ligand, but allows two loops adjacent to the ligand-binding pocket to remain disordered in the structure. These observations highlight the role structural flexibility plays in PXR’s promiscuous responses to xenobiotics.
PNU-142721 bound into the active site of PXR (Green surface=hydrophobic, Purple=H-bond, Blue =Polar)
Aryl hydrocarbon receptor (AhR)
The aryl hydrocarbon receptor (AhR) is a member of the basic helix–loop–helix (bHLH)-Per-ARNT-Sim (PAS) family of transcriptional regulators that control a variety of developmental and physiological events. Known ligands include ligands such as TCDD, coplanar polychlorinated biphenyls (PCBs) and dibenzo[a]antracene (DBMA). Ligand binding to the AhR is presumed to produce conformational changes in the AhR protein which result in the exposure of an AhR nuclear localization signal and the translocation of the whole complex into the nucleus where at the activated transcription factor complex is formed. This then binds to specific sequences of DNA resulting in increased transcription. Whilst CYP1A1 was the first cytochrome shown to be induced by TCDD an “Ahr gene battery” of Phase I and Phase II metabolizing enzymes consisting of CYP1A1, CYP1A2, CYP1B1, NQO1, ALDH3A1, UGT1A2 and GSTA1 have subsequently been identified.
obacco smoking has also been shown to induce CYP1A via activation of AhR. AhR also interacts with other signalling pathways such as those mediated by Estrogen Receptor and other Hormone receptors, Hypoxia, NF-KappaB and Rb and it appears to be directly involved in the reproductive toxicity caused by a range of aryl hydrocarbons, including the highly toxic compound 2,3,7,8-tetrachlor-odibenzo-p-dioxin (TCDD), DOI : 10.1530/rep.1.00294
In silico models have been developed Jacobs et al used PLS and VolSurf descriptors for the development of a QSAR model for PXR/AhR interaction DOI. Modeling of the Aryl Hydrocarbon Receptor (AhR) ligand binding domain and its utility in virtual ligand screening to predict new AhR ligands has been described DOI, this work also compared different species.
Constitutive androstane receptor (CAR)
The xenobiotic receptor constitutive androstane receptor (CAR) mediates the well-studied induction of CYP2B genes and other drug-metabolizing enzymes by phenobarbital (PB), an antiepileptic drug that has been shown to alter thyroid hormone (TH) levels. Unlike most nuclear receptors, this transcriptional regulator is constitutively active in the absence of ligand but is regulated by both agonists and inverse agonists. Ligand binding results in translocation of this protein to the nucleus, where it activates or represses target gene transcription.
Efavirenz is metabolised in the liver, and is both a substrate and inducer of the 2B6 and 3A4 isoforms of the cytochrome P450 system.
a 3D-QSAR model has been developed DOI using a combination of protein structure and ligand based methods.