ABSTRACT
Aggrecanase-1 and -2 (ADAMTS-4 and ADAMTS-5) are zinc metalloproteases involved in the degradation of aggrecan in cartilage. Inhibitors could provide a means of altering the progression of osteoarthritis. We report the identification of 7 which had good oral pharmacokinetics in rats and showed efficacy in a rat chemical model of osteoarthritis. The projected human dose required to achieve sustained plasma levels ≥10 times the hADAMTS-5 IC50 is 5 mg q.d.
Subject(s)
ADAMTS4 Protein/antagonists & inhibitors , ADAMTS5 Protein/antagonists & inhibitors , Enzyme Inhibitors/chemistry , Enzyme Inhibitors/therapeutic use , Hydantoins/chemistry , Hydantoins/therapeutic use , Osteoarthritis/drug therapy , ADAMTS4 Protein/metabolism , ADAMTS5 Protein/metabolism , Aggrecans/metabolism , Animals , Enzyme Inhibitors/blood , Enzyme Inhibitors/pharmacology , Humans , Hydantoins/blood , Hydantoins/pharmacology , Male , Molecular Docking Simulation , Osteoarthritis/enzymology , Osteoarthritis/metabolism , Rats , Rats, Inbred LewABSTRACT
With inadequate efficacy being the primary cause for the attrition of drug candidates in clinical development, the need to better predict clinical efficacy earlier in the drug development process has increased in importance in the pharmaceutical industry. Here, we review current applications of translational pharmacokinetic-pharmacodynamic (PK-PD) modeling of preclinical data in the pharmaceutical industry, including best practices. Preclinical translational PK-PD modeling has been used in many therapeutic areas and has been impactful to drug development. The role of preclinical translational PK-PD modeling in drug discovery and development will continue to evolve and broaden, given that its broad implementation in the pharmaceutical industry is relatively recent and many opportunities still exist for its further application.
Subject(s)
Drug Discovery/methods , Drug Industry/methods , Animals , Drug Evaluation, Preclinical/methods , Humans , Models, BiologicalABSTRACT
We describe a novel class of acidic mPGES-1 inhibitors with nanomolar enzymatic and human whole blood (HWB) potency. Rational design in conjunction with structure-based design led initially to the identification of anthranilic acid 5, an mPGES-1 inhibitor with micromolar HWB potency. Structural modifications of 5 improved HWB potency by over 1000×, reduced CYP2C9 single point inhibition, and improved rat clearance, which led to the selection of [(cyclopentyl)ethyl]benzoic acid compound 16 for clinical studies. Compound 16 showed an IC80 of 24nM for inhibition of PGE2 formation in vitro in LPS-stimulated HWB. A single oral dose resulted in plasma concentrations of 16 that exceeded its HWB IC80 in both rat (5mg/kg) and dog (3mg/kg) for over twelve hours.
Subject(s)
Benzoates/chemistry , Benzoates/pharmacology , Drug Discovery , Microsomes/drug effects , Prostaglandin-E Synthases/antagonists & inhibitors , Animals , Crystallography, X-Ray , Dogs , Microsomes/enzymology , Prostaglandin-E Synthases/chemistry , RatsABSTRACT
In an effort to develop a novel therapeutic agent aimed at addressing the unmet need of patients with osteoarthritis pain, we set out to develop an inhibitor for autotaxin with excellent potency and physical properties to allow for the clinical investigation of autotaxin-induced nociceptive and neuropathic pain. An initial hit identification campaign led to an aminopyrimidine series with an autotaxin IC50 of 500 nM. X-ray crystallography enabled the optimization to a lead compound that demonstrated favorable potency (IC50 = 2 nM), PK properties, and a robust PK/PD relationship.
ABSTRACT
The development of reliable relationships between in vivo target engagement, pharmacodynamic activity, and efficacy in chronic disease models is beneficial for enabling hypothesis-driven drug discovery and facilitating the development of patient-focused candidate selection criteria. Toward those ends, osmotic infusion pumps can be useful for overcoming limitations in the PK properties of proof-of-concept (POC) compounds to accelerate the development of such relationships. In this report, we describe the application of this strategy to the development of hydantoin-derived aggrecanase inhibitors (eg, 3) for the treatment of osteoarthiritis (OA). Potent, selective inhibitors were efficacious in both chemical and surgical models of OA when exposures were sustained in excess of 10 times the plasma IC50. The use of these data for establishing patient-focused candidate selection criteria is exemplified with the characterization of compound 8, which is projected to sustain the desired level of target engagement at a dose of 45 mg qd.
Subject(s)
Endopeptidases/metabolism , Enzyme Inhibitors/pharmacology , Enzyme Inhibitors/pharmacokinetics , Hydantoins/pharmacology , Hydantoins/pharmacokinetics , Osteoarthritis/drug therapy , Animals , Dose-Response Relationship, Drug , Enzyme Inhibitors/chemical synthesis , Enzyme Inhibitors/chemistry , Humans , Hydantoins/chemical synthesis , Hydantoins/chemistry , Male , Molecular Structure , Osmosis/drug effects , Osteoarthritis/metabolism , Rats , Rats, Inbred Lew , Structure-Activity RelationshipABSTRACT
The application of modeling and simulation techniques is increasingly common in preclinical stages of the drug discovery and development process. A survey focusing on preclinical pharmacokinetic/pharmacodynamics (PK/PD) analysis was conducted across pharmaceutical companies that are members of the International Consortium for Quality and Innovation in Pharmaceutical Development. Based on survey responses, ~68% of companies use preclinical PK/PD analysis in all therapeutic areas indicating its broad application. An important goal of preclinical PK/PD analysis in all pharmaceutical companies is for the selection/optimization of doses and/or dose regimens, including prediction of human efficacious doses. Oncology was the therapeutic area with the most PK/PD analysis support and where it showed the most impact. Consistent use of more complex systems pharmacology models and hybrid physiologically based pharmacokinetic models with PK/PD components was less common compared to traditional PK/PD models. Preclinical PK/PD analysis is increasingly being included in regulatory submissions with ~73% of companies including these data to some degree. Most companies (~86%) have seen impact of preclinical PK/PD analyses in drug development. Finally, ~59% of pharmaceutical companies have plans to expand their PK/PD modeling groups over the next 2 years indicating continued growth. The growth of preclinical PK/PD modeling groups in pharmaceutical industry is necessary to establish required resources and skills to further expand use of preclinical PK/PD modeling in a meaningful and impactful manner.
Subject(s)
Computer Simulation , Drug Evaluation, Preclinical/methods , Drug Industry/methods , Models, Biological , Data Collection , Dose-Response Relationship, Drug , Drug Design , Drug Discovery/methods , Drug Industry/statistics & numerical data , HumansABSTRACT
Nociceptin/OFQ (N/OFQ) is a 17 amino acid peptide that is the endogenous ligand for the ORL1/NOP receptor. Nociceptin appears to regulate a host of physiological functions such as biological reactions to stress, anxiety, mood, and drug abuse, in addition to feeding behaviors. To develop tools to study the function of nociceptin and NOP receptor, our research effort sought to identify orally available NOP antagonists. Our effort led to the discovery of a novel chemical series based on the dihydrospiro(piperidine-4,7'-thieno[2,3-c]pyran) scaffold. Herein we show that dihydrospiro(piperidine-4,7'-thieno[2,3-c]pyran)-derived compounds are potent NOP antagonists with high selectivity versus classical opioid receptors (µ, δ, and κ). Moreover, these compounds exhibit sufficient bioavailability to produce a high level of NOP receptor occupancy in the brain following oral administration in rats.
Subject(s)
Narcotic Antagonists , Pyrans/chemical synthesis , Administration, Oral , Animals , Drug Discovery , Male , Pyrans/pharmacokinetics , Pyrans/pharmacology , Rats , Rats, Sprague-Dawley , Receptors, Opioid , Structure-Activity Relationship , Nociceptin ReceptorABSTRACT
Currently, a lack of sufficient tools has limited the understanding of the relationship between neuropsychiatric disorders and the nociceptin/orphanin FQ (N/OFQ) peptide (NOP) receptor. Herein, we describe the discovery and development of an antagonist NOP receptor occupancy (RO) tracer and a novel positron emission tomography (PET) radioligand suitable to probe the NOP receptor in human clinical studies. A thorough structure-activity relationship (SAR) around the high-affinity 3-(2'-fluoro-4',5'-dihydrospiro[piperidine-4,7'-thieno[2,3-c]pyran]-1-yl)-2-(2-halobenzyl)-N-alkylpropanamide scaffold identified a series of subnanomolar, highly selective NOP antagonists. Subsequently, these unlabeled NOP ligands were evaluated in vivo by liquid chromatography-tandem mass spectrometry (LC-MS/MS) in rat to determine brain uptake, kinetics and specific binding. (S)-27 was identified as a suitable unlabeled preclinical RO tracer to accurately quantify NOP receptor engagement in rat brain. Three compounds were selected for evaluation in nonhuman primates as PET tracers: (-)-26, (-)-30, and (-)-33. Carbon-11 labeling of (+)-31 yielded [(11)C]-(S)-30, which exhibited minimal generation of central nervous system (CNS) penetrant radiometabolites, improved brain uptake, and was an excellent PET radioligand in both rat and monkey. Currently [(11)C]-(S)-30 is being evaluated as a PET radiotracer for the NOP receptor in human subjects.
Subject(s)
Radiopharmaceuticals/chemical synthesis , Receptors, Opioid/metabolism , Spiro Compounds/chemical synthesis , Thiophenes/chemical synthesis , Animals , Brain/diagnostic imaging , Brain/metabolism , CHO Cells , Carbon Radioisotopes , Chromatography, Liquid , Cricetinae , Cricetulus , HEK293 Cells , Humans , Macaca , Male , Narcotic Antagonists , Positron-Emission Tomography , Radioligand Assay , Radiopharmaceuticals/chemistry , Radiopharmaceuticals/pharmacokinetics , Rats , Rats, Sprague-Dawley , Receptors, Opioid/agonists , Spiro Compounds/chemistry , Spiro Compounds/pharmacokinetics , Stereoisomerism , Structure-Activity Relationship , Tandem Mass Spectrometry , Thiophenes/chemistry , Thiophenes/pharmacokinetics , Nociceptin ReceptorABSTRACT
The bioavailability, fraction of dose that reaches systemic circulation, of orally administered drugs is often limited by both physical barriers of the intestine (e.g., unstirred-water and mucosal layers, epithelial tight junctions) as well as biochemical barriers such as cytochromes P450 (CYP) and P-glycoprotein (P-gp). Highly expressed in intestine and liver, CYP and P-gp can limit the systemic-availability of parent-drug by metabolism and efflux, respectively, by means of similarly large and flexible active sites that accommodate a variety of structurally-diverse, lipophilic molecules over a wide-range of molecular weights. Consequently, many molecules that are substrates for CYP3A4 also demonstrate affinity for P-gp and numerous studies have reported that for these dual-substrates, CYP3A4 and P-gp afford an interplay that affects bioavailability and clearance in a manner that is non-linear. Several in vitro and in situ models of metabolism and permeability, including transfected cell lines, isolated tissues and perfused organs as well as computational models including physiologically-based pharmacokinetic models of such co-expressing systems have demonstrated this phenomenon of CYP3A/Pgp interplay. Furthermore, recent availability of ligand bound X-ray co-crystal structures of the CYP3A4 and P-gp binding sites coupled with computational docking techniques and other validated in silico models, provide medicinal chemists with tools to inform structural-design modifications that can modify the interaction with one or both proteins. This article provides a review of relevant in silico, in vitro, ex vivo and in situ models that allow for investigation of the extent to which clearance or bioavailability can be affected by CYP/P-gp interplay.
Subject(s)
ATP Binding Cassette Transporter, Subfamily B, Member 1/metabolism , Cytochrome P-450 CYP3A/metabolism , Models, Biological , ATP Binding Cassette Transporter, Subfamily B, Member 1/chemistry , Administration, Oral , Animals , Binding Sites , Biological Availability , Computer Simulation , Cytochrome P-450 CYP3A/chemistry , Humans , Pharmaceutical Preparations/administration & dosage , Pharmaceutical Preparations/metabolism , Protein Conformation , Tissue DistributionABSTRACT
Modern drug discovery chemical libraries contain a large number of molecular entities exhibiting low aqueous solubility, often necessitating the inclusion of solubilizing agents in preclinical models of absorption or metabolism. The objective of the present study was to investigate the effects of several commonly used polyethoxylated solubilizing agents on P450 (CYP) 3A and P-glycoprotein (P-gp) in the rat intestinal mucosa. Atenolol and verapamil were administered in the in situ perfused rat intestine or incubated with rat intestinal microsomes in the presence or absence of polyethylene glycol (PEG) 400 (2% or 20%, v/v) D-alpha-tocopheryl polyethylene glycol-1000 succinate (TPGS; 100 microg/mL), Cremophor EL (47.5 microg/mL) or polysorbate (Tween) 80 (25 microg/mL). Effects on the absorption of unchanged drug were minimal, with the exception of Tween 80 which caused a 5.0-fold increase in paracellular absorption. Rat intestinal CYP3A was significantly inhibited by PEG-400 and in situ, exceeded inhibition observed with ketoconazole. Cremophor and TPGS increased the fraction of norverapamil in the plasma, consistent with excipient-mediated inhibition of P-gp. These results suggest that caution be exercised when these solubilizing agents are included in preclinical oral dosing solutions as the perturbation of drug absorption barriers may heighten the risk of incorrectly classifying drug candidate PK-parameters.
Subject(s)
Excipients/pharmacology , Intestinal Absorption/drug effects , Intestinal Mucosa/drug effects , Intestinal Mucosa/metabolism , Pharmaceutical Preparations/metabolism , Polyethylene Glycols/pharmacology , ATP Binding Cassette Transporter, Subfamily B, Member 1/metabolism , Adrenergic beta-Antagonists/administration & dosage , Adrenergic beta-Antagonists/pharmacokinetics , Animals , Atenolol/administration & dosage , Atenolol/pharmacokinetics , Biotransformation , Calcium Channel Blockers/administration & dosage , Calcium Channel Blockers/pharmacokinetics , Chemistry, Pharmaceutical , Chromatography, High Pressure Liquid , Cytochrome P-450 CYP3A/metabolism , Excipients/chemistry , In Vitro Techniques , Indicators and Reagents , Male , Mass Spectrometry , Microsomes/metabolism , Permeability , Polyethylene Glycols/chemistry , Rats , Rats, Sprague-Dawley , Verapamil/administration & dosage , Verapamil/pharmacokineticsABSTRACT
Modulation of intestinal drug absorption barriers can have a profound impact on the bioavailability of orally administered compounds. With its commonality of use as an absorption model, it is valuable to assess the role of such barriers in the rat intestinal mucosa. In the present study, atenolol and verapamil were concomitantly delivered in the in situ perfused rat intestine in the presence or absence of inhibitors to simultaneously assess the function and modulation of passive diffusion barriers, cytochrome P450 (CYP)3A metabolism and P-glycoprotein (P-gp) efflux. A high performance liquid chromatography-tandem mass spectrometry method measured atenolol, verapamil and the CYP3A-mediated metabolite, norverapamil, with linearity (r(2) > 0.99), precision (CV Subject(s)
Intestinal Absorption/physiology
, Intestinal Mucosa/metabolism
, Pharmaceutical Preparations/metabolism
, ATP Binding Cassette Transporter, Subfamily B, Member 1/antagonists & inhibitors
, Algorithms
, Animals
, Chromatography, High Pressure Liquid
, Cytochrome P-450 CYP3A/metabolism
, Cytochrome P-450 CYP3A Inhibitors
, Enzyme Inhibitors/pharmacology
, Indicators and Reagents
, Intestinal Mucosa/drug effects
, Male
, Mass Spectrometry
, Microsomes/drug effects
, Microsomes/metabolism
, Perfusion
, Pharmacokinetics
, Rats
, Rats, Sprague-Dawley
, Reference Standards
, Reproducibility of Results
, Tight Junctions/drug effects
, Tight Junctions/metabolism
ABSTRACT
The absorption of drug molecules is often investigated using in vitro or in situ models of the intestinal mucosa; however, few studies have quantified the kinetics that limit absorption. The objective of this study was to quantify kinetic rates of rat intestinal absorption, metabolism, and efflux using nonlinear mixed effects modeling. A multicompartment model accurately described the absorption and distribution of atenolol and verapamil as well as the metabolism of verapamil and distribution of the metabolite, norverapamil. The accurate description of atenolol data required inclusion of an intermediate compartment in addition to paracellular clearance, whereas verapamil and norverapamil were modeled in the absence of paracellular clearance. The absorption of verapamil was well characterized by linear kinetics, whereas the formation and distribution of norverapamil were well characterized by Michaelis-Menten kinetics. The model identified EDTA as a modulator of physical barriers, ketoconazole as an inhibitor of cytochrome P450 3A and P-glycoprotein (P-gp), and PSC-833 and GF-120918 as specific P-gp inhibitors. These results demonstrate the utility of a physiologically based model to characterize (i) the drug distribution across the in situ perfused rat intestine and (ii) the effect of chemical modulators in this biological system.
Subject(s)
Intestinal Absorption/physiology , Intestinal Mucosa/metabolism , Pharmaceutical Preparations/metabolism , ATP Binding Cassette Transporter, Subfamily B, Member 1/antagonists & inhibitors , ATP Binding Cassette Transporter, Subfamily B, Member 1/metabolism , Acridines/pharmacology , Adrenergic beta-Antagonists/administration & dosage , Adrenergic beta-Antagonists/pharmacokinetics , Algorithms , Animals , Atenolol/administration & dosage , Atenolol/pharmacokinetics , Biotransformation , Computer Simulation , Cyclosporins/pharmacology , Cytochrome P-450 CYP3A Inhibitors , Enzyme Inhibitors/pharmacology , Ketoconazole/pharmacology , Models, Statistical , Rats , Tetrahydroisoquinolines/pharmacology , Verapamil/administration & dosage , Verapamil/analogs & derivatives , Verapamil/metabolism , Verapamil/pharmacokineticsABSTRACT
Fenbuconazole, a triazole fungicide, has been associated with an increase in the incidence of liver adenomas in female mice following long-term dietary exposure. The aim of this study was to evaluate whether the mode of action for liver tumor formation by fenbuconazole is similar to that of phenobarbital. Treatment of CD1 mice with 0, 20, 60, 180 or 1300 ppm fenbuconazole for up to 4 weeks caused a dose-dependent increase in liver weight that was associated with centrilobular hepatocellular hypertrophy, cytoplasmic eosinophilia and panlobular hepatocellular vacuolation, as well as an initial increase in the cell proliferation labeling index. Fenbuconazole also caused a dose-dependent increase in liver microsomal cytochromes b(5) and P450 and the levels of immunoreactive CYP2B10 and its associated activity 7-pentoxyresorufin O-dealkylation (PROD). Treatment of mice with 1000 ppm phenobarbital elicited the same effects as treatment of mice with 1300 ppm fenbuconazole, except that phenobarbital was more effective than fenbuconazole at inducing PROD activity, even though fenbuconazole induced CYP2B10 to the same extent as did phenobarbital. This difference was attributed to the ability of fenbuconazole to bind tightly to CYP2B10 and partially mask its catalytic activity in liver microsomes, which is characteristic of several azole-containing drugs. All hepatocellular changes and induced enzyme activity returned to control levels within 4 weeks of discontinuing treatment with fenbuconazole or phenobarbital, indicating that the observed changes were fully reversible. We conclude that fenbuconazole is a phenobarbital-type inducer of mouse liver cytochrome P450, and the mode of action by which fenbuconazole induces liver adenomas in mice is similar to that of phenobarbital.
Subject(s)
Cell Proliferation/drug effects , Enzyme Induction/drug effects , Liver/drug effects , Nitriles/toxicity , Triazoles/toxicity , Animals , Aryl Hydrocarbon Hydroxylases/metabolism , Blotting, Western , Body Weight/drug effects , Cell Enlargement/drug effects , Cytochrome P-450 CYP2B1/metabolism , Cytochrome P450 Family 2 , Cytochromes b5/metabolism , Dose-Response Relationship, Drug , Female , Fungicides, Industrial/administration & dosage , Fungicides, Industrial/toxicity , Hepatocytes/drug effects , Hepatocytes/metabolism , Hepatocytes/pathology , Liver/metabolism , Liver/pathology , Liver Neoplasms/chemically induced , Liver Neoplasms/metabolism , Mice , Nitriles/administration & dosage , Organ Size/drug effects , Phenobarbital/administration & dosage , Phenobarbital/toxicity , Steroid Hydroxylases/metabolism , Time Factors , Triazoles/administration & dosageABSTRACT
We have measured cytochrome P450 (CYP) activity in nearly 150 samples of human liver microsomes and 64 samples of cryopreserved human hepatocytes, and we have performed induction studies in over 90 preparations of cultured human hepatocytes. We have analyzed these data to examine whether the expression of CYP enzyme activity in liver microsomes and isolated hepatocytes or the inducibility of CYP enzymes in cultured hepatocytes is influenced by the gender, age, or ethnicity of the donor (the latter being limited to Caucasians, African Americans, and Hispanics due to a paucity of livers from Asian donors). In human liver microsomes, there were no statistically significant differences (P > 0.05) in CYP activity as a function of age, gender, or ethnicity with one exception. 7-Ethoxyresorufin O-dealkylase (CYP1A2) activity was greater in males than females, which is consistent with clinical observation. Liver microsomal testosterone 6beta-hydroxylase (CYP3A4) activity was slightly greater in females than males, but the difference was not significant. However, in cryopreserved human hepatocytes, the gender difference in CYP3A4 activity (females = twice males) did reach statistical significance, which supports the clinical observation that females metabolize certain CYP3A4 substrates faster than do males. Compared with those from Caucasians and African Americans, liver microsomes from Hispanics had about twice the average activity of CYP2A6, CYP2B6, and CYP2C8 and half the activity of CYP1A2, although this apparent ethnic difference may be a consequence of the relatively low number of Hispanic donors. Primary cultures of hepatocytes were treated with beta-naphthoflavone, an inducer of CYP1A2, phenobarbital or rifampin, both of which induce CYP2B6, CYP2C9, CYP2C19, and CYP3A4, albeit it to different extents. Induction of these CYP enzymes in freshly cultured hepatocytes did not appear to be influenced by the gender or age of the donor. Furthermore, CYP3A4 induction in hepatocytes isolated from cirrhotic liver was comparable to that in normal hepatocytes, which supports the "healthy hepatocyte, sick environment" hypothesis of liver cirrhosis. This review summarizes these findings and discusses their implications for the use of human liver microsomes and hepatocytes for in vitro studies of drug metabolism and enzyme induction, which play a key role in drug development.
Subject(s)
Cytochrome P-450 Enzyme System/metabolism , Hepatocytes/enzymology , Liver Cirrhosis/enzymology , Microsomes, Liver/enzymology , Aging/physiology , Alcohol Drinking/metabolism , Animals , Cells, Cultured , Cytochrome P-450 Enzyme System/biosynthesis , Cytochrome P-450 Enzyme System/genetics , Enzyme Induction/drug effects , Ethnicity , Female , Humans , Isoenzymes/biosynthesis , Isoenzymes/genetics , Isoenzymes/metabolism , Male , Sex Characteristics , Smoking/metabolismABSTRACT
Cultured human hepatocytes are a valuable in vitro system for evaluating new molecular entities as inducers of cytochrome P450 (P450) enzymes. The present study summarizes data obtained from 62 preparations of cultured human hepatocytes that were treated with vehicles (saline or dimethylsulfoxide, 0.1%), beta-naphthoflavone (33 microM), phenobarbital (100 or 250 microM), isoniazid (100 microM) and/or rifampin (20 or 50 microM), and examined for the expression of P450 enzymes based on microsomal activity toward marker substrates, or in the case of CYP2C8, the level of immunoreactive protein. The results show that CYP1A2 activity was markedly induced by beta-naphthoflavone (on average 13-fold, n = 28 preparations), and weakly induced by phenobarbital (1.9-fold, n = 25) and rifampin (2.3-fold, n = 22); CYP2A6 activity tended to be increased with phenobarbital (n = 7) and rifampin (n = 3) treatments, but the effects were not statistically significant; CYP2B6 was induced by phenobarbital (6.5-fold, n = 13) and rifampin (13-fold, n = 14); CYP2C8 was induced by phenobarbital (4.0-fold, n = 4) and rifampin (5.2-fold, n = 4); CYP2C9 was induced by phenobarbital (1.8-fold, n = 14) and rifampin (3.5-fold, n = 10); CYP2C19 was markedly induced by rifampin (37-fold, n = 10), but relatively modestly by phenobarbital (7-fold, n = 9); CYP2D6 was not significantly induced by phenobarbital (n = 5) or rifampin (n = 5); CYP2E1 was induced by phenobarbital (1.7-fold, n = 5), rifampin (2.2-fold, n = 5), and isoniazid (2.3-fold, n = 5); and, CYP3A4 was induced by phenobarbital (3.3-fold, n = 42) and rifampin (10-fold, n = 61), but not by beta-naphthoflavone. Based on these observations, we generalize that beta-naphthoflavone induces CYP1A2 and isoniazid induces CYP2E1, whereas rifampin and, to a lesser extent phenobarbital, tend to significantly and consistently induce enzymes of the CYP2A, CYP2B, CYP2C, CYP2E, and CYP3A subfamilies but not the 2D subfamily.