CYP2B6 and Drug Interactions FDA Advisory Committee for Pharmaceutical Sciences Clinical Pharmacology Subcommittee November 18th, 2003. Zeruesenay Desta PhD David A Flockhart MD, PhD Indiana University School of Medicine

Outline CYP2B6 expression CYP2B6 substrates CYP2B6 inhibitors In vitro In vivo? CYP2B6 inhibitors CYP inducers Relevant in vivo data

CYP2B6 expression: Late 1990s Low level of protein expression Minor component of the total hepatic CYPs (<1% of total P450 in the liver) Thought to play a relatively minor role in human drug metabolism

CYP2B6 Expression: Initial Studies Detection (%) Pmol/mg protein (range) References 13/13 (100%)**   Yamano et al 1989 12/12 (100%)* Forrester et al., 1992 12/14 (86%), Thai Kirby et al 1993 12/50 (24%) 1.41.5 (0-7) Mimura et al 1993 9/30 (30%) (Japanese) 0.25 (0-2) Shimada et al 1994 26/30 (87%), Caucasian 1.0 (0-2.5) 2/10 (20%) Baker et al 1995 ?/40, Japanese 0.140.62 Guengerich et al 1995 ?/32, Caucasian 1.41.8 ?/16 3 (0-17) Imaoka et al 1996 8/30 (27%) Boobis et al 1996 High level in Thai Kim et al 1997 7/10 (70%) 19.3623.9) Tateishi et al 1997 2.655.94 6/30 (20%) Edwards et al 1998

CYP2B6 Expression: Recent Studies Detection (%) Pmol/mg protein (range) Variability References 17/17 (100) 1521 (0.5-70) 247-fold Code et al. 1997 24/26 (92) 10.99.5 (0-28)   Yang et al., 1998 19/19 (100) 9.7717.6 (0.7-71.1) 107-fold Ekins et al 1998 28/28 (100) 24.9 18.2 (2-82) 41-fold Stresser et al, 1999 43/48 (90) 1.7 (0.4-8, n=43 livers) 20 Gervot et al 1999 15/16 (94) 100-fold (n=15) Faucette et al 2000 12/12 (100) 44.6 (1.5-148.4) 99-fold Hesse et al 2000 136/136 (100) 16.916.6 (0.5-95.5) 191-fold Lang et al. 2001

CYP2B6 Expression: 2003 New mono- and polyclonal antibodies of higher sensitivity and specificity for CYP2B6 protein immuno-quantification: Greater frequency of detection (almost in all human livers tested) Higher protein quantities: Average: ~6% of the total liver CYP450 content Absolute maximum amounts: up to 25 to 43.5%

The CYP2B6 gene CYP2B subfamily: CYP2B6 gene (functional) CYP2B7 gene (nonfunctional) CYP2B7-like pseudogene) Mapped to chromosome 19 [19q12 and 19q13.2] Contains 9 exons - encodes a protein with 491 amino acids

CYP2B6: substrate drugs (in vitro) HIV medications: Nevirapine Efavirenz DPC 963 Anticancer drugs: Cyclophosphamide Ifosfamide Tamoxifen ThioTEPA CNS acting drugs: S-mephobarbital Propofol Bupropion Ketamine Selegiline Methadone S-mephenytoin Other drugs: RP 73401 Artemisinin Benzphetamine Cinnarizine Diazepam Clopidogrel Lidocaine N-deethylation Ticlopidine Dextromethorphan R-mianserin Benzyloxyresorufin Antipyrine 4-Chloromethyl-7-ethoxycoumarin 3-Cyano-7-ethoxycoumarin 7-ethoxycoumarin 7-Ethoxy-4-trifluoromethylcoumarin Imipramine methoxyflurane Sertraline

CYP2B6: Xenobiotics CYP2B6: endogenous substances Testosterone Estrone Styrene 6-aminochrysene Dibenzo[a,h]anthracene Aflatoxin B Nicotine Methylenedioxymethamphetamine (ecstasy) 2-dibromoethane benzo[a]pyrene Phenanthrene Methoxychlor 1,3-butadiene 2-chloro-1,1-difluoroethene azinphos-methyl (AZIN) chlorpyrifos (CPF) diazinon (DIA) parathion (PAR), CYP2B6: endogenous substances Testosterone Estrone 17beta-estadiol

Human metabolism of efavirenz (Ward et al., J Pharmacol Exp Ther 2003)

CYP2B6 catalyses the metabolism of efavirienz (1µM)

Effect of rifampin on efavirenz in healthy volunteers

CYP2B6 Catalyses metabolism of an efavirenz analog: DPC 963 Drug Metab Dispos. 2003 Jan;31(1):122-32.

CYP2B6 is a low affinity catalyst of S-mephenytoin metabolism to Nirvanol Km = 564µM Heyn et al Drug Metab Dispos 1996;24:948-954

N-Demethylation of S-mephenytoin (200µM) by CYP2C9 and CYP2B6 Ko, Desta and Flockhart. Drug Metab Dispos 1998;26(8):775-778

Human Metabolism of Mephenytoin

R-mephenytoin as CYP2B6 substrate probe?

Inhibitors of CYP2B6 Inhibitors: Antidepressants (e.g. paroxetine and sertraline) Antiretrovirals (e.g. nelfinavir and ritonavir) Ticlopidine and clopidogrel ThioTEPA

ThioTEPA is a specific cytochrome P450 inhibitor in vitro IC50 5µM

Inhibition of CYP2B6 by thioTEPA Ki=4.8 ± 0.3 µM (HLMs) Ki=6.2 ± 0.7 µM (CYP2B6) ThioTEPA therapeutic concentration: 1.1-18.6 µM

Cyclophosphamide activation

Effect of thioTEPA on the PK of CPA (Huitema et al. Cancer Chemother Pharmacol 2000;46:119-127)

HMG-CoA reductase inhibitors Inducers of CYP2B6 Rifampin Hyperforin Phenobarbital Ritonavir Phenytoin Carbamazepine HMG-CoA reductase inhibitors Nevirapine Efavirenz Clotrimazole Artemisinin

Conclusions CYP2B6 is a significant contributor to hepatic CYP expression The number of substrate drugs for CYP2B6 is growing rapidly Efavirenz and buproprion are specific in vitro probes for CYP2B6 activity ThioTEPA is a specific in vitro inhibitor of CYP2B6 No valuable, specific inhibitors of CYP2B6 in vivo have been demonstrated to date Efavirenz is a potentially valuable in vivo probe for CYP2B6 activity

Effect of rifampin on efavirenz in healthy volunteers