Time dependent inhibition of CYP3A4 (unknown origin) at 10 uM by LC/MS system
—
Inhibition < 10.0 %
Reference
Title : Combination of GSH trapping and time-dependent inhibition assays as a predictive method of drugs generating highly reactive metabolites. Authors : Nakayama S, Takakusa H, Watanabe A, Miyaji Y, Suzuki W, Sugiyama D, Shiosakai K, Honda K, Okudaira N, Izumi T, Okazaki O. Publication Year :
2011 Journal : Drug Metab. Dispos. Abstract : Covalent binding (CB) of reactive metabolites (RMs) is potentially involved in severe adverse drug reactions. Because the CB assay is of low throughput and costly, a qualitative trapping assay using agents such as [(35)S]GSH is often performed in the early stages of drug discovery. However, trapping methods alone cannot replace the CB assay. We hypothesized that the time-dependent inhibition (TDI) assay might be complementary to the [(35)S]GSH trapping assay in detecting RMs. We performed CB assays, [(35)S]GSH trapping assays, and TDI assays for 42 structurally diverse compounds. First, we showed that the [(35)S]GSH trapping assay alone does not correlate with the extent of CB. Four compounds that the [(35)S]GSH trapping assay failed to detect but that showed high extent of CB were inactivators of the enzyme in the TDI assay. There was a tendency for compounds judged as positive in the TDI assay to show a high degree of CB irrespective of the result of the [(35)S]GSH trapping assay. Finally, to combine parameters from the two assays, we introduced intrinsic clearance to describe the formation of RMs (CL(int, RMs)). The Spearman rank correlation coefficient between the extent of CB and CL(int, RMs) was 0.77 (p < 0.0001), which was better than that for the formation rates of [(35)S]GSH adducts. Therefore, we demonstrated that a combination of the [(35)S]GSH trapping and TDI assays is an effective method for detecting compounds potentially capable of generating highly reactive metabolites in the early stages of drug discovery. DOI : 10.1124/dmd.111.039180
ACETAMINOPHEN
Inhibition of CYP3A4 (unknown origin) in baculosomes preincubated for 20 mins followed by addition of CYP enzyme-specific substrate and NADP+ and measured after 30 mins by fluorescence based analysis relative to control
—
Inhibition = 50.0 %
Reference
Title : A novel pipeline of 2-(benzenesulfonamide)-N-(4-hydroxyphenyl) acetamide analgesics that lack hepatotoxicity and retain antipyresis. Authors : Bazan HA,Bhattacharjee S,Burgos C,Recio J,Abet V,Pahng AR,Jun B,Heap J,Ledet AJ,Gordon WC,Edwards S,Paul D,Alvarez-Builla J,Bazan NG Publication Year :
2020 Journal : Eur J Med Chem Abstract : Although acetaminophen (ApAP) is one of the most commonly used medicines worldwide, hepatotoxicity is a risk with overdose or in patients with compromised liver function. ApAP overdose is the most common cause of acute fulminant hepatic failure. Oxidation of ApAP to N-acetyl-p-benzoquinone imine (NAPQI) is the mechanism for hepatotoxicity. 1 is a non-hepatotoxic, metabolically unstable lipophilic ApAP analog that is not antipyretic. The newly synthesized 3 is a non-hepatotoxic ApAP analog that is stable, lipophilic, and retains analgesia and antipyresis. Intraperitoneal or po administration of the new chemical entities (NCEs), 3b and 3r, in concentrations equal to a toxic dose of ApAP did not result in the formation of NAPQI. Unlike livers from NCE-treated mice, the livers from ApAP-treated mice demonstrated large amounts of nitrotyrosine, a marker of mitochondrial free radical formation, and loss of hepatic tight junction integrity. Given the widespread use of ApAP, hepatotoxicity risk with overuse, and the ongoing opioid epidemic, these NCEs represent a novel, non-narcotic therapeutic pipeline. DOI : 10.1016/j.ejmech.2020.112600
AMPRENAVIR
Mechanism based inhibition of human cytochrome P450 3A4 measured by testosterone hydroxylation
—
Ki = 260.0 nM
Reference
Title : Cytochrome p450 enzymes mechanism based inhibitors: common sub-structures and reactivity. Authors : Fontana E, Dansette PM, Poli SM. Publication Year :
2005 Journal : Curr. Drug Metab. Abstract : The inhibition of human cytochrome P450s (CYPs) is one of the most common mechanisms which can lead to drug-drug interactions. The inhibition of CYPs can be reversible (competitive or non-competitive) or irreversible. Irreversible inhibition usually derives from activation of a drug by CYPs into a reactive metabolite, which tightly binds to the enzyme active site, leading to a long lasting inactivation. This process is called "mechanism based inhibition" or "suicide inhibition". The irreversible inactivation usually implies the formation of a covalent bond between the metabolite and the enzyme, which can lead to hapten formation and can in some cases trigger an autoimmune-response. For these reasons it is of utmost importance to study the mechanism of the CYP inhibition of new potential drugs as early as possible during the drug discovery process. The literature on CYPs is vast and covers numerous aspects of their biology and biochemistry, however to our knowledge there is no general and systematic review focusing on mechanism-based inhibitors; we have reviewed the literature and compiled all the available data on chemical entities, which are known to be CYP suicide inhibitors. Each compound is reported together with its chemical structure, the CYP isoform and the parameters describing the inactivation. Literature references are reported together with their PMID (PubMed ID number) to allow a fast retrieval of the papers. This review offers a quick reference to help predict liabilities of new chemical entities without carrying out extensive in vitro work, and will hopefully help in designing safer drugs. DOI : 10.2174/138920005774330639
AMPRENAVIR
Time dependent inhibition of CYP3A4-mediated testosterone-6-beta hydroxylation in human liver microsome
Liver
Ki = 370.0 nM
Reference
Title : Mechanism-based inactivation (MBI) of cytochrome P450 enzymes: structure-activity relationships and discovery strategies to mitigate drug-drug interaction risks. Authors : Orr ST, Ripp SL, Ballard TE, Henderson JL, Scott DO, Obach RS, Sun H, Kalgutkar AS. Publication Year :
2012 Journal : J. Med. Chem. DOI : 10.1021/jm300065h
APREPITANT
Inhibition of human CYP3A4
—
IC50 = 46.0 nM
Reference
Title : Pyrrolidine-carboxamides and oxadiazoles as potent hNK1 antagonists. Authors : Young JR, Eid R, Turner C, DeVita RJ, Kurtz MM, Tsao KL, Chicchi GG, Wheeldon A, Carlson E, Mills SG. Publication Year :
2007 Journal : Bioorg. Med. Chem. Lett. Abstract : The preparation and structure-activity-relationships of novel pyrrolidine-carboxamides and oxadiazoles are described. Compounds in this series were found to be potent hNK(1) antagonists in vitro and efficacious in vivo with minimal interactions with P(450) liver enzymes. Oxadiazole analog 22 was determined to have excellent hNK(1) binding affinity, functional activity, and a good PD response in vivo. DOI : 10.1016/j.bmcl.2007.08.028
CARBAMAZEPINE
Time dependent inhibition of CYP3A4 (unknown origin) at 10 uM by LC/MS system
—
Inhibition < 10.0 %
Reference
Title : Combination of GSH trapping and time-dependent inhibition assays as a predictive method of drugs generating highly reactive metabolites. Authors : Nakayama S, Takakusa H, Watanabe A, Miyaji Y, Suzuki W, Sugiyama D, Shiosakai K, Honda K, Okudaira N, Izumi T, Okazaki O. Publication Year :
2011 Journal : Drug Metab. Dispos. Abstract : Covalent binding (CB) of reactive metabolites (RMs) is potentially involved in severe adverse drug reactions. Because the CB assay is of low throughput and costly, a qualitative trapping assay using agents such as [(35)S]GSH is often performed in the early stages of drug discovery. However, trapping methods alone cannot replace the CB assay. We hypothesized that the time-dependent inhibition (TDI) assay might be complementary to the [(35)S]GSH trapping assay in detecting RMs. We performed CB assays, [(35)S]GSH trapping assays, and TDI assays for 42 structurally diverse compounds. First, we showed that the [(35)S]GSH trapping assay alone does not correlate with the extent of CB. Four compounds that the [(35)S]GSH trapping assay failed to detect but that showed high extent of CB were inactivators of the enzyme in the TDI assay. There was a tendency for compounds judged as positive in the TDI assay to show a high degree of CB irrespective of the result of the [(35)S]GSH trapping assay. Finally, to combine parameters from the two assays, we introduced intrinsic clearance to describe the formation of RMs (CL(int, RMs)). The Spearman rank correlation coefficient between the extent of CB and CL(int, RMs) was 0.77 (p < 0.0001), which was better than that for the formation rates of [(35)S]GSH adducts. Therefore, we demonstrated that a combination of the [(35)S]GSH trapping and TDI assays is an effective method for detecting compounds potentially capable of generating highly reactive metabolites in the early stages of drug discovery. DOI : 10.1124/dmd.111.039180
CELECOXIB
Time dependent inhibition of CYP3A4 (unknown origin) at 10 uM by LC/MS system
—
Inhibition < 10.0 %
Reference
Title : Combination of GSH trapping and time-dependent inhibition assays as a predictive method of drugs generating highly reactive metabolites. Authors : Nakayama S, Takakusa H, Watanabe A, Miyaji Y, Suzuki W, Sugiyama D, Shiosakai K, Honda K, Okudaira N, Izumi T, Okazaki O. Publication Year :
2011 Journal : Drug Metab. Dispos. Abstract : Covalent binding (CB) of reactive metabolites (RMs) is potentially involved in severe adverse drug reactions. Because the CB assay is of low throughput and costly, a qualitative trapping assay using agents such as [(35)S]GSH is often performed in the early stages of drug discovery. However, trapping methods alone cannot replace the CB assay. We hypothesized that the time-dependent inhibition (TDI) assay might be complementary to the [(35)S]GSH trapping assay in detecting RMs. We performed CB assays, [(35)S]GSH trapping assays, and TDI assays for 42 structurally diverse compounds. First, we showed that the [(35)S]GSH trapping assay alone does not correlate with the extent of CB. Four compounds that the [(35)S]GSH trapping assay failed to detect but that showed high extent of CB were inactivators of the enzyme in the TDI assay. There was a tendency for compounds judged as positive in the TDI assay to show a high degree of CB irrespective of the result of the [(35)S]GSH trapping assay. Finally, to combine parameters from the two assays, we introduced intrinsic clearance to describe the formation of RMs (CL(int, RMs)). The Spearman rank correlation coefficient between the extent of CB and CL(int, RMs) was 0.77 (p < 0.0001), which was better than that for the formation rates of [(35)S]GSH adducts. Therefore, we demonstrated that a combination of the [(35)S]GSH trapping and TDI assays is an effective method for detecting compounds potentially capable of generating highly reactive metabolites in the early stages of drug discovery. DOI : 10.1124/dmd.111.039180
CLOBETASOL PROPIONATE
Inhibition of human recombinant CYP3A4 expressed in supersomes assessed as decrease in formation of D-luciferin using luciferin-IPA as substrate incubated for 10 mins in presence of NADPH by P450-Glo luminescence assay
—
IC50 = 206.0 nM
Reference
Title : Clobetasol Propionate Is a Heme-Mediated Selective Inhibitor of Human Cytochrome P450 3A5. Authors : Wright WC, Chenge J, Wang J, Girvan HM, Yang L, Chai SC, Huber AD, Wu J, Oladimeji PO, Munro AW, Chen T. Publication Year :
2020 Journal : J Med Chem Abstract : The human cytochrome P450 (CYP) enzymes CYP3A4 and CYP3A5 metabolize most drugs and have high similarities in their structure and substrate preference. Whereas CYP3A4 is predominantly expressed in the liver, CYP3A5 is upregulated in cancer, contributing to drug resistance. Selective inhibitors of CYP3A5 are, therefore, critical to validating it as a therapeutic target. Here we report clobetasol propionate (clobetasol) as a potent and selective CYP3A5 inhibitor identified by high-throughput screening using enzymatic and cell-based assays. Molecular dynamics simulations suggest a close proximity of clobetasol to the heme in CYP3A5 but not in CYP3A4. UV-visible spectroscopy and electron paramagnetic resonance analyses confirmed the formation of an inhibitory type I heme-clobetasol complex in CYP3A5 but not in CYP3A4, thus explaining the CYP3A5 selectivity of clobetasol. Our results provide a structural basis for selective CYP3A5 inhibition, along with mechanistic insights, and highlight clobetasol as an important chemical tool for target validation. DOI : 10.1021/acs.jmedchem.9b02067
CLOTRIMAZOLE
Inhibition of CYP3A4 in human liver microsomes using midazolam as substrate after 8 mins by LC-MS/MS analysis
Liver
IC50 < 100.0 nM
Reference
Title : Identifying a selective substrate and inhibitor pair for the evaluation of CYP2J2 activity. Authors : Lee CA, Jones JP, Katayama J, Kaspera R, Jiang Y, Freiwald S, Smith E, Walker GS, Totah RA. Publication Year :
2012 Journal : Drug Metab. Dispos. Abstract : CYP2J2, an arachidonic acid epoxygenase, is recognized for its role in the first-pass metabolism of astemizole and ebastine. To fully assess the role of CYP2J2 in drug metabolism, a selective substrate and potent specific chemical inhibitor are essential. In this study, we report amiodarone 4-hydoxylation as a specific CYP2J2-catalyzed reaction with no CYP3A4, or other drug-metabolizing enzyme, involvement. Amiodarone 4-hydroxylation enabled the determination of liver relative activity factor and intersystem extrapolation factor for CYP2J2. Amiodarone 4-hydroxylation correlated with astemizole O-demethylation but not with CYP2J2 protein content in a sample of human liver microsomes. To identify a specific CYP2J2 inhibitor, 138 drugs were screened using terfenadine and astemizole as probe substrates with recombinant CYP2J2. Forty-two drugs inhibited CYP2J2 activity by ≥50% at 30 μM, but inhibition was substrate-dependent. Of these, danazol was a potent inhibitor of both hydroxylation of terfenadine (IC(50) = 77 nM) and O-demethylation of astemizole (K(i) = 20 nM), and inhibition was mostly competitive. Danazol inhibited CYP2C9, CYP2C8, and CYP2D6 with IC(50) values of 1.44, 1.95, and 2.74 μM, respectively. Amiodarone or astemizole were included in a seven-probe cocktail for cytochrome P450 (P450) drug-interaction screening potential, and astemizole demonstrated a better profile because it did not appreciably interact with other P450 probes. Thus, danazol, amiodarone, and astemizole will facilitate the ability to determine the metabolic role of CYP2J2 in hepatic and extrahepatic tissues. DOI : 10.1124/dmd.111.043505
CLOTRIMAZOLE
Inhibition of recombinant human CYP3A4 using Luciferin-PPXE as substrate preincubated for 10 mins followed by NADPH addition measured after 15 mins by luminometric method
—
IC50 = 67.0 nM
Reference
Title : Antifungal activity, mode of action variability, and subcellular distribution of coumarin-based antifungal azoles. Authors : Elias R, Benhamou RI, Jaber QZ, Dorot O, Zada SL, Oved K, Pichinuk E, Fridman M. Publication Year :
2019 Journal : Eur J Med Chem Abstract : Azole antifungals inhibit the biosynthesis of ergosterol, the fungal equivalent of cholesterol in mammalian cells. Here we report an investigation of the activity of coumarin-substituted azole antifungals. Screening against a panel of Candida pathogens, including a mutant lacking CYP51, the target of antifungal azoles, revealed that this enzyme is inhibited by triazole-based antifungals, whereas imidazole-based derivatives have more than one mode of action. The imidazole-bearing antifungals more effectively reduced trailing growth associated with persistence and/or recurrence of fungal infections than triazole-based derivatives. The imidazole derivatives were more toxic to mammalian cells and more potently inhibited the activity of CYP3A4, which is one of the main causes of azole toxicity. Using live cell imaging, we showed that regardless of the type of azole ring fluorescent 7-diethylaminocoumarin-based azoles localized to the endoplasmic reticulum, the organelle that harbors CYP51. This study suggests that the coumarin is a promising scaffold for development of novel azole-based antifungals that effectively localize to the fungal cell endoplasmic reticulum. DOI : 10.1016/j.ejmech.2019.07.003
CLOTRIMAZOLE
Inhibition of CYP3A4 (unknown origin)
—
IC50 = 50.0 nM
Reference
Title : Strategies for the development of highly selective cytochrome P450 inhibitors: Several CYP targets in current research. Authors : Zhao L, Sun N, Tian L, Zhao S, Sun B, Sun Y, Zhao D. Publication Year :
2019 Journal : Bioorg Med Chem Lett Abstract : Cytochromes P450 (CYPs) play an important role in the metabolism of endogenic and xenobiotic substances, especially drugs. In addition, many CYPs may serve as targets for disease treatment. However, due to the presence of a common heme, the hydrophobicity of the CYP binding cavity, and the high homology within the binding pocket, most CYP inhibitors lack selectivity, which often leads to drug-drug interactions. Therefore, it is meaningful to develop highly selective CYP inhibitors. In this review, we summarize some of the strategies that have been used to develop highly selective CYP inhibitors, such as the weakening of the heme-binding group interaction, reduction of molecular lipophilicity and introduction of small structural changes within compounds. DOI : 10.1016/j.bmcl.2019.06.040
CLOZAPINE
Time dependent inhibition of CYP3A4 (unknown origin) at 100 uM by LC/MS system
—
Inhibition = 21.0 %
Reference
Title : Combination of GSH trapping and time-dependent inhibition assays as a predictive method of drugs generating highly reactive metabolites. Authors : Nakayama S, Takakusa H, Watanabe A, Miyaji Y, Suzuki W, Sugiyama D, Shiosakai K, Honda K, Okudaira N, Izumi T, Okazaki O. Publication Year :
2011 Journal : Drug Metab. Dispos. Abstract : Covalent binding (CB) of reactive metabolites (RMs) is potentially involved in severe adverse drug reactions. Because the CB assay is of low throughput and costly, a qualitative trapping assay using agents such as [(35)S]GSH is often performed in the early stages of drug discovery. However, trapping methods alone cannot replace the CB assay. We hypothesized that the time-dependent inhibition (TDI) assay might be complementary to the [(35)S]GSH trapping assay in detecting RMs. We performed CB assays, [(35)S]GSH trapping assays, and TDI assays for 42 structurally diverse compounds. First, we showed that the [(35)S]GSH trapping assay alone does not correlate with the extent of CB. Four compounds that the [(35)S]GSH trapping assay failed to detect but that showed high extent of CB were inactivators of the enzyme in the TDI assay. There was a tendency for compounds judged as positive in the TDI assay to show a high degree of CB irrespective of the result of the [(35)S]GSH trapping assay. Finally, to combine parameters from the two assays, we introduced intrinsic clearance to describe the formation of RMs (CL(int, RMs)). The Spearman rank correlation coefficient between the extent of CB and CL(int, RMs) was 0.77 (p < 0.0001), which was better than that for the formation rates of [(35)S]GSH adducts. Therefore, we demonstrated that a combination of the [(35)S]GSH trapping and TDI assays is an effective method for detecting compounds potentially capable of generating highly reactive metabolites in the early stages of drug discovery. DOI : 10.1124/dmd.111.039180
COBICISTAT
Inhibition of human CYP3A4-mdiated midazolam 1'-hydroxylase activity
—
IC50 = 150.0 nM
Reference
Title : Structure-activity relationships of diamine inhibitors of cytochrome P450 (CYP) 3A as novel pharmacoenhancers. Part II: P2/P3 region and discovery of cobicistat (GS-9350). Authors : Xu L, Liu H, Hong A, Vivian R, Murray BP, Callebaut C, Choi YC, Lee MS, Chau J, Tsai LK, Stray KM, Strickley RG, Wang J, Tong L, Swaminathan S, Rhodes GR, Desai MC. Publication Year :
2014 Journal : Bioorg. Med. Chem. Lett. Abstract : The HIV protease inhibitor (PI) ritonavir (RTV) has been widely used as a pharmacoenhancer for other PIs, which are substrates of cytochrome P450 3A (CYP3A). However the potent anti-HIV activity of ritonavir may limit its use as a pharmacoenhancer with other classes of anti-HIV agents. Ritonavir is also associated with limitations such as poor physicochemical properties. To address these issues a series of compounds with replacements at the P2 and/or P3 region was designed and evaluated as novel CYP3A inhibitors. Through these efforts, a potent and selective inhibitor of CYP3A, GS-9350 (cobicistat) with improved physiochemical properties was discovered. DOI : 10.1016/j.bmcl.2013.12.057
COLCHICINE
Inhibition of CYP3A4 (unknown origin) at 3 uM
—
Inhibition = 9.1 %
Reference
Title : New cytotoxic benzosuberene analogs. Synthesis, molecular modeling and biological evaluation. Authors : Chen Z, Maderna A, Sukuru SC, Wagenaar M, O'Donnell CJ, Lam MH, Musto S, Loganzo F. Publication Year :
2013 Journal : Bioorg. Med. Chem. Lett. Abstract : In this Letter we describe the synthesis and biological evaluation of new benzosuberene analogs with structural modifications on the B-ring. The focus was initially to probe the chemical space around the B-ring C-8 position. This position was readily available for derivatization chemistry using our recently developed new synthesis for this compound class. Furthermore, we describe two new B-ring analogs, one containing a diene and the other a cyclic ether group. Both new analogs show excellent potencies in tumor cell proliferation assays. In addition, we describe molecular modeling studies that provide a binding rationale for reference compound 8 in the colchicine binding site using the known colchicine crystal structure. We also examine whether the cell based potency data obtained with selected new analogs are supported by modeling results. DOI : 10.1016/j.bmcl.2013.10.039
DICLOFENAC
Time dependent inhibition of CYP3A4 (unknown origin) at 100 uM by LC/MS system
—
Inhibition < 10.0 %
Reference
Title : Combination of GSH trapping and time-dependent inhibition assays as a predictive method of drugs generating highly reactive metabolites. Authors : Nakayama S, Takakusa H, Watanabe A, Miyaji Y, Suzuki W, Sugiyama D, Shiosakai K, Honda K, Okudaira N, Izumi T, Okazaki O. Publication Year :
2011 Journal : Drug Metab. Dispos. Abstract : Covalent binding (CB) of reactive metabolites (RMs) is potentially involved in severe adverse drug reactions. Because the CB assay is of low throughput and costly, a qualitative trapping assay using agents such as [(35)S]GSH is often performed in the early stages of drug discovery. However, trapping methods alone cannot replace the CB assay. We hypothesized that the time-dependent inhibition (TDI) assay might be complementary to the [(35)S]GSH trapping assay in detecting RMs. We performed CB assays, [(35)S]GSH trapping assays, and TDI assays for 42 structurally diverse compounds. First, we showed that the [(35)S]GSH trapping assay alone does not correlate with the extent of CB. Four compounds that the [(35)S]GSH trapping assay failed to detect but that showed high extent of CB were inactivators of the enzyme in the TDI assay. There was a tendency for compounds judged as positive in the TDI assay to show a high degree of CB irrespective of the result of the [(35)S]GSH trapping assay. Finally, to combine parameters from the two assays, we introduced intrinsic clearance to describe the formation of RMs (CL(int, RMs)). The Spearman rank correlation coefficient between the extent of CB and CL(int, RMs) was 0.77 (p < 0.0001), which was better than that for the formation rates of [(35)S]GSH adducts. Therefore, we demonstrated that a combination of the [(35)S]GSH trapping and TDI assays is an effective method for detecting compounds potentially capable of generating highly reactive metabolites in the early stages of drug discovery. DOI : 10.1124/dmd.111.039180
DOLUTEGRAVIR
Inhibition of CYP3A4 in human liver microsomes using nifedipine as substrate at 100 uM preincubated for 20 mins with substrate prior to initiation of reaction with NADPH by HPLC analysis
Liver
Inhibition = 20.0 %
Reference
Title : In vitro investigations into the roles of drug transporters and metabolizing enzymes in the disposition and drug interactions of dolutegravir, a HIV integrase inhibitor. Authors : Reese MJ, Savina PM, Generaux GT, Tracey H, Humphreys JE, Kanaoka E, Webster LO, Harmon KA, Clarke JD, Polli JW. Publication Year :
2013 Journal : Drug Metab. Dispos. Abstract : Dolutegravir (DTG; S/GSK1349572) is a potent HIV-1 integrase inhibitor with a distinct resistance profile and a once-daily dose regimen that does not require pharmacokinetic boosting. This work investigated the in vitro drug transport and metabolism of DTG and assessed the potential for clinical drug-drug interactions. DTG is a substrate for the efflux transporters P-glycoprotein (Pgp) and human breast cancer resistance protein (BCRP). Its high intrinsic membrane permeability limits the impact these transporters have on DTG's intestinal absorption. UDP-glucuronosyltransferase (UGT) 1A1 is the main enzyme responsible for the metabolism of DTG in vivo, with cytochrome P450 (P450) 3A4 being a notable pathway and UGT1A3 and UGT1A9 being only minor pathways. DTG demonstrated little or no inhibition (IC(50) values > 30 μM) in vitro of the transporters Pgp, BCRP, multidrug resistance protein 2, organic anion transporting polypeptide 1B1/3, organic cation transporter (OCT) 1, or the drug metabolizing enzymes CYP1A2, 2A6, 2B6, 2C8, 2C9, 2C19, 2D6, 3A4, UGT1A1, or 2B7. Further, DTG did not induce CYP1A2, 2B6, or 3A4 mRNA in vitro using human hepatocytes. DTG does inhibit the renal OCT2 (IC(50) = 1.9 μM) transporter, which provides a mechanistic basis for the mild increases in serum creatinine observed in clinical studies. These in vitro studies demonstrate a low propensity for DTG to be a perpetrator of clinical drug interactions and provide a basis for predicting when other drugs could result in a drug interaction with DTG. DOI : 10.1124/dmd.112.048918
ERYTHROMYCIN
Time dependent inhibition of CYP3A4 (unknown origin) at 100 uM by LC/MS system
—
Inhibition = 45.0 %
Reference
Title : Combination of GSH trapping and time-dependent inhibition assays as a predictive method of drugs generating highly reactive metabolites. Authors : Nakayama S, Takakusa H, Watanabe A, Miyaji Y, Suzuki W, Sugiyama D, Shiosakai K, Honda K, Okudaira N, Izumi T, Okazaki O. Publication Year :
2011 Journal : Drug Metab. Dispos. Abstract : Covalent binding (CB) of reactive metabolites (RMs) is potentially involved in severe adverse drug reactions. Because the CB assay is of low throughput and costly, a qualitative trapping assay using agents such as [(35)S]GSH is often performed in the early stages of drug discovery. However, trapping methods alone cannot replace the CB assay. We hypothesized that the time-dependent inhibition (TDI) assay might be complementary to the [(35)S]GSH trapping assay in detecting RMs. We performed CB assays, [(35)S]GSH trapping assays, and TDI assays for 42 structurally diverse compounds. First, we showed that the [(35)S]GSH trapping assay alone does not correlate with the extent of CB. Four compounds that the [(35)S]GSH trapping assay failed to detect but that showed high extent of CB were inactivators of the enzyme in the TDI assay. There was a tendency for compounds judged as positive in the TDI assay to show a high degree of CB irrespective of the result of the [(35)S]GSH trapping assay. Finally, to combine parameters from the two assays, we introduced intrinsic clearance to describe the formation of RMs (CL(int, RMs)). The Spearman rank correlation coefficient between the extent of CB and CL(int, RMs) was 0.77 (p < 0.0001), which was better than that for the formation rates of [(35)S]GSH adducts. Therefore, we demonstrated that a combination of the [(35)S]GSH trapping and TDI assays is an effective method for detecting compounds potentially capable of generating highly reactive metabolites in the early stages of drug discovery. DOI : 10.1124/dmd.111.039180
ERYTHROMYCIN
Inhibition of human CYP3A4 expressed in Escherichia coli using diethoxyfluorescein substrate measured in 25 to 30 mins by time dependent inhibition assay
—
IC50 = 190.0 nM
Reference
Title : Discovery of N-(3-fluorophenyl)-1-[(4-([(3S)-3-methyl-1-piperazinyl]methyl)phenyl)acetyl]-4-piperidinamine (GSK962040), the first small molecule motilin receptor agonist clinical candidate. Authors : Westaway SM, Brown SL, Fell SC, Johnson CN, MacPherson DT, Mitchell DJ, Myatt JW, Stanway SJ, Seal JT, Stemp G, Thompson M, Lawless K, McKay F, Muir AI, Barford JM, Cluff C, Mahmood SR, Matthews KL, Mohamed S, Smith B, Stevens AJ, Bolton VJ, Jarvie EM, Sanger GJ. Publication Year :
2009 Journal : J. Med. Chem. Abstract : N-(3-fluorophenyl)-1-[(4-([(3S)-3-methyl-1-piperazinyl]methyl)phenyl)acetyl]-4-piperidinamine 12 (GSK962040) is a novel small molecule motilin receptor agonist. It possesses excellent activity at the recombinant human motilin receptor and also at the native rabbit motilin receptor where its agonist activity results in potentiation of the amplitude of neuronal-mediated contractions of isolated gastric antrum tissue. Compound 12 also possesses highly promising pharmacokinetic profiles in both rat and dog, and these results, in combination with further profiling in human native tissue and an in vivo model of gastrointestinal transit in the rabbit, have led to its selection as a candidate for further development. DOI : 10.1021/jm801332q
ETHINYL ESTRADIOL
Time dependent inhibition of CYP3A4 (unknown origin) at 50 uM by LC/MS system
—
Inhibition = 65.0 %
Reference
Title : Combination of GSH trapping and time-dependent inhibition assays as a predictive method of drugs generating highly reactive metabolites. Authors : Nakayama S, Takakusa H, Watanabe A, Miyaji Y, Suzuki W, Sugiyama D, Shiosakai K, Honda K, Okudaira N, Izumi T, Okazaki O. Publication Year :
2011 Journal : Drug Metab. Dispos. Abstract : Covalent binding (CB) of reactive metabolites (RMs) is potentially involved in severe adverse drug reactions. Because the CB assay is of low throughput and costly, a qualitative trapping assay using agents such as [(35)S]GSH is often performed in the early stages of drug discovery. However, trapping methods alone cannot replace the CB assay. We hypothesized that the time-dependent inhibition (TDI) assay might be complementary to the [(35)S]GSH trapping assay in detecting RMs. We performed CB assays, [(35)S]GSH trapping assays, and TDI assays for 42 structurally diverse compounds. First, we showed that the [(35)S]GSH trapping assay alone does not correlate with the extent of CB. Four compounds that the [(35)S]GSH trapping assay failed to detect but that showed high extent of CB were inactivators of the enzyme in the TDI assay. There was a tendency for compounds judged as positive in the TDI assay to show a high degree of CB irrespective of the result of the [(35)S]GSH trapping assay. Finally, to combine parameters from the two assays, we introduced intrinsic clearance to describe the formation of RMs (CL(int, RMs)). The Spearman rank correlation coefficient between the extent of CB and CL(int, RMs) was 0.77 (p < 0.0001), which was better than that for the formation rates of [(35)S]GSH adducts. Therefore, we demonstrated that a combination of the [(35)S]GSH trapping and TDI assays is an effective method for detecting compounds potentially capable of generating highly reactive metabolites in the early stages of drug discovery. DOI : 10.1124/dmd.111.039180
FLUTAMIDE
Time dependent inhibition of CYP3A4 (unknown origin) at 100 uM by LC/MS system
—
Inhibition < 10.0 %
Reference
Title : Combination of GSH trapping and time-dependent inhibition assays as a predictive method of drugs generating highly reactive metabolites. Authors : Nakayama S, Takakusa H, Watanabe A, Miyaji Y, Suzuki W, Sugiyama D, Shiosakai K, Honda K, Okudaira N, Izumi T, Okazaki O. Publication Year :
2011 Journal : Drug Metab. Dispos. Abstract : Covalent binding (CB) of reactive metabolites (RMs) is potentially involved in severe adverse drug reactions. Because the CB assay is of low throughput and costly, a qualitative trapping assay using agents such as [(35)S]GSH is often performed in the early stages of drug discovery. However, trapping methods alone cannot replace the CB assay. We hypothesized that the time-dependent inhibition (TDI) assay might be complementary to the [(35)S]GSH trapping assay in detecting RMs. We performed CB assays, [(35)S]GSH trapping assays, and TDI assays for 42 structurally diverse compounds. First, we showed that the [(35)S]GSH trapping assay alone does not correlate with the extent of CB. Four compounds that the [(35)S]GSH trapping assay failed to detect but that showed high extent of CB were inactivators of the enzyme in the TDI assay. There was a tendency for compounds judged as positive in the TDI assay to show a high degree of CB irrespective of the result of the [(35)S]GSH trapping assay. Finally, to combine parameters from the two assays, we introduced intrinsic clearance to describe the formation of RMs (CL(int, RMs)). The Spearman rank correlation coefficient between the extent of CB and CL(int, RMs) was 0.77 (p < 0.0001), which was better than that for the formation rates of [(35)S]GSH adducts. Therefore, we demonstrated that a combination of the [(35)S]GSH trapping and TDI assays is an effective method for detecting compounds potentially capable of generating highly reactive metabolites in the early stages of drug discovery. DOI : 10.1124/dmd.111.039180
FUROSEMIDE
Time dependent inhibition of CYP3A4 (unknown origin) at 100 uM by LC/MS system
—
Inhibition < 10.0 %
Reference
Title : Combination of GSH trapping and time-dependent inhibition assays as a predictive method of drugs generating highly reactive metabolites. Authors : Nakayama S, Takakusa H, Watanabe A, Miyaji Y, Suzuki W, Sugiyama D, Shiosakai K, Honda K, Okudaira N, Izumi T, Okazaki O. Publication Year :
2011 Journal : Drug Metab. Dispos. Abstract : Covalent binding (CB) of reactive metabolites (RMs) is potentially involved in severe adverse drug reactions. Because the CB assay is of low throughput and costly, a qualitative trapping assay using agents such as [(35)S]GSH is often performed in the early stages of drug discovery. However, trapping methods alone cannot replace the CB assay. We hypothesized that the time-dependent inhibition (TDI) assay might be complementary to the [(35)S]GSH trapping assay in detecting RMs. We performed CB assays, [(35)S]GSH trapping assays, and TDI assays for 42 structurally diverse compounds. First, we showed that the [(35)S]GSH trapping assay alone does not correlate with the extent of CB. Four compounds that the [(35)S]GSH trapping assay failed to detect but that showed high extent of CB were inactivators of the enzyme in the TDI assay. There was a tendency for compounds judged as positive in the TDI assay to show a high degree of CB irrespective of the result of the [(35)S]GSH trapping assay. Finally, to combine parameters from the two assays, we introduced intrinsic clearance to describe the formation of RMs (CL(int, RMs)). The Spearman rank correlation coefficient between the extent of CB and CL(int, RMs) was 0.77 (p < 0.0001), which was better than that for the formation rates of [(35)S]GSH adducts. Therefore, we demonstrated that a combination of the [(35)S]GSH trapping and TDI assays is an effective method for detecting compounds potentially capable of generating highly reactive metabolites in the early stages of drug discovery. DOI : 10.1124/dmd.111.039180
IMATINIB MESYLATE
Activity of CYP3A4 in human liver microsomes assessed as inhibition of ketoconazole-mediated N-desmethylimatinib formation at 0.1 uM after 5 mins by LC/MS analysis
Liver
Inhibition = 80.0 %
Reference
Title : Autoinhibition of CYP3A4 leads to important role of CYP2C8 in imatinib metabolism: variability in CYP2C8 activity may alter plasma concentrations and response. Authors : Filppula AM, Neuvonen M, Laitila J, Neuvonen PJ, Backman JT. Publication Year :
2013 Journal : Drug Metab. Dispos. Abstract : Recent data suggest that the role of CYP3A4 in imatinib metabolism is smaller than presumed. This study aimed to evaluate the quantitative importance of different cytochrome P450 (P450) enzymes in imatinib pharmacokinetics. First, the metabolism of imatinib was investigated using recombinant P450 enzymes and human liver microsomes with P450 isoform-selective inhibitors. Thereafter, an in silico model for imatinib was constructed to perform pharmacokinetic simulations to assess the roles of P450 enzymes in imatinib elimination at clinically used imatinib doses. In vitro, CYP2C8 inhibitors and CYP3A4 inhibitors inhibited the depletion of 0.1 µM imatinib by 45 and 80%, respectively, and the formation of the main metabolite of imatinib, N-desmethylimatinib, by >50%. Likewise, recombinant CYP2C8 and CYP3A4 metabolized imatinib extensively, whereas other isoforms had minor effect on imatinib concentrations. In the beginning of imatinib treatment, the fractions of its hepatic clearance mediated by CYP2C8 and CYP3A4 were predicted to approximate 40 and 60%, respectively. During long-term treatment with imatinib 400 mg once or twice daily, up to 65 or 75% of its hepatic elimination was predicted to occur via CYP2C8, and only about 35 or 25% by CYP3A4, due to dose- and time-dependent autoinactivation of CYP3A4 by imatinib. Thus, although CYP2C8 and CYP3A4 are the main enzymes in imatinib metabolism in vitro, in silico predictions indicate that imatinib inhibits its own CYP3A4-mediated metabolism, assigning a key role for CYP2C8. During multiple dosing, pharmacogenetic polymorphisms and drug interactions affecting CYP2C8 activity may cause marked interindividual variation in the exposure and response to imatinib. DOI : 10.1124/dmd.112.048017
IMATINIB MESYLATE
Activity of CYP3A4 in human liver microsomes assessed as inhibition of troleandomycin-mediated N-desmethylimatinib formation at 0.1 uM after 5 mins by LC/MS analysis
Liver
Inhibition = 80.0 %
Reference
Title : Autoinhibition of CYP3A4 leads to important role of CYP2C8 in imatinib metabolism: variability in CYP2C8 activity may alter plasma concentrations and response. Authors : Filppula AM, Neuvonen M, Laitila J, Neuvonen PJ, Backman JT. Publication Year :
2013 Journal : Drug Metab. Dispos. Abstract : Recent data suggest that the role of CYP3A4 in imatinib metabolism is smaller than presumed. This study aimed to evaluate the quantitative importance of different cytochrome P450 (P450) enzymes in imatinib pharmacokinetics. First, the metabolism of imatinib was investigated using recombinant P450 enzymes and human liver microsomes with P450 isoform-selective inhibitors. Thereafter, an in silico model for imatinib was constructed to perform pharmacokinetic simulations to assess the roles of P450 enzymes in imatinib elimination at clinically used imatinib doses. In vitro, CYP2C8 inhibitors and CYP3A4 inhibitors inhibited the depletion of 0.1 µM imatinib by 45 and 80%, respectively, and the formation of the main metabolite of imatinib, N-desmethylimatinib, by >50%. Likewise, recombinant CYP2C8 and CYP3A4 metabolized imatinib extensively, whereas other isoforms had minor effect on imatinib concentrations. In the beginning of imatinib treatment, the fractions of its hepatic clearance mediated by CYP2C8 and CYP3A4 were predicted to approximate 40 and 60%, respectively. During long-term treatment with imatinib 400 mg once or twice daily, up to 65 or 75% of its hepatic elimination was predicted to occur via CYP2C8, and only about 35 or 25% by CYP3A4, due to dose- and time-dependent autoinactivation of CYP3A4 by imatinib. Thus, although CYP2C8 and CYP3A4 are the main enzymes in imatinib metabolism in vitro, in silico predictions indicate that imatinib inhibits its own CYP3A4-mediated metabolism, assigning a key role for CYP2C8. During multiple dosing, pharmacogenetic polymorphisms and drug interactions affecting CYP2C8 activity may cause marked interindividual variation in the exposure and response to imatinib. DOI : 10.1124/dmd.112.048017
ITRACONAZOLE
Inhibition of human liver microsome CYP3A4 at 1 uM expressed in baculosomes using fluorogenic-DBOMF as substrate preincubated for 10 mins followed by substrate addition and measured after 60 mins by fluorescence based assay
Liver
Inhibition = 84.0 %
Reference
Title : Novel Tetrazole-Containing Analogues of Itraconazole as Potent Antiangiogenic Agents with Reduced Cytochrome P450 3A4 Inhibition. Authors : Li Y, Pasunooti KK, Li RJ, Liu W, Head SA, Shi WQ, Liu JO. Publication Year :
2018 Journal : J Med Chem Abstract : Itraconazole has been found to possess potent antiangiogenic activity, exhibiting promising antitumor activity in several human clinical studies. The wider use of itraconazole in the treatment of cancer, however, has been limited by its potent inhibition of the drug metabolizing enzyme cytochrome P450 3A4 (CYP3A4). In an effort to eliminate the CYP3A4 inhibition while retaining its antiangiogenic activity, we designed and synthesized a series of derivatives in which the 1,2,4-triazole ring is replaced with various azoles and nonazoles. Among these analogues, 15n with tetrazole in place of 1,2,4-triazole exhibited optimal inhibition of human umbilical vein endothelial cell proliferation with an IC50 of 73 nM without a significant effect on CYP3A4 (EC50 > 20 μM). Similar to itraconazole, 15n induced Niemann-Pick C phenotype (NPC phenotype) and blocked AMPK/mechanistic target of rapamycin signaling. These results suggest that 15n is a promising angiogenesis inhibitor that can be used in combination with most other known anticancer drugs. DOI : 10.1021/acs.jmedchem.8b01252
ITRACONAZOLE
Inhibition of CYP3A4 (unknown origin) using midazolam as substrate by UPLC-MS/MS analysis
—
Inhibition = 0.04 %
Reference
Title : Structure-Activity Relationships for Itraconazole-Based Triazolone Analogues as Hedgehog Pathway Inhibitors. Authors : Pace JR, Teske KA, Chau LQ, Dash RC, Zaino AM, Wechsler-Reya RJ, Hadden MK. Publication Year :
2019 Journal : J Med Chem Abstract : The Food and Drug Administration-approved antifungal agent, itraconazole (ITZ), has been increasingly studied for its novel biological properties. In particular, ITZ inhibits the hedgehog (Hh) signaling pathway and has the potential to serve as an anticancer chemotherapeutic against several Hh-dependent malignancies. We have extended our studies on ITZ analogues as Hh pathway inhibitors through the design, synthesis, and evaluation of novel des-triazole ITZ analogues that incorporate modifications to the triazolone/side chain region of the scaffold. Our overall results suggest that the triazolone/side chain region can be replaced with various functionalities (hydrazine carboxamides and meta-substituted amides) resulting in improved potency when compared to ITZ. Our studies also indicate that the stereochemical orientation of the dioxolane ring is important for both potent Hh pathway inhibition and compound stability. Finally, our studies suggest that the ITZ scaffold can be successfully modified in terms of functionality and stereochemistry to further improve its anti-Hh potency and physicochemical properties. DOI : 10.1021/acs.jmedchem.8b01283
ITRACONAZOLE
Inhibition of CYP3A4 in human hepatocytes using testosterone as substrate by HPLC/MS/MS method
—
IC50 = 70.0 nM
Reference
Title : Design and optimization of highly-selective fungal CYP51 inhibitors. Authors : Hoekstra WJ, Garvey EP, Moore WR, Rafferty SW, Yates CM, Schotzinger RJ. Publication Year :
2014 Journal : Bioorg. Med. Chem. Lett. Abstract : While the orally-active azoles such as voriconazole and itraconazole are effective antifungal agents, they potently inhibit a broad range of off-target human cytochrome P450 enzymes (CYPs) leading to various safety issues (e.g., drug-drug interactions, liver toxicity). Herein, we describe rationally-designed, broad-spectrum antifungal agents that are more selective for the target fungal enzyme, CYP51, than related human CYP enzymes such as CYP3A4. Using proprietary methodology, the triazole metal-binding group found in current clinical agents was replaced with novel, less avid metal-binding groups in concert with potency-enhancing molecular scaffold modifications. This process produced a unique series of fungal CYP51-selective inhibitors that included the oral antifungal 7d (VT-1161), now in Phase 2 clinical trials. This series exhibits excellent potency against key yeast and dermatophyte strains. The chemical methodology described is potentially applicable to the design of new and more effective metalloenzyme inhibitor treatments for a broad array of diseases. DOI : 10.1016/j.bmcl.2014.05.068
ITRACONAZOLE
Inhibition of CYP3A4 in human liver microsomes using testosterone as substrate after 10 mins by LC/MS/MS analysis
Liver
IC50 = 50.4 nM
Reference
Title : Repurposing the Clinically Efficacious Antifungal Agent Itraconazole as an Anticancer Chemotherapeutic. Authors : Pace JR, DeBerardinis AM, Sail V, Tacheva-Grigorova SK, Chan KA, Tran R, Raccuia DS, Wechsler-Reya RJ, Hadden MK. Publication Year :
2016 Journal : J. Med. Chem. Abstract : Itraconazole (ITZ) is an FDA-approved member of the triazole class of antifungal agents. Two recent drug repurposing screens identified ITZ as a promising anticancer chemotherapeutic that inhibits both the angiogenesis and hedgehog (Hh) signaling pathways. We have synthesized and evaluated first- and second-generation ITZ analogues for their anti-Hh and antiangiogenic activities to probe more fully the structural requirements for these anticancer properties. Our overall results suggest that the triazole functionality is required for ITZ-mediated inhibition of angiogenesis but that it is not essential for inhibition of Hh signaling. The synthesis and evaluation of stereochemically defined des-triazole ITZ analogues also provides key information as to the optimal configuration around the dioxolane ring of the ITZ scaffold. Finally, the results from our studies suggest that two distinct cellular mechanisms of action govern the anticancer properties of the ITZ scaffold. DOI : 10.1021/acs.jmedchem.5b01718
ITRACONAZOLE
Inhibition of CYP3A4 in human liver microsomes assessed as reduction in nifedipine oxidation incubated for 10 mins
Liver
IC50 = 32.6 nM
Reference
Title : Discovery of biphenyl imidazole derivatives as potent antifungal agents: Design, synthesis, and structure-activity relationship studies. Authors : Zhao D, Zhao S, Zhao L, Zhang X, Wei P, Liu C, Hao C, Sun B, Su X, Cheng M. Publication Year :
2017 Journal : Bioorg Med Chem Abstract : Fungal infections have became a serious medical problem due to their high incidence and mortality. We describe the discovery and structure-activity relationships studies (SARs) of a series of novel biphenyl imidazole derivatives with excellent antifungal activities against Candida albicans and Cryptococcus neoformans. The most promising compounds 12f-g and 19a-b exhibited excellent activity with minimum inhibitory concentration (MIC) values in the range of 0.03125-2μg/mL. Preliminary mechanism studies showed that the potent antifungal activity of compound 12g stemed from inhibition of CYP51 in Candida albicans. Furthermore, compounds 12g and 19b exhibited low inhibition profiles for various human cytochrome P450 isoforms. The SARs and binding mode established in this study will be useful for further lead optimization. DOI : 10.1016/j.bmc.2016.11.051
ITRACONAZOLE
Inhibition of CYP3A4 in human liver microsomes by LC-MS/MS analysis
Liver
IC50 = 40.0 nM
Reference
Title : Development of posaconazole-based analogues as hedgehog signaling pathway inhibitors. Authors : Teske KA, Dash RC, Morel SR, Chau LQ, Wechsler-Reya RJ, Hadden MK. Publication Year :
2019 Journal : Eur J Med Chem Abstract : Inhibition of the hedgehog (Hh) signaling pathway has been validated as a therapeutic strategy to treat basal cell carcinoma and holds potential for several other forms of human cancer. Itraconazole and posaconazole are clinically useful triazole anti-fungals that are being repurposed as anti-cancer agents based on their ability to inhibit the Hh pathway. We have previously demonstrated that removal of the triazole from itraconazole does not affect its ability to inhibit the Hh pathway while abolishing its primary side effect, potent inhibition of Cyp3A4. To develop structure-activity relationships for the related posaconazole scaffold, we synthesized and evaluated a series of des-triazole analogues designed through both ligand- and structure-based methods. These compounds demonstrated improved anti-Hh properties compared to posaconazole and enhanced stability without inhibiting Cyp3A4. In addition, we utilized a series of molecular dynamics and binding energy studies to probe specific interactions between the compounds and their proposed binding site on Smoothened. These studies strongly suggest that the tetrahydrofuran region of the scaffold projects out of the binding site and that π-π interactions between the compound and Smoothened play a key role in stabilizing the bound analogues. DOI : 10.1016/j.ejmech.2018.11.056
ITRACONAZOLE
Inhibition of CYP3A4 in human liver microsome using Nifedipine as a substrate incubated for 10 mins in the presence of NADPH by LC-MS/MS analysis
Liver
IC50 = 32.6 nM
Reference
Title : Authors : Publication Year : Journal :
KETOCONAZOLE
Inhibition of CYP3A4 in human liver microsome
Liver
Inhibition = 103.0 %
Reference
Title : Synthesis of novel ketoconazole derivatives as inhibitors of the human Pregnane X Receptor (PXR; NR1I2; also termed SXR, PAR). Authors : Das BC, Madhukumar AV, Anguiano J, Kim S, Sinz M, Zvyaga TA, Power EC, Ganellin CR, Mani S. Publication Year :
2008 Journal : Bioorg. Med. Chem. Lett. Abstract : PXR, pregnane X receptor, in its activated state, is a validated target for controlling certain drug-drug interactions in humans. In this context, there is a paucity of inhibitors directed toward activated PXR. Using prior observations with ketoconazole as a PXR inhibitor, the target compound 3 was synthesized from (s)-glycidol with overall 56% yield. (+)-Glycidol was reacted with 4-bromophenol and potassium carbonate in DMF to yield the ring opened compound 6. This was then heated to reflux in benzene along with 2', 4'-difluoroacetophenone and catalytic amount of para-toluene sulfonic acid to yield 8. The resultant acetal 8 was then functionalized using Palladium chemistry to yield the target compound 3. The activity of the compound was compared with ketoconazole and UCL2158H. However, in contrast with ketoconazole (IC(50) approximately 0.020 microM; approximately 100% inhibition), 3 has negligible effects on inhibition of microsomal CYP450 (maximum approximately 20% inhibition) at concentrations >40 microM. In vitro, micromolar concentration of ketoconazole is toxic to passaged human cell lines, while 3 does not exhibit cytotoxicity up to concentrations approximately 100 microM (viability >85%). This is the first demonstration of a chemical analog of a PXR inhibitor that retains activity against activated PXR. Furthermore, in contrast with ketoconazole, 3 is less toxic in human cell lines and has negligible CYP450 activity. DOI : 10.1016/j.bmcl.2008.06.018
KETOCONAZOLE
Inhibition of human CYP3A4 at 10 uM
—
Inhibition > 93.0 %
Reference
Title : Synthesis and evaluation of non-dimeric HCV NS5A inhibitors. Authors : Amblard F, Zhang H, Zhou L, Shi J, Bobeck DR, Nettles JH, Chavre S, McBrayer TR, Tharnish P, Whitaker T, Coats SJ, Schinazi RF. Publication Year :
2013 Journal : Bioorg. Med. Chem. Lett. Abstract : Based on the symmetrical bidentate structure of the NS5A inhibitor BMS-790052, a series of new monodentate molecules were designed. The synthesis of 36 new non-dimeric NS5A inhibitors is reported along with their ability to block HCV replication in an HCV 1b replicon system. Among them compound 5a showed picomolar range activity along with an excellent selectivity index (SI > 90,000). DOI : 10.1016/j.bmcl.2013.02.023
KETOCONAZOLE
Inhibition of CYP3A4 in human liver microsomes assessed as midazolam hydroxylation to 1'-hydroxymidazolam at 1 uM after 10 mins relative to control
Liver
Inhibition = 95.0 %
Reference
Title : Rational development of 4-aminopyridyl-based inhibitors targeting Trypanosoma cruzi CYP51 as anti-chagas agents. Authors : Choi JY, Calvet CM, Gunatilleke SS, Ruiz C, Cameron MD, McKerrow JH, Podust LM, Roush WR. Publication Year :
2013 Journal : J. Med. Chem. Abstract : A new series of 4-aminopyridyl-based lead inhibitors targeting Trypanosoma cruzi CYP51 (TcCYP51) has been developed using structure-based drug design as well as structure-property relationship (SPR) analyses. The screening hit starting point, LP10 (KD ≤ 42 nM; EC50 = 0.65 μM), has been optimized to give the potential leads 14t, 27i, 27q, 27r, and 27t, which have low-nanomolar binding affinity to TcCYP51 and significant activity against T. cruzi amastigotes cultured in human myoblasts (EC50 = 14-18 nM for 27i and 27r). Many of the optimized compounds have improved microsome stability, and most are selective against human CYPs 1A2, 2D6, and 3A4 (<50% inhibition at 1 μM). A rationale for the improvement in microsome stability and selectivity of inhibitors against human metabolic CYP enzymes is presented. In addition, the binding mode of 14t with the Trypanosoma brucei CYP51 (TbCYP51) orthologue has been characterized by X-ray structure analysis. DOI : 10.1021/jm401067s
KETOCONAZOLE
Inhibition of CYP3A4 (unknown origin) at 10 uM relative to control
—
Inhibition = 95.34 %
Reference
Title : Potent Hepatitis C Virus NS5A Inhibitors Containing a Benzidine Core. Authors : Bae IH, Choi JK, Chough C, Keum SJ, Kim H, Jang SK, Kim BM. Publication Year :
2014 Journal : ACS Med. Chem. Lett. Abstract : Here we report the discovery of a series of potent hepatitis C virus (HCV) NS5A inhibitors based on the benzidine prolinamide backbone. Taking a simple synthetic route, we developed a novel inhibitor structure, which allows easy modification, and through optimization of the capping group, we identified compound 6 with highly potent anti-HCV activity. Compound 6 is nontoxic and is anticipated to be an effective HCV drug candidate. DOI : 10.1021/ml4003293
KETOCONAZOLE
Inhibition of human CYP3A4 at 10 uM
—
Inhibition = 99.0 %
Reference
Title : Development of (E)-2-((1,4-dimethylpiperazin-2-ylidene)amino)-5-nitro-N-phenylbenzamide, ML336: Novel 2-amidinophenylbenzamides as potent inhibitors of venezuelan equine encephalitis virus. Authors : Schroeder CE, Yao T, Sotsky J, Smith RA, Roy S, Chu YK, Guo H, Tower NA, Noah JW, McKellip S, Sosa M, Rasmussen L, Smith LH, White EL, Aubé J, Jonsson CB, Chung D, Golden JE. Publication Year :
2014 Journal : J. Med. Chem. Abstract : Venezuelan equine encephalitis virus (VEEV) is an emerging pathogenic alphavirus that can cause significant disease in humans. Given the absence of therapeutic options available and the significance of VEEV as a weaponized agent, an optimization effort was initiated around a quinazolinone screening hit 1 with promising cellular antiviral activity (EC50 = 0.8 μM), limited cytotoxic liability (CC50 > 50 μM), and modest in vitro efficacy in reducing viral progeny (63-fold at 5 μM). Scaffold optimization revealed a novel rearrangement affording amidines, specifically compound 45, which was found to potently inhibit several VEEV strains in the low nanomolar range without cytotoxicity (EC50 = 0.02-0.04 μM, CC50 > 50 μM) while limiting in vitro viral replication (EC90 = 0.17 μM). Brain exposure was observed in mice with 45. Significant protection was observed in VEEV-infected mice at 5 mg kg(-1) day(-1) and viral replication appeared to be inhibited through interference of viral nonstructural proteins. DOI : 10.1021/jm501203v
KETOCONAZOLE
Inhibition of human recombinant CYP3A4 expressed in insect cell microsomes preincubated for 10 mins at 0.1 uM by fluorescence assay
—
Inhibition = 33.9 %
Reference
Title : Investigating the binding interactions of the anti-Alzheimer's drug donepezil with CYP3A4 and P-glycoprotein. Authors : McEneny-King A, Edginton AN, Rao PP. Publication Year :
2015 Journal : Bioorg. Med. Chem. Lett. Abstract : The anti-Alzheimer's agent donepezil is known to bind to the hepatic enzyme CYP3A4, but its relationship with the efflux transporter P-glycoprotein (P-gp) is not as well elucidated. We conducted in vitro inhibition studies of donepezil using human recombinant CYP3A4 and P-gp. These studies show that donepezil is a weak inhibitor of CYP3A4 (IC50=54.68±1.00μM) whereas the reference agent ketoconazole exhibited potent inhibition (CYP3A4 IC50=0.20±0.01μM). P-gp inhibition studies indicate that donepezil exhibits better inhibition relative to CYP3A4 (P-gp EC50=34.85±4.63μM) although it was less potent compared to ketoconazole (P-gp EC50=9.74±1.23μM). At higher concentrations, donepezil exhibited significant inhibition of CYP3A4 (69%, 84% and 87% inhibition at 100, 250 and 500μM, respectively). This indicates its potential to cause drug-drug interactions with other CYP3A4 substrates upon co-administration; however, this scenario is unlikely in vivo due to the low therapeutic concentrations of donepezil. Similarly, donepezil co-administration with P-gp substrates or inhibitors is unlikely to result in beneficial or adverse drug interactions. The molecular docking studies show that the 5,6-dimethoxyindan-1-one moiety of donepezil was oriented closer to the heme center in CYP3A4 whereas in the P-gp binding site, the protonated benzylpiperidine pharmacophore of donepezil played a major role in its binding ability. Energy parameters indicate that donepezil complex with both CYP3A4 and P-gp was less stable (CDOCKER energies=-15.05 and -4.91kcal/mol, respectively) compared to the ketoconazole-CYP3A4 and P-gp complex (CDOCKER energies=-41.89 and -20.03kcal/mol, respectively). DOI : 10.1016/j.bmcl.2014.11.046
KETOCONAZOLE
Inhibition of human recombinant CYP3A4 expressed in insect cell microsomes preincubated for 10 mins at 1 uM by fluorescence assay
—
Inhibition = 87.2 %
Reference
Title : Investigating the binding interactions of the anti-Alzheimer's drug donepezil with CYP3A4 and P-glycoprotein. Authors : McEneny-King A, Edginton AN, Rao PP. Publication Year :
2015 Journal : Bioorg. Med. Chem. Lett. Abstract : The anti-Alzheimer's agent donepezil is known to bind to the hepatic enzyme CYP3A4, but its relationship with the efflux transporter P-glycoprotein (P-gp) is not as well elucidated. We conducted in vitro inhibition studies of donepezil using human recombinant CYP3A4 and P-gp. These studies show that donepezil is a weak inhibitor of CYP3A4 (IC50=54.68±1.00μM) whereas the reference agent ketoconazole exhibited potent inhibition (CYP3A4 IC50=0.20±0.01μM). P-gp inhibition studies indicate that donepezil exhibits better inhibition relative to CYP3A4 (P-gp EC50=34.85±4.63μM) although it was less potent compared to ketoconazole (P-gp EC50=9.74±1.23μM). At higher concentrations, donepezil exhibited significant inhibition of CYP3A4 (69%, 84% and 87% inhibition at 100, 250 and 500μM, respectively). This indicates its potential to cause drug-drug interactions with other CYP3A4 substrates upon co-administration; however, this scenario is unlikely in vivo due to the low therapeutic concentrations of donepezil. Similarly, donepezil co-administration with P-gp substrates or inhibitors is unlikely to result in beneficial or adverse drug interactions. The molecular docking studies show that the 5,6-dimethoxyindan-1-one moiety of donepezil was oriented closer to the heme center in CYP3A4 whereas in the P-gp binding site, the protonated benzylpiperidine pharmacophore of donepezil played a major role in its binding ability. Energy parameters indicate that donepezil complex with both CYP3A4 and P-gp was less stable (CDOCKER energies=-15.05 and -4.91kcal/mol, respectively) compared to the ketoconazole-CYP3A4 and P-gp complex (CDOCKER energies=-41.89 and -20.03kcal/mol, respectively). DOI : 10.1016/j.bmcl.2014.11.046
KETOCONAZOLE
Inhibition of human recombinant CYP3A4 expressed in insect cell microsomes preincubated for 10 mins at 10 uM by fluorescence assay
—
Inhibition = 92.3 %
Reference
Title : Investigating the binding interactions of the anti-Alzheimer's drug donepezil with CYP3A4 and P-glycoprotein. Authors : McEneny-King A, Edginton AN, Rao PP. Publication Year :
2015 Journal : Bioorg. Med. Chem. Lett. Abstract : The anti-Alzheimer's agent donepezil is known to bind to the hepatic enzyme CYP3A4, but its relationship with the efflux transporter P-glycoprotein (P-gp) is not as well elucidated. We conducted in vitro inhibition studies of donepezil using human recombinant CYP3A4 and P-gp. These studies show that donepezil is a weak inhibitor of CYP3A4 (IC50=54.68±1.00μM) whereas the reference agent ketoconazole exhibited potent inhibition (CYP3A4 IC50=0.20±0.01μM). P-gp inhibition studies indicate that donepezil exhibits better inhibition relative to CYP3A4 (P-gp EC50=34.85±4.63μM) although it was less potent compared to ketoconazole (P-gp EC50=9.74±1.23μM). At higher concentrations, donepezil exhibited significant inhibition of CYP3A4 (69%, 84% and 87% inhibition at 100, 250 and 500μM, respectively). This indicates its potential to cause drug-drug interactions with other CYP3A4 substrates upon co-administration; however, this scenario is unlikely in vivo due to the low therapeutic concentrations of donepezil. Similarly, donepezil co-administration with P-gp substrates or inhibitors is unlikely to result in beneficial or adverse drug interactions. The molecular docking studies show that the 5,6-dimethoxyindan-1-one moiety of donepezil was oriented closer to the heme center in CYP3A4 whereas in the P-gp binding site, the protonated benzylpiperidine pharmacophore of donepezil played a major role in its binding ability. Energy parameters indicate that donepezil complex with both CYP3A4 and P-gp was less stable (CDOCKER energies=-15.05 and -4.91kcal/mol, respectively) compared to the ketoconazole-CYP3A4 and P-gp complex (CDOCKER energies=-41.89 and -20.03kcal/mol, respectively). DOI : 10.1016/j.bmcl.2014.11.046
KETOCONAZOLE
Inhibition of recombinant human CYP3A4 preincubated at 10 uM for 5 mins before fluorescent substrate addition by fluorescence assay
—
Inhibition = 14.4 %
Reference
Title : Synthesis and biological evaluation of 3-phenethylazetidine derivatives as triple reuptake inhibitors. Authors : Yun J, Han M, Song C, Cheon SH, Choi K, Hahn HG. Publication Year :
2014 Journal : Bioorg. Med. Chem. Lett. Abstract : We report the synthesis of 3-phenethylazetidine derivatives 2 and their biological activities against 5-HT, NE and DA transporters as well as microsomal stability, CYP inhibition, and hERG inhibition profiles. Compound 2at showed most potent triple reuptake inhibitor with good selectivity as a candidate for depression. DOI : 10.1016/j.bmcl.2014.06.026
KETOCONAZOLE
Inhibition of human CYP3A4 at 10 uM preincubated for 10 mins with cofactor followed by mixture of enzyme-substrate addition by fluorescence assay relative to control
—
Inhibition = 95.2 %
Reference
Title : Osthol and curcumin as inhibitors of human Pgp and multidrug efflux pumps of Staphylococcus aureus: reversing the resistance against frontline antibacterial drugs Authors : Joshi P, Singh S, Wani A, Sharma S, Jain SK, Singh B, Gupta BD, Satti NK, Koul S, Khan IA, Kumar A, Bharate SB, Vishwakarma RA Publication Year :
2014 Journal : MedChemComm DOI : 10.1039/C4MD00196F
KETOCONAZOLE
Inhibition of CYP3A4 (unknown origin) at 20 uM by luminescent readout-based method
—
Inhibition = 95.37 %
Reference
Title : Phenoxymethyl 1,3-oxazoles and 1,2,4-oxadiazoles as potent and selective agonists of free fatty acid receptor 1 (GPR40). Authors : Zahanich I, Kondratov I, Naumchyk V, Kheylik Y, Platonov M, Zozulya S, Krasavin M. Publication Year :
2015 Journal : Bioorg. Med. Chem. Lett. Abstract : A screening hit that showed a weak (EC50 = 18 μM), partial agonistic effect on GPR40 was used a prototype for expedited hit expansion effort using a set of advanced building blocks. The latter yielded several 1,3-oxazoles and 1,2,4-oxadiazoles with significantly improved potency (best EC50 = 0.058 μM). The lead compounds in each chemotype showed a very good ADME profile (aqueous solubility, plasma protein binding, microsomal stability and membrane permeability) and no appreciable inhibition of key cytochromes P450. The compounds reported are significant new starting points for further preclinical development of future diabetic agents with a mechanism of action for which a first-in-class agent is yet to be approved. DOI : 10.1016/j.bmcl.2015.06.018
KETOCONAZOLE
Inhibition of CYP3A4 (unknown origin) at 20 uM by P450-glo assay
—
Inhibition = 95.37 %
Reference
Title : Novel free fatty acid receptor 1 (GPR40) agonists based on 1,3,4-thiadiazole-2-carboxamide scaffold. Authors : Krasavin M, Lukin A, Zhurilo N, Kovalenko A, Zahanich I, Zozulya S, Moore D, Tikhonova IG. Publication Year :
2016 Journal : Bioorg. Med. Chem. Abstract : Free fatty acid receptor 1 (FFA1), previously known as GPR40 is a G protein-coupled receptor and a new target for treatment of type 2 diabetes. Two series of FFA1 agonists utilizing a 1,3,4-thiadiazole-2-caboxamide scaffold were synthetized. Both series offered significant improvement of the potency compared to the previously described 1,3,4-thiadiazole-based FFA1 agonists and high selectivity for FFA1. Molecular docking predicts new aromatic interactions with the receptor that improve agonist potency. The most potent compounds from both series were profiled for in vitro ADME properties (plasma and metabolic stability, LogD, plasma protein binding, hERG binding and CYP inhibition). One series suffered very rapid degradation in plasma and in presence of mouse liver microsomes. However, the other series delivered a lead compound that displayed a reasonable ADME profile together with the improved FFA1 potency. DOI : 10.1016/j.bmc.2016.04.065
KETOCONAZOLE
Inhibition of CYP3A4 in human liver microsomes using midazolam as substrate at 10 uM preincubated for 5 mins followed by NADPH addition measured after 5 mins by LC-MS/MS analysis relative to control
Liver
Inhibition = 100.0 %
Reference
Title : Potent and Selective Human Neuronal Nitric Oxide Synthase Inhibition by Optimization of the 2-Aminopyridine-Based Scaffold with a Pyridine Linker. Authors : Wang HY, Qin Y, Li H, Roman LJ, Martásek P, Poulos TL, Silverman RB. Publication Year :
2016 Journal : J. Med. Chem. Abstract : Neuronal nitric oxide synthase (nNOS) is an important therapeutic target for the treatment of various neurodegenerative disorders. A major challenge in the design of nNOS inhibitors focuses on potency in humans and selectivity over other NOS isoforms. Here we report potent and selective human nNOS inhibitors based on the 2-aminopyridine scaffold with a central pyridine linker. Compound 14j, the most promising inhibitor in this study, exhibits excellent potency for rat nNOS (Ki = 16 nM) with 828-fold n/e and 118-fold n/i selectivity with a Ki value of 13 nM against human nNOS with 1761-fold human n/e selectivity. Compound 14j also displayed good metabolic stability in human liver microsomes, low plasma protein binding, and minimal binding to cytochromes P450 (CYPs), although it had little to no Caco-2 permeability. DOI : 10.1021/acs.jmedchem.6b00273
KETOCONAZOLE
Inhibition of CYP3A4 in human liver microsomes using midazolam as substrate assessed as reduction in 1-OH-midazolam formation at 0.005 uM by LC/MS/MS analysis
Liver
Inhibition = 34.0 %
Reference
Title : Identification of (R)-(2-Chloro-3-(trifluoromethyl)phenyl)(1-(5-fluoropyridin-2-yl)-4-methyl-6,7-dihydro-1H-imidazo[4,5-c]pyridin-5(4H)-yl)methanone (JNJ 54166060), a Small Molecule Antagonist of the P2X7 receptor. Authors : Swanson DM, Savall BM, Coe KJ, Schoetens F, Koudriakova T, Skaptason J, Wall J, Rech J, Deng X, De Angelis M, Everson A, Lord B, Wang Q, Ao H, Scott B, Sepassi K, Lovenberg TW, Carruthers NI, Bhattacharya A, Letavic MA. Publication Year :
2016 Journal : J Med Chem Abstract : The synthesis and SAR of a series of 4,5,6,7-tetrahydro-imidazo[4,5-c]pyridine P2X7 antagonists are described. Addressing P2X7 affinity and liver microsomal stability issues encountered with this template afforded methyl substituted 4,5,6,7-tetrahydro-imidazo[4,5-c]pyridines ultimately leading to the identification of 1 (JNJ 54166060). 1 is a potent P2X7 antagonist with an ED50 = 2.3 mg/kg in rats, high oral bioavailability and low-moderate clearance in preclinical species, acceptable safety margins in rats, and a predicted human dose of 120 mg of QD. Additionally, 1 possesses a unique CYP profile and was found to be a regioselective inhibitor of midazolam CYP3A metabolism. DOI : 10.1021/acs.jmedchem.6b00989
KETOCONAZOLE
Inhibition of CYP3A4 in human liver microsomes using midazolam as substrate assessed as reduction in 1-OH-midazolam formation at 0.01 uM by LC/MS/MS analysis
Liver
Inhibition = 53.0 %
Reference
Title : Identification of (R)-(2-Chloro-3-(trifluoromethyl)phenyl)(1-(5-fluoropyridin-2-yl)-4-methyl-6,7-dihydro-1H-imidazo[4,5-c]pyridin-5(4H)-yl)methanone (JNJ 54166060), a Small Molecule Antagonist of the P2X7 receptor. Authors : Swanson DM, Savall BM, Coe KJ, Schoetens F, Koudriakova T, Skaptason J, Wall J, Rech J, Deng X, De Angelis M, Everson A, Lord B, Wang Q, Ao H, Scott B, Sepassi K, Lovenberg TW, Carruthers NI, Bhattacharya A, Letavic MA. Publication Year :
2016 Journal : J Med Chem Abstract : The synthesis and SAR of a series of 4,5,6,7-tetrahydro-imidazo[4,5-c]pyridine P2X7 antagonists are described. Addressing P2X7 affinity and liver microsomal stability issues encountered with this template afforded methyl substituted 4,5,6,7-tetrahydro-imidazo[4,5-c]pyridines ultimately leading to the identification of 1 (JNJ 54166060). 1 is a potent P2X7 antagonist with an ED50 = 2.3 mg/kg in rats, high oral bioavailability and low-moderate clearance in preclinical species, acceptable safety margins in rats, and a predicted human dose of 120 mg of QD. Additionally, 1 possesses a unique CYP profile and was found to be a regioselective inhibitor of midazolam CYP3A metabolism. DOI : 10.1021/acs.jmedchem.6b00989
KETOCONAZOLE
Inhibition of CYP3A4 in human liver microsomes using midazolam as substrate assessed as reduction in 1-OH-midazolam formation at 0.02 uM by LC/MS/MS analysis
Liver
Inhibition = 70.0 %
Reference
Title : Identification of (R)-(2-Chloro-3-(trifluoromethyl)phenyl)(1-(5-fluoropyridin-2-yl)-4-methyl-6,7-dihydro-1H-imidazo[4,5-c]pyridin-5(4H)-yl)methanone (JNJ 54166060), a Small Molecule Antagonist of the P2X7 receptor. Authors : Swanson DM, Savall BM, Coe KJ, Schoetens F, Koudriakova T, Skaptason J, Wall J, Rech J, Deng X, De Angelis M, Everson A, Lord B, Wang Q, Ao H, Scott B, Sepassi K, Lovenberg TW, Carruthers NI, Bhattacharya A, Letavic MA. Publication Year :
2016 Journal : J Med Chem Abstract : The synthesis and SAR of a series of 4,5,6,7-tetrahydro-imidazo[4,5-c]pyridine P2X7 antagonists are described. Addressing P2X7 affinity and liver microsomal stability issues encountered with this template afforded methyl substituted 4,5,6,7-tetrahydro-imidazo[4,5-c]pyridines ultimately leading to the identification of 1 (JNJ 54166060). 1 is a potent P2X7 antagonist with an ED50 = 2.3 mg/kg in rats, high oral bioavailability and low-moderate clearance in preclinical species, acceptable safety margins in rats, and a predicted human dose of 120 mg of QD. Additionally, 1 possesses a unique CYP profile and was found to be a regioselective inhibitor of midazolam CYP3A metabolism. DOI : 10.1021/acs.jmedchem.6b00989
KETOCONAZOLE
Inhibition of CYP3A4 in human liver microsomes using midazolam as substrate assessed as reduction in 1-OH-midazolam formation at 0.04 uM by LC/MS/MS analysis
Liver
Inhibition = 82.0 %
Reference
Title : Identification of (R)-(2-Chloro-3-(trifluoromethyl)phenyl)(1-(5-fluoropyridin-2-yl)-4-methyl-6,7-dihydro-1H-imidazo[4,5-c]pyridin-5(4H)-yl)methanone (JNJ 54166060), a Small Molecule Antagonist of the P2X7 receptor. Authors : Swanson DM, Savall BM, Coe KJ, Schoetens F, Koudriakova T, Skaptason J, Wall J, Rech J, Deng X, De Angelis M, Everson A, Lord B, Wang Q, Ao H, Scott B, Sepassi K, Lovenberg TW, Carruthers NI, Bhattacharya A, Letavic MA. Publication Year :
2016 Journal : J Med Chem Abstract : The synthesis and SAR of a series of 4,5,6,7-tetrahydro-imidazo[4,5-c]pyridine P2X7 antagonists are described. Addressing P2X7 affinity and liver microsomal stability issues encountered with this template afforded methyl substituted 4,5,6,7-tetrahydro-imidazo[4,5-c]pyridines ultimately leading to the identification of 1 (JNJ 54166060). 1 is a potent P2X7 antagonist with an ED50 = 2.3 mg/kg in rats, high oral bioavailability and low-moderate clearance in preclinical species, acceptable safety margins in rats, and a predicted human dose of 120 mg of QD. Additionally, 1 possesses a unique CYP profile and was found to be a regioselective inhibitor of midazolam CYP3A metabolism. DOI : 10.1021/acs.jmedchem.6b00989
KETOCONAZOLE
Inhibition of CYP3A4 in human liver microsomes using midazolam as substrate assessed as reduction in 1-OH-midazolam formation at 0.08 uM by LC/MS/MS analysis
Liver
Inhibition = 90.0 %
Reference
Title : Identification of (R)-(2-Chloro-3-(trifluoromethyl)phenyl)(1-(5-fluoropyridin-2-yl)-4-methyl-6,7-dihydro-1H-imidazo[4,5-c]pyridin-5(4H)-yl)methanone (JNJ 54166060), a Small Molecule Antagonist of the P2X7 receptor. Authors : Swanson DM, Savall BM, Coe KJ, Schoetens F, Koudriakova T, Skaptason J, Wall J, Rech J, Deng X, De Angelis M, Everson A, Lord B, Wang Q, Ao H, Scott B, Sepassi K, Lovenberg TW, Carruthers NI, Bhattacharya A, Letavic MA. Publication Year :
2016 Journal : J Med Chem Abstract : The synthesis and SAR of a series of 4,5,6,7-tetrahydro-imidazo[4,5-c]pyridine P2X7 antagonists are described. Addressing P2X7 affinity and liver microsomal stability issues encountered with this template afforded methyl substituted 4,5,6,7-tetrahydro-imidazo[4,5-c]pyridines ultimately leading to the identification of 1 (JNJ 54166060). 1 is a potent P2X7 antagonist with an ED50 = 2.3 mg/kg in rats, high oral bioavailability and low-moderate clearance in preclinical species, acceptable safety margins in rats, and a predicted human dose of 120 mg of QD. Additionally, 1 possesses a unique CYP profile and was found to be a regioselective inhibitor of midazolam CYP3A metabolism. DOI : 10.1021/acs.jmedchem.6b00989
KETOCONAZOLE
Inhibition of CYP3A4 in human liver microsomes using midazolam as substrate assessed as reduction in 1-OH-midazolam formation at 0.16 uM by LC/MS/MS analysis
Liver
Inhibition = 94.0 %
Reference
Title : Identification of (R)-(2-Chloro-3-(trifluoromethyl)phenyl)(1-(5-fluoropyridin-2-yl)-4-methyl-6,7-dihydro-1H-imidazo[4,5-c]pyridin-5(4H)-yl)methanone (JNJ 54166060), a Small Molecule Antagonist of the P2X7 receptor. Authors : Swanson DM, Savall BM, Coe KJ, Schoetens F, Koudriakova T, Skaptason J, Wall J, Rech J, Deng X, De Angelis M, Everson A, Lord B, Wang Q, Ao H, Scott B, Sepassi K, Lovenberg TW, Carruthers NI, Bhattacharya A, Letavic MA. Publication Year :
2016 Journal : J Med Chem Abstract : The synthesis and SAR of a series of 4,5,6,7-tetrahydro-imidazo[4,5-c]pyridine P2X7 antagonists are described. Addressing P2X7 affinity and liver microsomal stability issues encountered with this template afforded methyl substituted 4,5,6,7-tetrahydro-imidazo[4,5-c]pyridines ultimately leading to the identification of 1 (JNJ 54166060). 1 is a potent P2X7 antagonist with an ED50 = 2.3 mg/kg in rats, high oral bioavailability and low-moderate clearance in preclinical species, acceptable safety margins in rats, and a predicted human dose of 120 mg of QD. Additionally, 1 possesses a unique CYP profile and was found to be a regioselective inhibitor of midazolam CYP3A metabolism. DOI : 10.1021/acs.jmedchem.6b00989
KETOCONAZOLE
Inhibition of CYP3A4 in human liver microsomes using midazolam as substrate assessed as reduction in 4-OH-midazolam formation at 0.005 uM by LC/MS/MS analysis
Liver
Inhibition = 37.0 %
Reference
Title : Identification of (R)-(2-Chloro-3-(trifluoromethyl)phenyl)(1-(5-fluoropyridin-2-yl)-4-methyl-6,7-dihydro-1H-imidazo[4,5-c]pyridin-5(4H)-yl)methanone (JNJ 54166060), a Small Molecule Antagonist of the P2X7 receptor. Authors : Swanson DM, Savall BM, Coe KJ, Schoetens F, Koudriakova T, Skaptason J, Wall J, Rech J, Deng X, De Angelis M, Everson A, Lord B, Wang Q, Ao H, Scott B, Sepassi K, Lovenberg TW, Carruthers NI, Bhattacharya A, Letavic MA. Publication Year :
2016 Journal : J Med Chem Abstract : The synthesis and SAR of a series of 4,5,6,7-tetrahydro-imidazo[4,5-c]pyridine P2X7 antagonists are described. Addressing P2X7 affinity and liver microsomal stability issues encountered with this template afforded methyl substituted 4,5,6,7-tetrahydro-imidazo[4,5-c]pyridines ultimately leading to the identification of 1 (JNJ 54166060). 1 is a potent P2X7 antagonist with an ED50 = 2.3 mg/kg in rats, high oral bioavailability and low-moderate clearance in preclinical species, acceptable safety margins in rats, and a predicted human dose of 120 mg of QD. Additionally, 1 possesses a unique CYP profile and was found to be a regioselective inhibitor of midazolam CYP3A metabolism. DOI : 10.1021/acs.jmedchem.6b00989
KETOCONAZOLE
Inhibition of CYP3A4 in human liver microsomes using midazolam as substrate assessed as reduction in 4-OH-midazolam formation at 0.01 uM by LC/MS/MS analysis
Liver
Inhibition = 57.0 %
Reference
Title : Identification of (R)-(2-Chloro-3-(trifluoromethyl)phenyl)(1-(5-fluoropyridin-2-yl)-4-methyl-6,7-dihydro-1H-imidazo[4,5-c]pyridin-5(4H)-yl)methanone (JNJ 54166060), a Small Molecule Antagonist of the P2X7 receptor. Authors : Swanson DM, Savall BM, Coe KJ, Schoetens F, Koudriakova T, Skaptason J, Wall J, Rech J, Deng X, De Angelis M, Everson A, Lord B, Wang Q, Ao H, Scott B, Sepassi K, Lovenberg TW, Carruthers NI, Bhattacharya A, Letavic MA. Publication Year :
2016 Journal : J Med Chem Abstract : The synthesis and SAR of a series of 4,5,6,7-tetrahydro-imidazo[4,5-c]pyridine P2X7 antagonists are described. Addressing P2X7 affinity and liver microsomal stability issues encountered with this template afforded methyl substituted 4,5,6,7-tetrahydro-imidazo[4,5-c]pyridines ultimately leading to the identification of 1 (JNJ 54166060). 1 is a potent P2X7 antagonist with an ED50 = 2.3 mg/kg in rats, high oral bioavailability and low-moderate clearance in preclinical species, acceptable safety margins in rats, and a predicted human dose of 120 mg of QD. Additionally, 1 possesses a unique CYP profile and was found to be a regioselective inhibitor of midazolam CYP3A metabolism. DOI : 10.1021/acs.jmedchem.6b00989
KETOCONAZOLE
Inhibition of CYP3A4 in human liver microsomes using midazolam as substrate assessed as reduction in 4-OH-midazolam formation at 0.02 uM by LC/MS/MS analysis
Liver
Inhibition = 74.0 %
Reference
Title : Identification of (R)-(2-Chloro-3-(trifluoromethyl)phenyl)(1-(5-fluoropyridin-2-yl)-4-methyl-6,7-dihydro-1H-imidazo[4,5-c]pyridin-5(4H)-yl)methanone (JNJ 54166060), a Small Molecule Antagonist of the P2X7 receptor. Authors : Swanson DM, Savall BM, Coe KJ, Schoetens F, Koudriakova T, Skaptason J, Wall J, Rech J, Deng X, De Angelis M, Everson A, Lord B, Wang Q, Ao H, Scott B, Sepassi K, Lovenberg TW, Carruthers NI, Bhattacharya A, Letavic MA. Publication Year :
2016 Journal : J Med Chem Abstract : The synthesis and SAR of a series of 4,5,6,7-tetrahydro-imidazo[4,5-c]pyridine P2X7 antagonists are described. Addressing P2X7 affinity and liver microsomal stability issues encountered with this template afforded methyl substituted 4,5,6,7-tetrahydro-imidazo[4,5-c]pyridines ultimately leading to the identification of 1 (JNJ 54166060). 1 is a potent P2X7 antagonist with an ED50 = 2.3 mg/kg in rats, high oral bioavailability and low-moderate clearance in preclinical species, acceptable safety margins in rats, and a predicted human dose of 120 mg of QD. Additionally, 1 possesses a unique CYP profile and was found to be a regioselective inhibitor of midazolam CYP3A metabolism. DOI : 10.1021/acs.jmedchem.6b00989
KETOCONAZOLE
Inhibition of CYP3A4 in human liver microsomes using midazolam as substrate assessed as reduction in 4-OH-midazolam formation at 0.04 uM by LC/MS/MS analysis
Liver
Inhibition = 85.0 %
Reference
Title : Identification of (R)-(2-Chloro-3-(trifluoromethyl)phenyl)(1-(5-fluoropyridin-2-yl)-4-methyl-6,7-dihydro-1H-imidazo[4,5-c]pyridin-5(4H)-yl)methanone (JNJ 54166060), a Small Molecule Antagonist of the P2X7 receptor. Authors : Swanson DM, Savall BM, Coe KJ, Schoetens F, Koudriakova T, Skaptason J, Wall J, Rech J, Deng X, De Angelis M, Everson A, Lord B, Wang Q, Ao H, Scott B, Sepassi K, Lovenberg TW, Carruthers NI, Bhattacharya A, Letavic MA. Publication Year :
2016 Journal : J Med Chem Abstract : The synthesis and SAR of a series of 4,5,6,7-tetrahydro-imidazo[4,5-c]pyridine P2X7 antagonists are described. Addressing P2X7 affinity and liver microsomal stability issues encountered with this template afforded methyl substituted 4,5,6,7-tetrahydro-imidazo[4,5-c]pyridines ultimately leading to the identification of 1 (JNJ 54166060). 1 is a potent P2X7 antagonist with an ED50 = 2.3 mg/kg in rats, high oral bioavailability and low-moderate clearance in preclinical species, acceptable safety margins in rats, and a predicted human dose of 120 mg of QD. Additionally, 1 possesses a unique CYP profile and was found to be a regioselective inhibitor of midazolam CYP3A metabolism. DOI : 10.1021/acs.jmedchem.6b00989
KETOCONAZOLE
Inhibition of CYP3A4 in human liver microsomes using midazolam as substrate assessed as reduction in 4-OH-midazolam formation at 0.08 uM by LC/MS/MS analysis
Liver
Inhibition = 92.0 %
Reference
Title : Identification of (R)-(2-Chloro-3-(trifluoromethyl)phenyl)(1-(5-fluoropyridin-2-yl)-4-methyl-6,7-dihydro-1H-imidazo[4,5-c]pyridin-5(4H)-yl)methanone (JNJ 54166060), a Small Molecule Antagonist of the P2X7 receptor. Authors : Swanson DM, Savall BM, Coe KJ, Schoetens F, Koudriakova T, Skaptason J, Wall J, Rech J, Deng X, De Angelis M, Everson A, Lord B, Wang Q, Ao H, Scott B, Sepassi K, Lovenberg TW, Carruthers NI, Bhattacharya A, Letavic MA. Publication Year :
2016 Journal : J Med Chem Abstract : The synthesis and SAR of a series of 4,5,6,7-tetrahydro-imidazo[4,5-c]pyridine P2X7 antagonists are described. Addressing P2X7 affinity and liver microsomal stability issues encountered with this template afforded methyl substituted 4,5,6,7-tetrahydro-imidazo[4,5-c]pyridines ultimately leading to the identification of 1 (JNJ 54166060). 1 is a potent P2X7 antagonist with an ED50 = 2.3 mg/kg in rats, high oral bioavailability and low-moderate clearance in preclinical species, acceptable safety margins in rats, and a predicted human dose of 120 mg of QD. Additionally, 1 possesses a unique CYP profile and was found to be a regioselective inhibitor of midazolam CYP3A metabolism. DOI : 10.1021/acs.jmedchem.6b00989
KETOCONAZOLE
Inhibition of CYP3A4 in human liver microsomes using midazolam as substrate assessed as reduction in 4-OH-midazolam formation at 0.16 uM by LC/MS/MS analysis
Liver
Inhibition = 95.0 %
Reference
Title : Identification of (R)-(2-Chloro-3-(trifluoromethyl)phenyl)(1-(5-fluoropyridin-2-yl)-4-methyl-6,7-dihydro-1H-imidazo[4,5-c]pyridin-5(4H)-yl)methanone (JNJ 54166060), a Small Molecule Antagonist of the P2X7 receptor. Authors : Swanson DM, Savall BM, Coe KJ, Schoetens F, Koudriakova T, Skaptason J, Wall J, Rech J, Deng X, De Angelis M, Everson A, Lord B, Wang Q, Ao H, Scott B, Sepassi K, Lovenberg TW, Carruthers NI, Bhattacharya A, Letavic MA. Publication Year :
2016 Journal : J Med Chem Abstract : The synthesis and SAR of a series of 4,5,6,7-tetrahydro-imidazo[4,5-c]pyridine P2X7 antagonists are described. Addressing P2X7 affinity and liver microsomal stability issues encountered with this template afforded methyl substituted 4,5,6,7-tetrahydro-imidazo[4,5-c]pyridines ultimately leading to the identification of 1 (JNJ 54166060). 1 is a potent P2X7 antagonist with an ED50 = 2.3 mg/kg in rats, high oral bioavailability and low-moderate clearance in preclinical species, acceptable safety margins in rats, and a predicted human dose of 120 mg of QD. Additionally, 1 possesses a unique CYP profile and was found to be a regioselective inhibitor of midazolam CYP3A metabolism. DOI : 10.1021/acs.jmedchem.6b00989
KETOCONAZOLE
Inhibition of recombinant human CYP3A4 expressed in insect cells at >= 1 uM using DBOMF as substrate pretreated for 10 mins followed by substrate addition measured every minute for 15 mins in presence of NADP+ by fluorescence assay relative to control
—
Inhibition > 85.0 %
Reference
Title : Cytochrome P450 binding studies of novel tacrine derivatives: Predicting the risk of hepatotoxicity. Authors : McEneny-King A, Osman W, Edginton AN, Rao PPN. Publication Year :
2017 Journal : Bioorg Med Chem Lett Abstract : The 1,2,3,4-tetrahydroacridine derivative tacrine was the first drug approved to treat Alzheimer's disease (AD). It is known to act as a potent cholinesterase inhibitor. However, tacrine was removed from the market due to its hepatotoxicity concerns as it undergoes metabolism to toxic quinonemethide species through the cytochrome P450 enzyme CYP1A2. Despite these challenges, tacrine serves as a useful template in the development of novel multi-targeting anti-AD agents. In this regard, we sought to evaluate the risk of hepatotoxicity in a series of C9 substituted tacrine derivatives that exhibit cholinesterase inhibition properties. The hepatotoxic potential of tacrine derivatives was evaluated using recombinant cytochrome (CYP) P450 CYP1A2 and CYP3A4 enzymes. Molecular docking studies were conducted to predict their binding modes and potential risk of forming hepatotoxic metabolites. Tacrine derivatives compound 1 (N-(3,4-dimethoxybenzyl)-1,2,3,4-tetrahydroacridin-9-amine) and 2 (6-chloro-N-(3,4-dimethoxybenzyl)-1,2,3,4-tetrahydroacridin-9-amine) which possess a C9 3,4-dimethoxybenzylamino substituent exhibited weak binding to CYP1A2 enzyme (1, IC50=33.0µM; 2, IC50=8.5µM) compared to tacrine (CYP1A2 IC50=1.5µM). Modeling studies show that the presence of a bulky 3,4-dimethoxybenzylamino C9 substituent prevents the orientation of the 1,2,3,4-tetrahydroacridine ring close to the heme-iron center of CYP1A2 thereby reducing the risk of forming hepatotoxic species. DOI : 10.1016/j.bmcl.2017.04.006
KETOCONAZOLE
Inhibition of recombinant human Cytochrome P450 3A4 with BFC [7-benzyloxy-4-trifluoromethylcoumarin] after 15 minutes
—
IC50 = 13.0 nM
Reference
Title : Fluorine substitution can block CYP3A4 metabolism-dependent inhibition: identification of (S)-N-[1-(4-fluoro-3- morpholin-4-ylphenyl)ethyl]-3- (4-fluorophenyl)acrylamide as an orally bioavailable KCNQ2 opener devoid of CYP3A4 metabolism-dependent inhibition. Authors : Wu YJ, Davis CD, Dworetzky S, Fitzpatrick WC, Harden D, He H, Knox RJ, Newton AE, Philip T, Polson C, Sivarao DV, Sun LQ, Tertyshnikova S, Weaver D, Yeola S, Zoeckler M, Sinz MW. Publication Year :
2003 Journal : J. Med. Chem. Abstract : The formation of a reactive intermediate was found to be responsible for CYP3A4 metabolism-dependent inhibition (MDI) observed with (S)-N-[1-(3-morpholin-4-ylphenyl)ethyl]-3-phenyl-acrylamide (1). Structure-3A4 MDI relationship studies culminated in the discovery of a difluoro analogue, (S)-N-[1-(4-fluoro-3-morpholin-4-ylphenyl)ethyl]-3-(4-fluoro-phenyl)acrylamide (2), as an orally bioavailable KCNQ2 opener free of CYP3A4 MDI. DOI : 10.1021/jm034111v
KETOCONAZOLE
Inhibition of recombinant human Cytochrome P450 3A4 using BFC [7-benzyloxy-4-trifluoromethylcoumarin] after 30 minutes
—
IC50 = 17.0 nM
Reference
Title : Fluorine substitution can block CYP3A4 metabolism-dependent inhibition: identification of (S)-N-[1-(4-fluoro-3- morpholin-4-ylphenyl)ethyl]-3- (4-fluorophenyl)acrylamide as an orally bioavailable KCNQ2 opener devoid of CYP3A4 metabolism-dependent inhibition. Authors : Wu YJ, Davis CD, Dworetzky S, Fitzpatrick WC, Harden D, He H, Knox RJ, Newton AE, Philip T, Polson C, Sivarao DV, Sun LQ, Tertyshnikova S, Weaver D, Yeola S, Zoeckler M, Sinz MW. Publication Year :
2003 Journal : J. Med. Chem. Abstract : The formation of a reactive intermediate was found to be responsible for CYP3A4 metabolism-dependent inhibition (MDI) observed with (S)-N-[1-(3-morpholin-4-ylphenyl)ethyl]-3-phenyl-acrylamide (1). Structure-3A4 MDI relationship studies culminated in the discovery of a difluoro analogue, (S)-N-[1-(4-fluoro-3-morpholin-4-ylphenyl)ethyl]-3-(4-fluoro-phenyl)acrylamide (2), as an orally bioavailable KCNQ2 opener free of CYP3A4 MDI. DOI : 10.1021/jm034111v
KETOCONAZOLE
Inhibition of recombinant human Cytochrome P450 3A4 with BFC [7-benzyloxy-4-trifluoromethylcoumarin] after 45 minutes
—
IC50 = 22.0 nM
Reference
Title : Fluorine substitution can block CYP3A4 metabolism-dependent inhibition: identification of (S)-N-[1-(4-fluoro-3- morpholin-4-ylphenyl)ethyl]-3- (4-fluorophenyl)acrylamide as an orally bioavailable KCNQ2 opener devoid of CYP3A4 metabolism-dependent inhibition. Authors : Wu YJ, Davis CD, Dworetzky S, Fitzpatrick WC, Harden D, He H, Knox RJ, Newton AE, Philip T, Polson C, Sivarao DV, Sun LQ, Tertyshnikova S, Weaver D, Yeola S, Zoeckler M, Sinz MW. Publication Year :
2003 Journal : J. Med. Chem. Abstract : The formation of a reactive intermediate was found to be responsible for CYP3A4 metabolism-dependent inhibition (MDI) observed with (S)-N-[1-(3-morpholin-4-ylphenyl)ethyl]-3-phenyl-acrylamide (1). Structure-3A4 MDI relationship studies culminated in the discovery of a difluoro analogue, (S)-N-[1-(4-fluoro-3-morpholin-4-ylphenyl)ethyl]-3-(4-fluoro-phenyl)acrylamide (2), as an orally bioavailable KCNQ2 opener free of CYP3A4 MDI. DOI : 10.1021/jm034111v
KETOCONAZOLE
Inhibition of recombinant human Cytochrome P450 3A4 with BFC [7-benzyloxy-4-trifluoromethylcoumarin] after 5 minutes
—
IC50 = 16.0 nM
Reference
Title : Fluorine substitution can block CYP3A4 metabolism-dependent inhibition: identification of (S)-N-[1-(4-fluoro-3- morpholin-4-ylphenyl)ethyl]-3- (4-fluorophenyl)acrylamide as an orally bioavailable KCNQ2 opener devoid of CYP3A4 metabolism-dependent inhibition. Authors : Wu YJ, Davis CD, Dworetzky S, Fitzpatrick WC, Harden D, He H, Knox RJ, Newton AE, Philip T, Polson C, Sivarao DV, Sun LQ, Tertyshnikova S, Weaver D, Yeola S, Zoeckler M, Sinz MW. Publication Year :
2003 Journal : J. Med. Chem. Abstract : The formation of a reactive intermediate was found to be responsible for CYP3A4 metabolism-dependent inhibition (MDI) observed with (S)-N-[1-(3-morpholin-4-ylphenyl)ethyl]-3-phenyl-acrylamide (1). Structure-3A4 MDI relationship studies culminated in the discovery of a difluoro analogue, (S)-N-[1-(4-fluoro-3-morpholin-4-ylphenyl)ethyl]-3-(4-fluoro-phenyl)acrylamide (2), as an orally bioavailable KCNQ2 opener free of CYP3A4 MDI. DOI : 10.1021/jm034111v
KETOCONAZOLE
Inhibitory concentration against human hepatic cytochrome P450 3A4 enzyme
—
IC50 = 50.0 nM
Reference
Title : Heteroaryl-substituted naphthalenes and structurally modified derivatives: selective inhibitors of CYP11B2 for the treatment of congestive heart failure and myocardial fibrosis. Authors : Voets M, Antes I, Scherer C, Müller-Vieira U, Biemel K, Barassin C, Marchais-Oberwinkler S, Hartmann RW. Publication Year :
2005 Journal : J. Med. Chem. Abstract : Recently we proposed inhibition of aldosterone synthase (CYP11B2) as a novel strategy for the treatment of congestive heart failure and myocardial fibrosis. In this study the synthesis and biological evaluation of heteroaryl-substituted naphthalenes and quinolines (1-31) is described. Key step for the preparation of the compounds was a Suzuki cross-coupling. Activity of the compounds was determined in vitro using human CYP11B2 and selectivity was evaluated toward the human steroidogenic enzymes CYP11B1, CYP19, and CYP17. A large number of highly active and selective inhibitors of CYP11B2 was identified. The most active inhibitor was the 6-cyano compound 8 (IC50 = 3 nM) showing a competitive type of inhibition (K(i) value = 1.9 nM). The 6-ethoxy derivative 5 was found to be the most selective CYP11B2 inhibitor (IC50 = 12 nM; K(i) value = 8 nM; CYP11B1 IC50 = 5419 nM; selectivity factor = 451), showing no inhibition of human CYP3A4 (50 nM) and CYP2D6 (20 nM). Docking and molecular dynamics studies using our homology modeled CYP11B2 structure with selected compounds were performed. Caco-2 cell experiments revealed a large number of medium and highly permeable compounds and metabolic studies with 2 using rat liver microsomes showed sufficient stability. DOI : 10.1021/jm0503704
KETOCONAZOLE
Inhibition of human CYP3A4
—
IC50 = 50.0 nM
Reference
Title : Synthesis and evaluation of heteroaryl-substituted dihydronaphthalenes and indenes: potent and selective inhibitors of aldosterone synthase (CYP11B2) for the treatment of congestive heart failure and myocardial fibrosis. Authors : Voets M, Antes I, Scherer C, Müller-Vieira U, Biemel K, Marchais-Oberwinkler S, Hartmann RW. Publication Year :
2006 Journal : J. Med. Chem. Abstract : In this study, the synthesis and biological evaluation of heteroaryl-substituted dihydronaphthalenes and indenes (1-16) is described. The compounds were tested for activity by use of human CYP11B2 expressed in fission yeast and V79 MZh cells and for selectivity by use of human CYP11B1, CYP17, and CYP19. The most active inhibitor was the 6-methoxydihydronaphthalene 4 (IC(50) = 2 nM), showing a K(i) value of 1.3 nM and a competitive type of inhibition. The 5-methoxyindene 3 was found to be the most selective CYP11B2 inhibitor (IC(50) = 4 nM; CYP11B1 IC(50) = 5684 nM), which also showed only marginal inhibition of human CYP3A4 and CYP2D6. Docking and molecular dynamics studies using our homology-modeled CYP11B2 structure were performed to understand some structure-activity relationships. Caco-2 cell experiments revealed highly cell-permeable compounds, and metabolic studies with 4 using rat liver microsomes showed sufficient stability. DOI : 10.1021/jm060055x
KETOCONAZOLE
Inhibition of human CYP3A4 expressed in Escherichia coli assessed as inhibition of nifedipine oxidation
—
IC50 = 210.0 nM
Reference
Title : Cytochrome P3A4 inhibitors and other constituents of Fibraurea tinctoria. Authors : Su CR, Ueng YF, Dung NX, Vijaya Bhaskar Reddy M, Wu TS. Publication Year :
2007 Journal : J. Nat. Prod. Abstract : Four new furanoditerpenoids, fibrauretin A ( 1), fibrauretinoside A ( 2), epi-fibrauretinoside A ( 3), and epi-12-palmatoside G ( 4), and a new ecdysteroid glucoside, fibraurecdyside A ( 5), together with seven known compounds including two furanoditerpenoids ( 6 and 7), an ecdysteroid ( 8), and four quaternary protoberberine alkaloids ( 9- 12) were isolated from the stems of Fibraurea tinctoria. The structures of 1- 5 were established on the basis of spectroscopic evidence. Among these compounds, palmatine ( 9) and jatrorrhizine ( 10) showed inhibitory effects against cytochrome P450 3A4 (CYP3A4) with IC 50 values of 0.9 and 2.1 microM, respectively. DOI : 10.1021/np0704248
KETOCONAZOLE
Inhibition of human CYP3A4
—
IC50 = 54.0 nM
Reference
Title : Design, synthesis, and biological evaluation of (hydroxyphenyl)naphthalene and -quinoline derivatives: potent and selective nonsteroidal inhibitors of 17beta-hydroxysteroid dehydrogenase type 1 (17beta-HSD1) for the treatment of estrogen-dependent diseases. Authors : Frotscher M, Ziegler E, Marchais-Oberwinkler S, Kruchten P, Neugebauer A, Fetzer L, Scherer C, Müller-Vieira U, Messinger J, Thole H, Hartmann RW. Publication Year :
2008 Journal : J. Med. Chem. Abstract : Human 17beta-hydroxysteroid dehydrogenase type 1 (17beta-HSD1) catalyzes the reduction of the weak estrogen estrone (E1) to the highly potent estradiol (E2). This reaction takes place in the target cell where the estrogenic effect is exerted via the estrogen receptor (ER). Estrogens, especially E2, are known to stimulate the proliferation of hormone-dependent diseases. 17beta-HSD1 is overexpressed in many breast tumors. Thus, it is an attractive target for the treatment of these diseases. Ligand- and structure-based drug design led to the discovery of novel, selective, and potent inhibitors of 17beta-HSD1. Phenyl-substituted bicyclic moieties were synthesized as mimics of the steroidal substrate. Computational methods were used to obtain insight into their interactions with the protein. Compound 5 turned out to be a highly potent inhibitor of 17beta-HSD1 showing good selectivity (17beta-HSD2, ERalpha and beta), medium cell permeation, reasonable metabolic stability (rat hepatic microsomes), and little inhibition of hepatic CYP enzymes. DOI : 10.1021/jm701447v
KETOCONAZOLE
Inhibition of CYP3A4 in human liver microsome
Liver
IC50 = 20.0 nM
Reference
Title : Synthesis of novel ketoconazole derivatives as inhibitors of the human Pregnane X Receptor (PXR; NR1I2; also termed SXR, PAR). Authors : Das BC, Madhukumar AV, Anguiano J, Kim S, Sinz M, Zvyaga TA, Power EC, Ganellin CR, Mani S. Publication Year :
2008 Journal : Bioorg. Med. Chem. Lett. Abstract : PXR, pregnane X receptor, in its activated state, is a validated target for controlling certain drug-drug interactions in humans. In this context, there is a paucity of inhibitors directed toward activated PXR. Using prior observations with ketoconazole as a PXR inhibitor, the target compound 3 was synthesized from (s)-glycidol with overall 56% yield. (+)-Glycidol was reacted with 4-bromophenol and potassium carbonate in DMF to yield the ring opened compound 6. This was then heated to reflux in benzene along with 2', 4'-difluoroacetophenone and catalytic amount of para-toluene sulfonic acid to yield 8. The resultant acetal 8 was then functionalized using Palladium chemistry to yield the target compound 3. The activity of the compound was compared with ketoconazole and UCL2158H. However, in contrast with ketoconazole (IC(50) approximately 0.020 microM; approximately 100% inhibition), 3 has negligible effects on inhibition of microsomal CYP450 (maximum approximately 20% inhibition) at concentrations >40 microM. In vitro, micromolar concentration of ketoconazole is toxic to passaged human cell lines, while 3 does not exhibit cytotoxicity up to concentrations approximately 100 microM (viability >85%). This is the first demonstration of a chemical analog of a PXR inhibitor that retains activity against activated PXR. Furthermore, in contrast with ketoconazole, 3 is less toxic in human cell lines and has negligible CYP450 activity. DOI : 10.1016/j.bmcl.2008.06.018
KETOCONAZOLE
Inhibition of human recombinant CYP3A4 expressed in insect microsomes
—
IC50 = 57.0 nM
Reference
Title : Overcoming undesirable CYP1A2 inhibition of pyridylnaphthalene-type aldosterone synthase inhibitors: influence of heteroaryl derivatization on potency and selectivity. Authors : Heim R, Lucas S, Grombein CM, Ries C, Schewe KE, Negri M, Müller-Vieira U, Birk B, Hartmann RW. Publication Year :
2008 Journal : J. Med. Chem. Abstract : Recently, we reported on the development of potent and selective inhibitors of aldosterone synthase (CYP11B2) for the treatment of congestive heart failure and myocardial fibrosis. A major drawback of these nonsteroidal compounds was a strong inhibition of the hepatic drug-metabolizing enzyme CYP1A2. In the present study, we examined the influence of substituents in the heterocycle of lead structures with a naphthalene molecular scaffold to overcome this unwanted side effect. With respect to CYP11B2 inhibition, some substituents induced a dramatic increase in inhibitory potency. The methoxyalkyl derivatives 22 and 26 are the most potent CYP11B2 inhibitors up to now (IC50 = 0.2 nM). Most compounds also clearly discriminated between CYP11B2 and CYP11B1, and the CYP1A2 potency significantly decreased in some cases (e.g., isoquinoline derivative 30 displayed only 6% CYP1A2 inhibition at 2 microM concentration). Furthermore, isoquinoline derivative 28 proved to be capable of passing the gastrointestinal tract and reached the general circulation after peroral administration to male Wistar rats. DOI : 10.1021/jm800377h
KETOCONAZOLE
Inhibition of human recombinant CYP3A4 expressed in baculovirus-infected insect microsomes
—
IC50 = 57.0 nM
Reference
Title : Novel aldosterone synthase inhibitors with extended carbocyclic skeleton by a combined ligand-based and structure-based drug design approach. Authors : Lucas S, Heim R, Negri M, Antes I, Ries C, Schewe KE, Bisi A, Gobbi S, Hartmann RW. Publication Year :
2008 Journal : J. Med. Chem. Abstract : Pharmacophore modeling of a series of aldosterone synthase (CYP11B2) inhibitors triggered the design of compounds 11 and 12 by extending a previously established naphthalene molecular scaffold (e.g., present in molecules 1 and 2) via introduction of a phenyl or benzyl residue in 3-position. These additional aromatic moieties have been hypothesized to fit into the newly identified hydrophobic pharmacophore feature HY3. Subsequent docking studies in our refined CYP11B2 protein model have been performed prior to synthesis to estimate the inhibitory properties of the proposed molecules. While phenyl-substituted compound 11 (IC50 > 500 nM) did not dock under the given pharmacophore constraint (i.e., the Fe(heme)-N(ligand) interaction), benzyl-substituted compound 12 (IC50 = 154 nM) was found to exploit a previously unexplored subpocket of the inhibitor binding site. By structural optimization based on the pharmacophore hypothesis, 25 novel compounds were synthesized, among them highly potent CYP11B2 inhibitors (e.g., 17, IC50 = 2.7 nM) with pronounced selectivity toward the most important steroidogenic and hepatic CYP enzymes. DOI : 10.1021/jm800683c
KETOCONAZOLE
Inhibition of human liver microsome CYP3A4 in assessed as [14C]formaldehyde formation
Liver
IC50 = 720.0 nM
Reference
Title : Potent CYP3A4 inhibitory constituents of Piper cubeba. Authors : Usia T, Watabe T, Kadota S, Tezuka Y. Publication Year :
2005 Journal : J. Nat. Prod. Abstract : The EtOAc-soluble fraction of the water extract of Piper cubeba, having shown potent inhibitory activity on the metabolism mediated by CYP3A4, was subjected to activity-guided isolation to yield two new lignans, (8R,8'R)-4-hydroxycubebinone (1) and (8R,8'R,9'S)-5-methoxyclusin (2), and two new sesquiterpenes, (5 alpha,8 alpha)-2-oxo-1(10),3,7(11)-guaiatrien-12,8-olide (3) and (1 alpha,2 beta,5 alpha,8 alpha 10 alpha)-1,10-epoxy-2-hydroxy-3,7(11)-guaiadien-12,8-olide (4), along with 16 known compounds (5-20). The structures of the isolated compounds were elucidated on the basis of spectroscopic and chemical analyses. The isolated compounds were tested for their inhibitory activity on the metabolism mediated by CYP3A4 or CYP2D6 using [N-methyl-(14)C]erythromycin or [O-methyl-(14)C]dextromethorphan as a substrate, respectively. The compounds (8R,8'R,9'S)-5-methoxyclusin (2), (-)-clusin (10), (-)-yatein (13), ethoxyclusin (15), and (-)-dihydroclusin (17), having one methylenedioxyphenyl moiety in their structures, showed very potent and selective inhibitory activity against CYP3A4 with IC(50) values (0.44-1.0 microM) identical to that of the positive control, ketoconazole (IC(50), 0.72 microM). DOI : 10.1021/np0401765
KETOCONAZOLE
Inhibition of human recombinant CYP3A4
—
IC50 = 40.0 nM
Reference
Title : Rational modification of a candidate cancer drug for use against Chagas disease. Authors : Kraus JM, Verlinde CL, Karimi M, Lepesheva GI, Gelb MH, Buckner FS. Publication Year :
2009 Journal : J. Med. Chem. Abstract : Chagas disease is one of the major neglected diseases of the world. Existing drug therapies are limited, ineffective, and highly toxic. We describe a novel strategy of drug discovery of adapting an existing clinical compound with excellent pharmaceutical properties to target a pathogenic organism. The protein farnesyltransferase (PFT) inhibitor tipifarnib, now in phase III anticancer clinical trials, was previously found to kill Trypanosoma cruzi by blocking sterol 14 alpha-demethylase (14DM). We rationally developed tipifarnib analogues that display reduced affinity for human PFT to reduce toxicity while increasing affinity for parasite 14DM. The lead compound has picomolar activity against cultured T. cruzi and is efficacious in a mouse model of acute Chagas disease. DOI : 10.1021/jm801313t
KETOCONAZOLE
Inhibition of human CYP3A4 by radiometric assay
—
IC50 = 245.0 nM
Reference
Title : Sesquiterpenes and flavonol glycosides from Zingiber aromaticum and their CYP3A4 and CYP2D6 inhibitory activities. Authors : Usia T, Iwata H, Hiratsuka A, Watabe T, Kadota S, Tezuka Y. Publication Year :
2004 Journal : J. Nat. Prod. Abstract : Three new sesquiterpenes, (2R,3S,5R)-2,3-epoxy-6,9-humuladien-5-ol-8-one (1), (2R,3R,5R)-2,3-epoxy-6,9-humuladien-5-ol-8-one (2), and (5R)-2,6,9-humulatrien-5-ol-8-one (3), and two new flavonol glycosides, kaempferol-3-O-(2,3-di-O-acetyl-alpha-l-rhamnopyranoside) (4) and kaempferol-3-O-(2,3,4-tri-O-acetyl-alpha-l-rhamnopyranoside) (5), were isolated from the EtOAc-soluble fraction of the water extract of Zingiber aromaticum, along with 13 known compounds (6-18). The structures of the isolated compounds were elucidated on the basis of spectroscopic and chemical analyses. The isolated compounds were tested for their inhibitory activity on the metabolism mediated by CYP3A4 or CYP2D6 using [N-methyl-(14)C]erythromycin or [O-methyl-(14)C]dextromethorphan as a substrate, respectively. Kaempferol-3-O-(2,3,4-tri-O-acetyl-alpha-l-rhamnopyranoside) (5) showed the most potent inhibitory activity (IC(50), 14.4 microM) on the metabolism mediated by CYP3A4, and kaempferol-3-O-methyl ether (14) inhibited CYP2D6 most potently (IC(50), 4.63 microM). DOI : 10.1021/np030556a
KETOCONAZOLE
Inhibition of human CYP3A4 assessed as biotransformation of 7-benzyloxyquinoline
—
Ki = 24.0 nM
Reference
Title : Selective cytochrome P450 3A4 inhibitory activity of Amaryllidaceae alkaloids. Authors : McNulty J, Nair JJ, Singh M, Crankshaw DJ, Holloway AC, Bastida J. Publication Year :
2009 Journal : Bioorg. Med. Chem. Lett. Abstract : A library of natural and semi-synthetic Amaryllidaceae alkaloids was screened for cytochrome P450 3A4 (CYP3A4) inhibitory activity. Of the crinane, lycorane and galanthamine representatives examined two semi-synthetic silylated lycorane analogues, accessed via a chemoselective silylation strategy from lycorine, and the natural compound narciclasine exhibited low micromolar activities. Important pharmacological features uncovered include the lack of CYP3A4 inhibitory activity seen for galanthamine and the selective activity that is seen with narciclasine over pancratistatin. DOI : 10.1016/j.bmcl.2009.04.086
KETOCONAZOLE
Inhibition of human CYP3A4 assessed as biotransformation of 7-benzyloxyquinoline
—
Ki = 25.12 nM
Reference
Title : Selective cytochrome P450 3A4 inhibitory activity of Amaryllidaceae alkaloids. Authors : McNulty J, Nair JJ, Singh M, Crankshaw DJ, Holloway AC, Bastida J. Publication Year :
2009 Journal : Bioorg. Med. Chem. Lett. Abstract : A library of natural and semi-synthetic Amaryllidaceae alkaloids was screened for cytochrome P450 3A4 (CYP3A4) inhibitory activity. Of the crinane, lycorane and galanthamine representatives examined two semi-synthetic silylated lycorane analogues, accessed via a chemoselective silylation strategy from lycorine, and the natural compound narciclasine exhibited low micromolar activities. Important pharmacological features uncovered include the lack of CYP3A4 inhibitory activity seen for galanthamine and the selective activity that is seen with narciclasine over pancratistatin. DOI : 10.1016/j.bmcl.2009.04.086
KETOCONAZOLE
Inhibition of human recombinant CYP3A4 assessed as biotransformation of 7-benzyloxyquinoline to 7-hydroxyquinoline measured every 15 mins by fluorescence intensity analysis
—
Ki = 30.0 nM
Reference
Title : Structure-activity studies on seco-pancratistatin analogs: potent inhibitors of human cytochrome P450 3A4. Authors : McNulty J, Nair JJ, Singh M, Crankshaw DJ, Holloway AC. Publication Year :
2009 Journal : Bioorg. Med. Chem. Lett. Abstract : Two total syntheses of fully functionalized seco-analogs of the anticancer compound pancratistatin are reported. Structure-activity relationship (SAR) studies identified potent and selective inhibitors of human cytochrome P450 3A4 (CYP3A4) and revealed several core pharmacophoric elements. These studies identify potential roadblocks and will guide the further development of a viable selective clinical pancratistatin derivative. DOI : 10.1016/j.bmcl.2009.08.032
KETOCONAZOLE
Inhibition of human recombinant CYP3A4 assessed as biotransformation of 7-benzyloxyquinoline to 7-hydroxyquinoline measured every 15 mins by fluorescence intensity analysis
—
Ki = 33.11 nM
Reference
Title : Structure-activity studies on seco-pancratistatin analogs: potent inhibitors of human cytochrome P450 3A4. Authors : McNulty J, Nair JJ, Singh M, Crankshaw DJ, Holloway AC. Publication Year :
2009 Journal : Bioorg. Med. Chem. Lett. Abstract : Two total syntheses of fully functionalized seco-analogs of the anticancer compound pancratistatin are reported. Structure-activity relationship (SAR) studies identified potent and selective inhibitors of human cytochrome P450 3A4 (CYP3A4) and revealed several core pharmacophoric elements. These studies identify potential roadblocks and will guide the further development of a viable selective clinical pancratistatin derivative. DOI : 10.1016/j.bmcl.2009.08.032
KETOCONAZOLE
Inhibition of human CYP3A4
—
IC50 = 4.0 nM
Reference
Title : A novel class of highly potent multidrug resistance reversal agents: disubstituted adamantyl derivatives. Authors : Min KH, Xia Y, Kim EK, Jin Y, Kaur N, Kim ES, Kim DK, Jung HY, Choi Y, Park MK, Min YK, Lee K, Lee K. Publication Year :
2009 Journal : Bioorg. Med. Chem. Lett. Abstract : Novel disubstituted adamantyl derivatives were synthesized and evaluated in a P-glycoprotein dependent multidrug resistance cancer cell line. The hit to lead optimization provided potent MDR reversal agents. Some potent adamantyl derivatives were more than 10-fold more potent than verapamil without considerable intrinsic cytotoxicity. The 3-trifluorophenyl derivative 14f did not affect the metabolism of CYP450 3A4, whereas most of MDR revertants had a weak inhibitory effect. DOI : 10.1016/j.bmcl.2009.07.127
KETOCONAZOLE
Inhibition of human recombinant CYP3A4 assessed as biotransformation of 7-hydroxyquinoline to 7-benzyloxyquinoline measured every minute for 15 mins by fluorescence intensity assay
—
Ki = 30.0 nM
Reference
Title : Potent and selective inhibition of human cytochrome P450 3A4 by seco-pancratistatin structural analogs. Authors : McNulty J, Nair JJ, Singh M, Crankshaw DJ, Holloway AC. Publication Year :
2010 Journal : Bioorg. Med. Chem. Lett. Abstract : seco-Derivatives of the anticancer agent pancratistatin bearing the 2S,3S,4S,5S configuration were accessed via a novel, highly diastereoselective anti-aldol reaction. Structure-activity relationships reveal important insights into the seco-pancratistatin pharmacophore as a potent and selective inhibitor of human cytochrome P450 3A4 (CYP3A4), and highlight features of concern in advancing a potent, selective anticancer agent in the pancratistatin series. DOI : 10.1016/j.bmcl.2010.01.157
KETOCONAZOLE
Inhibition of human recombinant CYP3A4 assessed as biotransformation of 7-hydroxyquinoline to 7-benzyloxyquinoline measured every minute for 15 mins by fluorescence intensity assay
—
Ki = 33.11 nM
Reference
Title : Potent and selective inhibition of human cytochrome P450 3A4 by seco-pancratistatin structural analogs. Authors : McNulty J, Nair JJ, Singh M, Crankshaw DJ, Holloway AC. Publication Year :
2010 Journal : Bioorg. Med. Chem. Lett. Abstract : seco-Derivatives of the anticancer agent pancratistatin bearing the 2S,3S,4S,5S configuration were accessed via a novel, highly diastereoselective anti-aldol reaction. Structure-activity relationships reveal important insights into the seco-pancratistatin pharmacophore as a potent and selective inhibitor of human cytochrome P450 3A4 (CYP3A4), and highlight features of concern in advancing a potent, selective anticancer agent in the pancratistatin series. DOI : 10.1016/j.bmcl.2010.01.157
KETOCONAZOLE
Inhibition of human recombinant CYP3A4
—
IC50 = 11.0 nM
Reference
Title : Second generation analogues of the cancer drug clinical candidate tipifarnib for anti-Chagas disease drug discovery. Authors : Kraus JM, Tatipaka HB, McGuffin SA, Chennamaneni NK, Karimi M, Arif J, Verlinde CL, Buckner FS, Gelb MH. Publication Year :
2010 Journal : J. Med. Chem. Abstract : We previously reported that the cancer drug clinical candidate tipifarnib kills the causative agent of Chagas disease, Trypanosoma cruzi, by blocking ergosterol biosynthesis at the level of inhibition of lanosterol 14alpha-demethylase. Tipifarnib is an inhibitor of human protein farnesyltransferase. We synthesized tipifarnib analogues that no longer bind to protein farnesyltransferase and display increased potency for killing parasites. This was achieved in a structure-guided fashion by changing the substituents attached to the phenyl group at the 4-position of the quinoline ring of tipifarnib and by replacing the amino group by OMe. Several compounds that kill Trypanosoma cruzi at subnanomolar concentrations and are devoid of protein farnesyltransferase inhibition were discovered. The compounds are shown to be advantageous over other lanosterol 14alpha-demethylase inhibitors in that they show only modest potency for inhibition of human cytochrome P450 (3A4). Since tipifarnib displays high oral bioavailability and acceptable pharmacokinetic properties, the newly discovered tipifarnib analogues are ideal leads for the development of drugs to treat Chagas disease. DOI : 10.1021/jm9013136
KETOCONAZOLE
Inhibition of human CYP3A4
—
IC50 = 50.0 nM
Reference
Title : New drug-like hydroxyphenylnaphthol steroidomimetics as potent and selective 17β-hydroxysteroid dehydrogenase type 1 inhibitors for the treatment of estrogen-dependent diseases. Authors : Marchais-Oberwinkler S, Wetzel M, Ziegler E, Kruchten P, Werth R, Henn C, Hartmann RW, Frotscher M. Publication Year :
2011 Journal : J. Med. Chem. Abstract : Inhibition of 17β-hydroxysteroid dehydrogenase type 1 (17β-HSD1) is a novel and attractive approach to reduce the local levels of the active estrogen 17β-estradiol in patients with estrogen-dependent diseases like breast cancer or endometriosis. With the aim of optimizing the biological profile of 17β-HSD1 inhibitors from the hydroxyphenylnaphthol class, structural optimizations were performed at the 1-position of the naphthalene by introduction of different heteroaromatic rings as well as substituted phenyl groups. In the latter class of compounds, which were synthesized applying Suzuki-cross coupling, the 3-methanesulfonamide 15 turned out to be a highly potent 17β-HSD1 inhibitor (IC(50) = 15 nM in a cell-free assay). It was also very active in the cellular assay (T47D cells, IC(50) = 71 nM) and selective toward 17β-HSD2 and the estrogen receptors α and β. It showed a good membrane permeation and metabolic stability and was orally available in the rat. DOI : 10.1021/jm1009082
KETOCONAZOLE
Inhibition of human CYP3A4 expressed in baculovirus-infected insect microsomes
—
IC50 = 57.0 nM
Reference
Title : Fine-tuning the selectivity of aldosterone synthase inhibitors: structure-activity and structure-selectivity insights from studies of heteroaryl substituted 1,2,5,6-tetrahydropyrrolo[3,2,1-ij]quinolin-4-one derivatives. Authors : Lucas S, Negri M, Heim R, Zimmer C, Hartmann RW. Publication Year :
2011 Journal : J. Med. Chem. Abstract : Pyridine substituted 3,4-dihydro-1H-quinolin-2-ones (e.g., 1-3) constitute a class of highly potent and selective inhibitors of aldosterone synthase (CYP11B2), a promising target for the treatment of hyperaldosteronism, congestive heart failure, and myocardial fibrosis. Among these, ethyl-substituted 3 possesses high selectivity against CYP1A2. Rigidification of 3 by incorporation of the ethyl group into a 5- or 6-membered ring affords compounds with a pyrroloquinolinone or pyridoquinolinone molecular scaffold (e.g., 4 and 5). It was found that these molecules are even more potent and selective CYP11B2 inhibitors than their corresponding open-chain analogues. Moreover, pyrroloquinolinone 4 exhibits no inhibition of the six most important hepatic CYP enzymes as well as a bioavailability in the range of the marketed drug fadrozole. The SAR studies disclose that subtle changes in the heterocyclic moiety are responsible for either a strong or a weak inhibition of the highly homologous 11β-hydroxylase (CYP11B1). These results are not only important for fine-tuning the selectivity of CYP11B2 inhibitors but also for the development of selective CYP11B1 inhibitors that are of interest for the treatment of Cushing's syndrome and metabolic syndrome. DOI : 10.1021/jm101470k
KETOCONAZOLE
Inhibition of human CYP3A4 in liver microsomes
Liver
IC50 = 70.0 nM
Reference
Title : Preclinical metabolism of LB42908, a novel farnesyl transferase inhibitor, and its effects on the cytochrome P450 isozyme activities. Authors : Kim HJ, Lee SH, Koh JS, Lee SH. Publication Year :
2012 Journal : Bioorg. Med. Chem. Lett. Abstract : Metabolism of LB42908, a novel farnesyl transferase inhibitor, was investigated for preclinical development. In vitro hepatic metabolism of LB42908 gave rise to at least 9 metabolites via phase I biotransformation pathways, which were characterized by HPLC-UV, LC-MS, and LC-MS/MS analyses. N-Dealkylation was shown to be a major phase I metabolic pathway. Species-specific in vitro metabolism of LB42908 was studied in liver fractions of rat, dog, monkey, and human. Order of metabolic stability is human≈dog>rat≈monkey in both S9 and microsomal fractions. Tissue-specific metabolism of LB42908 in various tissue homogenates of rats demonstrated that the liver was the major organ responsible for phase I metabolism of LB42908. The results from both qualitative and quantitative metabolism studies such as metabolic profiling and metabolic clearance indicated that dog would be the animal model of choice for preclinical toxicology studies. In addition, LB42908 was a potent CYP3A4 inhibitor in human liver microsomes and induced the activities of several CYP isozymes, implying that it has the potential for drug-drug interactions. Repeated dosing of LB42908 in rats did not significantly affect its own metabolism, indicating that long-term administration of LB42908 would not alter its pharmacokinetic profiles. DOI : 10.1016/j.bmcl.2012.03.070
KETOCONAZOLE
Inhibition of CYP3A4 (unknown origin) by fluorescence assay
—
IC50 = 24.0 nM
Reference
Title : Discovery of a non-estrogenic irreversible inhibitor of 17β-hydroxysteroid dehydrogenase type 1 from 3-substituted-16β-(m-carbamoylbenzyl)-estradiol derivatives. Authors : Maltais R, Ayan D, Trottier A, Barbeau X, Lagüe P, Bouchard JE, Poirier D. Publication Year :
2014 Journal : J. Med. Chem. Abstract : 17β-Hydroxysteroid dehydrogenase type 1 (17β-HSD1) is thought to play a pivotal role in the progression of estrogen-sensitive breast cancer by transforming estrone (E1) into estradiol (E2). We designed three successive series of E2-derivatives at position C3 of the potent inhibitor 16β-(m-carbamoylbenzyl)-E2 to remove its unwanted estrogenic activity. We report the chemical synthesis and characterization of 20 new E2-derivatives, their evaluation as 17β-HSD1 inhibitors, and their proliferative (estrogenic) activity on estrogen-sensitive cells. The structure-activity relationship study provided a new potent and steroidal nonestrogenic inhibitor of 17β-HSD1 named 3-{[(16β,17β)-3-(2-bromoethyl)-17-hydroxyestra-1(10),2,4-trien-16-yl]methyl}benzamide (23b). In fact, this compound inhibited the transformation of E1 into E2 by 17β-HSD1 in T-47D cells (IC50 = 83 nM), did not inhibit 17β-HSD2, 17β-HSD7, 17β-HSD12, and CYP3A4, and did not stimulate the proliferation of estrogen-sensitive MCF-7 cells. We also discussed the results of kinetic and molecular modeling (docking) experiments, suggesting that compound 23b is a competitive and irreversible inhibitor of 17β-HSD1. DOI : 10.1021/jm401639v
KETOCONAZOLE
Inhibition of recombinant CYP3A4 (unknown origin) using 7-benzyloxy-4-trifluoromethylcoumarin as substrate
—
IC50 = 280.0 nM
Reference
Title : Investigation of a novel series of 2-hydroxyisoquinoline-1,3(2H,4H)-diones as human immunodeficiency virus type 1 integrase inhibitors. Authors : Suchaud V, Bailly F, Lion C, Calmels C, Andréola ML, Christ F, Debyser Z, Cotelle P. Publication Year :
2014 Journal : J. Med. Chem. Abstract : We report herein further insight into the biological activities displayed by a series of 2-hydroxyisoquinoline-1,3(2H,4H)-diones (HIDs). Substitution of the N-hydroxyimide two-metal binding pharmacophore at position 4 by carboxamido side chains was previously shown by us to be fruitful for this scaffold, since strong human immunodeficiency virus type 1 integrase (HIV-1 IN) inhibitors in the low nanomolar range associated with low micromolar anti-HIV activities were obtained. We investigated the influence of substitution at position 7 on biological activity. Introduction of electron-withdrawing functional groups such as the nitro moiety at position 7 led to a noticeable improvement of antiviral activity, down to low nanomolar anti-HIV potencies, with advantageous therapeutic indexes going close to those of the clinically used raltegravir and retained potencies against a panel of IN mutants. DOI : 10.1021/jm500109z
KETOCONAZOLE
Inhibition of CYP3A4 (unknown origin) using AMMC by fluorescence assay
—
IC50 = 25.0 nM
Reference
Title : Chemical synthesis, cytotoxicity, selectivity and bioavailability of 5α-androstane-3α,17β-diol derivatives. Authors : Ayan D, Maltais R, Hospital A, Poirier D. Publication Year :
2014 Journal : Bioorg. Med. Chem. Abstract : Aminosteroid derivatives represent a new family of compounds with promising antiproliferative activity over different cancer cell lines. Among all the aminosteroid derivatives synthesised in our laboratory, we have identified E-37P as one of the more potent when tested in vitro. Unfortunately, the pharmacokinetic properties of E-37P decrease its effectiveness when tested in vivo. To improve the bioavailability and increase the efficiency of aminosteroid E-37P, two series of analog compounds were synthesised by classic chemical synthesis, they were then characterized, and the concentration that inhibits 50% of cell proliferation (IC50) was determined on different cell lines. RM-133, a 5α-androstane-3α,17β-diol derivative with a quinoline nucleus at the end of the piperazine-proline side-chain at position 2β and an ethinyl at position 17α, showed very good antiproliferative activity among the five cancer cell lines studied (IC50=0.1, 0.1, 0.1, 2.0 and 1.1 μM for HL-60, MCF-7, T-47D, LNCaP and WEHI-3, respectively). Moreover, the plasmatic concentration of RM-133 at 3h, when injected subcutaneously in rats, was 2.3-fold higher than that of E-37P (151 vs 64.8 ng/mL). Furthermore, RM-133 weakly inhibited the two representative liver enzymes, CYP3A4 and CYP2D6, indicating a very low risk of drug-drug interactions. The cytotoxicity of RM-133 against normal cells was tested on peripheral blood lymphocytes (PBL) obtained from different donors and previously activated with phytohemagglutinin-L. PBL responded differently to treatment with RM-133, we observed a stimulation of cell proliferation and/or cytotoxicity in a dose-dependent manner. Based on these results, additional studies are currently underway to evaluate the selectivity of our lead compound against normal cell lines in a more detailed fashion. DOI : 10.1016/j.bmc.2014.09.026
KETOCONAZOLE
Inhibition of human recombinant CYP3A4 expressed in insect cell microsomes preincubated for 10 mins by fluorescence assay
—
IC50 = 200.0 nM
Reference
Title : Investigating the binding interactions of the anti-Alzheimer's drug donepezil with CYP3A4 and P-glycoprotein. Authors : McEneny-King A, Edginton AN, Rao PP. Publication Year :
2015 Journal : Bioorg. Med. Chem. Lett. Abstract : The anti-Alzheimer's agent donepezil is known to bind to the hepatic enzyme CYP3A4, but its relationship with the efflux transporter P-glycoprotein (P-gp) is not as well elucidated. We conducted in vitro inhibition studies of donepezil using human recombinant CYP3A4 and P-gp. These studies show that donepezil is a weak inhibitor of CYP3A4 (IC50=54.68±1.00μM) whereas the reference agent ketoconazole exhibited potent inhibition (CYP3A4 IC50=0.20±0.01μM). P-gp inhibition studies indicate that donepezil exhibits better inhibition relative to CYP3A4 (P-gp EC50=34.85±4.63μM) although it was less potent compared to ketoconazole (P-gp EC50=9.74±1.23μM). At higher concentrations, donepezil exhibited significant inhibition of CYP3A4 (69%, 84% and 87% inhibition at 100, 250 and 500μM, respectively). This indicates its potential to cause drug-drug interactions with other CYP3A4 substrates upon co-administration; however, this scenario is unlikely in vivo due to the low therapeutic concentrations of donepezil. Similarly, donepezil co-administration with P-gp substrates or inhibitors is unlikely to result in beneficial or adverse drug interactions. The molecular docking studies show that the 5,6-dimethoxyindan-1-one moiety of donepezil was oriented closer to the heme center in CYP3A4 whereas in the P-gp binding site, the protonated benzylpiperidine pharmacophore of donepezil played a major role in its binding ability. Energy parameters indicate that donepezil complex with both CYP3A4 and P-gp was less stable (CDOCKER energies=-15.05 and -4.91kcal/mol, respectively) compared to the ketoconazole-CYP3A4 and P-gp complex (CDOCKER energies=-41.89 and -20.03kcal/mol, respectively). DOI : 10.1016/j.bmcl.2014.11.046
KETOCONAZOLE
Inhibition of CYP3A4 in human liver microsome
Liver
IC50 < 62.0 nM
Reference
Title : Discovery of novel pyrimidine and malonamide derivatives as TGR5 agonists. Authors : Park EJ, Ahn YG, Jung SH, Bang HJ, Kim M, Hong DJ, Kim J, Suh KH, Kim YJ, Kim D, Kim EY, Lee K, Min KH. Publication Year :
2014 Journal : Bioorg. Med. Chem. Lett. Abstract : Takeda G-protein-coupled receptor 5 (TGR5) is a promising molecular target for metabolic diseases. A series of 4-(2,5-dichlorophenoxy)pyrimidine and cyclopropylmalonamide derivatives were synthesized as potent agonists of TGR5 based on a bioisosteric replacement strategy. Several compounds exhibited improved potency, compared to a reference compound with a pyridine scaffold. The pharmacokinetic profile of the representative compound 18 was considered moderate. DOI : 10.1016/j.bmcl.2014.07.026
KETOCONAZOLE
Inhibition of human recombinant CYP3A4 assessed as effect on conversion of beetle D-luciferin derivative into D-luciferin by firefly luciferase based luminescence assay
—
IC50 = 140.0 nM
Reference
Title : Design, synthesis and biological evaluation of new hybrid anticonvulsants derived from N-benzyl-2-(2,5-dioxopyrrolidin-1-yl)propanamide and 2-(2,5-dioxopyrrolidin-1-yl)butanamide derivatives. Authors : Kamiński K, Rapacz A, Łuszczki JJ, Latacz G, Obniska J, Kieć-Kononowicz K, Filipek B. Publication Year :
2015 Journal : Bioorg. Med. Chem. Abstract : The purpose of this study was to synthesize the library of 33 new N-benzyl-2-(2,5-dioxopyrrolidin-1-yl)propanamides, 2-(3-methyl-2,5-dioxopyrrolidin-1-yl)propanamides, and 2-(2,5-dioxopyrrolidin-1-yl)butanamides as potential new hybrid anticonvulsant agents. These hybrid molecules join the chemical fragments of well-known antiepileptic drugs (AEDs) such as ethosuximide, levetiracetam, and lacosamide. The coupling reaction of the 2-(2,5-dioxopyrrolidin-1-yl)propanoic acid, 2-(3-methyl-2,5-dioxopyrrolidin-1-yl)propanoic acid, or 2-(2,5-dioxopyrrolidin-1-yl)butanoic acid with the appropriately substituted benzylamines in the presence of the coupling reagent, N,N-carbonyldiimidazole (CDI) generated the final compounds 4-36. Spectral data acquired via (1)H NMR, (13)C NMR, and LC-MS confirmed the chemical structures of the newly prepared compounds. The initial anticonvulsant screening was performed in mice intraperitoneally (ip), using the maximal electroshock seizure (MES) and subcutaneous pentylenetetrazole (scPTZ) seizure tests. The rotarod test determined the acute neurological toxicity (NT). The results of preliminary pharmacological screening revealed that 25 compounds showed protection in half or more of the animals tested in the MES and/or scPTZ seizure models at the fixed dose of 100mg/kg. The broad spectra of activity across the preclinical seizure models displayed compounds 4, 7, 8, 13, 15-18, 24, and 26. The quantitative pharmacological studies in mice demonstrated the highest protection for compounds 4 (ED50 MES=67.65 mg/kg, ED50scPTZ=42.83 mg/kg); 8 (ED50 MES=54.90 mg/kg, ED50scPTZ=50.29 mg/kg); and 20 (ED50scPTZ=47.39 mg/kg). These compounds were distinctly more potent and provided better safety profiles in the rotarod test compared to valproic acid or ethosuximide, which were used as model AEDs. Compound 8 underwent only a slight metabolic change by the human liver microsomes (HLMs), and also did not affect the activity of human cytochrome P450 isoform, CYP3A4, in the in vitro assays. DOI : 10.1016/j.bmc.2015.03.038
KETOCONAZOLE
Inhibition of CYP3A4 in human liver microsomes using midazolam as substrate after 8 mins by LC-MS/MS analysis
Liver
IC50 < 100.0 nM
Reference
Title : Identifying a selective substrate and inhibitor pair for the evaluation of CYP2J2 activity. Authors : Lee CA, Jones JP, Katayama J, Kaspera R, Jiang Y, Freiwald S, Smith E, Walker GS, Totah RA. Publication Year :
2012 Journal : Drug Metab. Dispos. Abstract : CYP2J2, an arachidonic acid epoxygenase, is recognized for its role in the first-pass metabolism of astemizole and ebastine. To fully assess the role of CYP2J2 in drug metabolism, a selective substrate and potent specific chemical inhibitor are essential. In this study, we report amiodarone 4-hydoxylation as a specific CYP2J2-catalyzed reaction with no CYP3A4, or other drug-metabolizing enzyme, involvement. Amiodarone 4-hydroxylation enabled the determination of liver relative activity factor and intersystem extrapolation factor for CYP2J2. Amiodarone 4-hydroxylation correlated with astemizole O-demethylation but not with CYP2J2 protein content in a sample of human liver microsomes. To identify a specific CYP2J2 inhibitor, 138 drugs were screened using terfenadine and astemizole as probe substrates with recombinant CYP2J2. Forty-two drugs inhibited CYP2J2 activity by ≥50% at 30 μM, but inhibition was substrate-dependent. Of these, danazol was a potent inhibitor of both hydroxylation of terfenadine (IC(50) = 77 nM) and O-demethylation of astemizole (K(i) = 20 nM), and inhibition was mostly competitive. Danazol inhibited CYP2C9, CYP2C8, and CYP2D6 with IC(50) values of 1.44, 1.95, and 2.74 μM, respectively. Amiodarone or astemizole were included in a seven-probe cocktail for cytochrome P450 (P450) drug-interaction screening potential, and astemizole demonstrated a better profile because it did not appreciably interact with other P450 probes. Thus, danazol, amiodarone, and astemizole will facilitate the ability to determine the metabolic role of CYP2J2 in hepatic and extrahepatic tissues. DOI : 10.1124/dmd.111.043505
KETOCONAZOLE
Inhibition of CYP3A4 activity in human liver microsomes assessed as dibenzo fluuorescene oxidation up to 40 uM
Liver
IC50 = 9.8 nM
Reference
Title : Metabolites of PPI-2458, a selective, irreversible inhibitor of methionine aminopeptidase-2: structure determination and in vivo activity. Authors : Arico-Muendel CC, Belanger B, Benjamin D, Blanchette HS, Caiazzo TM, Centrella PA, DeLorey J, Doyle EG, Gradhand U, Griffin ST, Hill S, Labenski MT, Morgan BA, O'Donovan G, Prasad K, Skinner S, Taghizadeh N, Thompson CD, Wakefield J, Westlin W, White KF. Publication Year :
2013 Journal : Drug Metab. Dispos. Abstract : The natural product fumagillin exhibits potent antiproliferative and antiangiogenic properties. The semisynthetic analog PPI-2458, [(3R,4S,5S,6R)-5-methoxy-4-[(2R,3R)-2-methyl-3-(3-methylbut-2-enyl)oxiran-2-yl]-1-oxaspiro[2.5]octan-6-yl] N-[(2R)-1-amino-3-methyl-1-oxobutan-2-yl]carbamate, demonstrates rapid inactivation of its molecular target, methionine aminopeptidase-2 (MetAP2), and good efficacy in several rodent models of cancer and inflammation with oral dosing despite low apparent oral bioavailability. To probe the basis of its in vivo efficacy, the metabolism of PPI-2458 was studied in detail. Reaction phenotyping identified CYP3A4/5 as the major source of metabolism in humans. Six metabolites were isolated from liver microsomes and characterized by mass spectrometry and nuclear resonance spectroscopy, and their structures were confirmed by chemical synthesis. The synthetic metabolites showed correlated inhibition of MetAP2 enzymatic activity and vascular endothelial cell growth. In an ex vivo experiment, MetAP2 inhibition in white blood cells, thymus, and lymph nodes in rats after single dosing with PPI-2458 and the isolated metabolites was found to correlate with the in vitro activity of the individual species. In a phase 1 clinical study, PPI-2458 was administered to patients with non-Hodgkin lymphoma. At 15 mg administered orally every other day, MetAP2 in whole blood was 80% inactivated for up to 48 hours, although the exposure of the parent compound was only ∼10% that of the summed cytochrome P450 metabolites. Taken together, the data confirm the participation of active metabolites in the in vivo efficacy of PPI-2458. The structures define a metabolic pathway for PPI-2458 that is distinct from that of TNP-470 ([(3R,4S,5S,6R)-5-methoxy-4-[(2R,3R)-2-methyl-3-(3-methylbut-2-enyl)oxiran-2-yl]-1-oxaspiro[2.5]octan-6-yl] N-(2-chloroacetyl)carbamate). The high level of MetAP2 inhibition achieved in vivo supports the value of fumagillin-derived therapeutics for angiogenic diseases. DOI : 10.1124/dmd.112.048355
KETOCONAZOLE
Inhibition of CYP3A4 activity in human liver microsomes using midazolam as a substrate by LC/MS analysis
Liver
Ki = 7.0 nM
Reference
Title : Metabolites of PPI-2458, a selective, irreversible inhibitor of methionine aminopeptidase-2: structure determination and in vivo activity. Authors : Arico-Muendel CC, Belanger B, Benjamin D, Blanchette HS, Caiazzo TM, Centrella PA, DeLorey J, Doyle EG, Gradhand U, Griffin ST, Hill S, Labenski MT, Morgan BA, O'Donovan G, Prasad K, Skinner S, Taghizadeh N, Thompson CD, Wakefield J, Westlin W, White KF. Publication Year :
2013 Journal : Drug Metab. Dispos. Abstract : The natural product fumagillin exhibits potent antiproliferative and antiangiogenic properties. The semisynthetic analog PPI-2458, [(3R,4S,5S,6R)-5-methoxy-4-[(2R,3R)-2-methyl-3-(3-methylbut-2-enyl)oxiran-2-yl]-1-oxaspiro[2.5]octan-6-yl] N-[(2R)-1-amino-3-methyl-1-oxobutan-2-yl]carbamate, demonstrates rapid inactivation of its molecular target, methionine aminopeptidase-2 (MetAP2), and good efficacy in several rodent models of cancer and inflammation with oral dosing despite low apparent oral bioavailability. To probe the basis of its in vivo efficacy, the metabolism of PPI-2458 was studied in detail. Reaction phenotyping identified CYP3A4/5 as the major source of metabolism in humans. Six metabolites were isolated from liver microsomes and characterized by mass spectrometry and nuclear resonance spectroscopy, and their structures were confirmed by chemical synthesis. The synthetic metabolites showed correlated inhibition of MetAP2 enzymatic activity and vascular endothelial cell growth. In an ex vivo experiment, MetAP2 inhibition in white blood cells, thymus, and lymph nodes in rats after single dosing with PPI-2458 and the isolated metabolites was found to correlate with the in vitro activity of the individual species. In a phase 1 clinical study, PPI-2458 was administered to patients with non-Hodgkin lymphoma. At 15 mg administered orally every other day, MetAP2 in whole blood was 80% inactivated for up to 48 hours, although the exposure of the parent compound was only ∼10% that of the summed cytochrome P450 metabolites. Taken together, the data confirm the participation of active metabolites in the in vivo efficacy of PPI-2458. The structures define a metabolic pathway for PPI-2458 that is distinct from that of TNP-470 ([(3R,4S,5S,6R)-5-methoxy-4-[(2R,3R)-2-methyl-3-(3-methylbut-2-enyl)oxiran-2-yl]-1-oxaspiro[2.5]octan-6-yl] N-(2-chloroacetyl)carbamate). The high level of MetAP2 inhibition achieved in vivo supports the value of fumagillin-derived therapeutics for angiogenic diseases. DOI : 10.1124/dmd.112.048355
KETOCONAZOLE
Inhibition of CYP3A4 activity in human liver microsomes using nifedipine as a substrate by LC/MS analysis
Liver
Ki = 11.0 nM
Reference
Title : Metabolites of PPI-2458, a selective, irreversible inhibitor of methionine aminopeptidase-2: structure determination and in vivo activity. Authors : Arico-Muendel CC, Belanger B, Benjamin D, Blanchette HS, Caiazzo TM, Centrella PA, DeLorey J, Doyle EG, Gradhand U, Griffin ST, Hill S, Labenski MT, Morgan BA, O'Donovan G, Prasad K, Skinner S, Taghizadeh N, Thompson CD, Wakefield J, Westlin W, White KF. Publication Year :
2013 Journal : Drug Metab. Dispos. Abstract : The natural product fumagillin exhibits potent antiproliferative and antiangiogenic properties. The semisynthetic analog PPI-2458, [(3R,4S,5S,6R)-5-methoxy-4-[(2R,3R)-2-methyl-3-(3-methylbut-2-enyl)oxiran-2-yl]-1-oxaspiro[2.5]octan-6-yl] N-[(2R)-1-amino-3-methyl-1-oxobutan-2-yl]carbamate, demonstrates rapid inactivation of its molecular target, methionine aminopeptidase-2 (MetAP2), and good efficacy in several rodent models of cancer and inflammation with oral dosing despite low apparent oral bioavailability. To probe the basis of its in vivo efficacy, the metabolism of PPI-2458 was studied in detail. Reaction phenotyping identified CYP3A4/5 as the major source of metabolism in humans. Six metabolites were isolated from liver microsomes and characterized by mass spectrometry and nuclear resonance spectroscopy, and their structures were confirmed by chemical synthesis. The synthetic metabolites showed correlated inhibition of MetAP2 enzymatic activity and vascular endothelial cell growth. In an ex vivo experiment, MetAP2 inhibition in white blood cells, thymus, and lymph nodes in rats after single dosing with PPI-2458 and the isolated metabolites was found to correlate with the in vitro activity of the individual species. In a phase 1 clinical study, PPI-2458 was administered to patients with non-Hodgkin lymphoma. At 15 mg administered orally every other day, MetAP2 in whole blood was 80% inactivated for up to 48 hours, although the exposure of the parent compound was only ∼10% that of the summed cytochrome P450 metabolites. Taken together, the data confirm the participation of active metabolites in the in vivo efficacy of PPI-2458. The structures define a metabolic pathway for PPI-2458 that is distinct from that of TNP-470 ([(3R,4S,5S,6R)-5-methoxy-4-[(2R,3R)-2-methyl-3-(3-methylbut-2-enyl)oxiran-2-yl]-1-oxaspiro[2.5]octan-6-yl] N-(2-chloroacetyl)carbamate). The high level of MetAP2 inhibition achieved in vivo supports the value of fumagillin-derived therapeutics for angiogenic diseases. DOI : 10.1124/dmd.112.048355
KETOCONAZOLE
Inhibition of CYP3A4 activity in human liver microsomes using testosterone as a substrate by LC/MS analysis
Liver
Ki = 49.0 nM
Reference
Title : Metabolites of PPI-2458, a selective, irreversible inhibitor of methionine aminopeptidase-2: structure determination and in vivo activity. Authors : Arico-Muendel CC, Belanger B, Benjamin D, Blanchette HS, Caiazzo TM, Centrella PA, DeLorey J, Doyle EG, Gradhand U, Griffin ST, Hill S, Labenski MT, Morgan BA, O'Donovan G, Prasad K, Skinner S, Taghizadeh N, Thompson CD, Wakefield J, Westlin W, White KF. Publication Year :
2013 Journal : Drug Metab. Dispos. Abstract : The natural product fumagillin exhibits potent antiproliferative and antiangiogenic properties. The semisynthetic analog PPI-2458, [(3R,4S,5S,6R)-5-methoxy-4-[(2R,3R)-2-methyl-3-(3-methylbut-2-enyl)oxiran-2-yl]-1-oxaspiro[2.5]octan-6-yl] N-[(2R)-1-amino-3-methyl-1-oxobutan-2-yl]carbamate, demonstrates rapid inactivation of its molecular target, methionine aminopeptidase-2 (MetAP2), and good efficacy in several rodent models of cancer and inflammation with oral dosing despite low apparent oral bioavailability. To probe the basis of its in vivo efficacy, the metabolism of PPI-2458 was studied in detail. Reaction phenotyping identified CYP3A4/5 as the major source of metabolism in humans. Six metabolites were isolated from liver microsomes and characterized by mass spectrometry and nuclear resonance spectroscopy, and their structures were confirmed by chemical synthesis. The synthetic metabolites showed correlated inhibition of MetAP2 enzymatic activity and vascular endothelial cell growth. In an ex vivo experiment, MetAP2 inhibition in white blood cells, thymus, and lymph nodes in rats after single dosing with PPI-2458 and the isolated metabolites was found to correlate with the in vitro activity of the individual species. In a phase 1 clinical study, PPI-2458 was administered to patients with non-Hodgkin lymphoma. At 15 mg administered orally every other day, MetAP2 in whole blood was 80% inactivated for up to 48 hours, although the exposure of the parent compound was only ∼10% that of the summed cytochrome P450 metabolites. Taken together, the data confirm the participation of active metabolites in the in vivo efficacy of PPI-2458. The structures define a metabolic pathway for PPI-2458 that is distinct from that of TNP-470 ([(3R,4S,5S,6R)-5-methoxy-4-[(2R,3R)-2-methyl-3-(3-methylbut-2-enyl)oxiran-2-yl]-1-oxaspiro[2.5]octan-6-yl] N-(2-chloroacetyl)carbamate). The high level of MetAP2 inhibition achieved in vivo supports the value of fumagillin-derived therapeutics for angiogenic diseases. DOI : 10.1124/dmd.112.048355
KETOCONAZOLE
Inhibition of human recombinant CYP3A4 assessed as remaining enzyme activity by measuring conversion of luciferin-PPXE into D-luciferin by luminescence CYP3A4 P450-Glo assay relative to untreated control
—
IC50 = 140.0 nM
Reference
Title : Design, synthesis, and anticonvulsant activity of new hybrid compounds derived from 2-(2,5-dioxopyrrolidin-1-yl)propanamides and 2-(2,5-dioxopyrrolidin-1-yl)butanamides. Authors : Kamiński K, Zagaja M, Łuszczki JJ, Rapacz A, Andres-Mach M, Latacz G, Kieć-Kononowicz K. Publication Year :
2015 Journal : J. Med. Chem. Abstract : The library of 27 new 1-(4-phenylpiperazin-1-yl)- or 1-(morpholin-4-yl)-(2,5-dioxopyrrolidin-1-yl)propanamides and (2,5-dioxopyrrolidin-1-yl)butanamides as potential new hybrid anticonvulsant agents was synthesized. These hybrid molecules join the chemical fragments of well-known antiepileptic drugs (AEDs) such as ethosuximide, levetiracetam, and lacosamide. Compounds 5, 10, 11, and 24 displayed the broad spectra of activity across the preclinical seizure models, namely, the maximal electroshock (MES) test, the subcutaneous pentylenetetrazole (scPTZ) test, and the six-hertz (6 Hz) model of pharmacoresistant limbic seizures. The highest protection was demonstrated by 11 (ED50 MES = 88.4 mg/kg, ED50 scPTZ = 59.9 mg/kg, ED50 6 Hz = 21.0 mg/kg). This molecule did not impair the motor coordination of animals in the chimney test even at high doses (TD50 > 1500 mg/kg), yielding superb protective indexes (PI MES > 16.97, PI PTZ > 25.04, PI 6 Hz > 71.43). As a result, 11 displayed distinctly better safety profile than clinically relevant AEDs ethosuximide, lacosamide, or valproic acid. DOI : 10.1021/acs.jmedchem.5b00578
KETOCONAZOLE
Inhibition of CYP3A4 (unknown origin)
—
IC50 = 180.0 nM
Reference
Title : 4-Fluoro-3',4',5'-trimethoxychalcone as a new anti-invasive agent. From discovery to initial validation in an in vivo metastasis model. Authors : Roman BI, De Ryck T, Patronov A, Slavov SH, Vanhoecke BW, Katritzky AR, Bracke ME, Stevens CV. Publication Year :
2015 Journal : Eur. J. Med. Chem. Abstract : Invasion and metastasis are responsible for 90% of cancer-related mortality. Herein, we report on our quest for novel, clinically relevant inhibitors of local invasion, based on a broad screen of natural products in a phenotypic assay. Starting from micromolar chalcone hits, a predictive QSAR model for diaryl propenones was developed, and synthetic analogues with a 100-fold increase in potency were obtained. Two nanomolar hits underwent efficacy validation and eADMET profiling; one compound was shown to increase the survival time in an artificial metastasis model in nude mice. Although the molecular mechanism(s) by which these substances mediate efficacy remain(s) unrevealed, we were able to eliminate the major targets commonly associated with antineoplastic chalcones. DOI : 10.1016/j.ejmech.2015.06.029
KETOCONAZOLE
Inhibition of human recombinant CYP3A4 assessed as conversion of luciferin-PPXE to D-luciferin by luminescence Glo assay
—
IC50 = 140.0 nM
Reference
Title : (2-Arylethenyl)-1,3,5-triazin-2-amines as a novel histamine H4 receptor ligands. Authors : Kamińska K, Ziemba J, Ner J, Schwed JS, Łażewska D, Więcek M, Karcz T, Olejarz A, Latacz G, Kuder K, Kottke T, Zygmunt M, Sapa J, Karolak-Wojciechowska J, Stark H, Kieć-Kononowicz K. Publication Year :
2015 Journal : Eur. J. Med. Chem. Abstract : Within the constantly growing number of histamine H4 (H4R) receptor ligands there is a large group of azine derivatives. A series of novel compounds in the group of 4-methylpiperazine-1,3,5-triazine-2-amines were designed and obtained. Considered structures were modified at the triazine 6-position by introduction of variously substituted arylethenyl moieties. Their affinities to histamine H4 receptors were evaluated in radioligand binding assays with use of Sf9 cells, transiently expressing human H4R. Pharmacological studies results allowed to identify 4-[(E)-2-(3-chlorophenyl)ethenyl]-6-(4-methylpiperazin-1-yl)-1,3,5-triazin-2-amine (Ki = 253 nM) as the most potent compound in the present series. DOI : 10.1016/j.ejmech.2015.08.014
KETOCONAZOLE
Inhibition of CYP3A4 (unknown origin) using luciferin tagged substrate preincubated for 10 mins before substrate addition
—
IC50 = 60.0 nM
Reference
Title : Discovery of potent and selective cytotoxic activity of new quinazoline-ureas against TMZ-resistant glioblastoma multiforme (GBM). Authors : Elkamhawy A, Viswanath AN, Pae AN, Kim HY, Heo JC, Park WK, Lee CO, Yang H, Kim KH, Nam DH, Seol HJ, Cho H, Roh EJ. Publication Year :
2015 Journal : Eur. J. Med. Chem. Abstract : Herein, we report new quinazoline-urea based compounds with potent cytotoxic activities against TMZ-resistant glioblastoma multiforme (GBM) cells. Low micromolar IC₅₀ values were exhibited over a panel of three primary GBM patient-derived cell cultures belonging to proneural (GBM-1), mesenchymal (GBM-2), and classical (GBM-3) subtypes. Eight compounds showed excellent selectivity indices for GBM cells comparing to a normal astrocyte cell line. In JC-1 assay, analogues 11, 12, 20, 22, and 24 exerted promising rates of mPTP opening induction towards proneural GBM subtype. Compounds 11, 20, and 24 bound to the translocator protein 18 kDa (TSPO) in submicromolar range using [(3)H] PK-11195 binding affinity assay. A homology model was built and docked models of 11, 12, 20, 22 and 24 were generated for describing their plausible binding modes in TSPO. In 3D clonogenic assay, compound 20 manifested potent tumoricidal effects on TMZ-resistant GBM cells even at submicromolar concentrations. In addition, CYP450 and hERG assays presented a safe toxicity profile of 20. Taken as a whole, this report presents compound 20 as a potent, selective and safe GBM cytotoxic agent which constitutes a promising direction against TMZ-resistant GBM. DOI : 10.1016/j.ejmech.2015.08.001
KETOCONAZOLE
Inhibition of recombinant human CYP3A4 by luminescence -based microplate reader assay
—
IC50 = 140.0 nM
Reference
Title : Rational design in search for 5-phenylhydantoin selective 5-HT7R antagonists. Molecular modeling, synthesis and biological evaluation. Authors : Kucwaj-Brysz K, Warszycki D, Podlewska S, Witek J, Witek K, González Izquierdo A, Satała G, Loza MI, Lubelska A, Latacz G, Bojarski AJ, Castro M, Kieć-Kononowicz K, Handzlik J. Publication Year :
2016 Journal : Eur. J. Med. Chem. Abstract : A series of novel arylpiperazine 5-(4-fluorophenyl)-5-methylhydantoins with 2-hydroxypropyl linker (2-15) was synthesized and evaluated on their affinity towards serotonin 5-HT7 receptor (5-HT7R) in comparison to other closely related GPCRs: serotonin 5-HT1A, and dopamine D2 receptors. The functional activity studied through the measurement of 5-HT7R-mediated cyclic AMP production in Human Embryonic Kidney 293 cells (HEK293) stably expressing human 5-HT7 proved their antagonistic properties. The lead structure was also examined in the preliminary metabolic stability study using human liver microsomes (HMLs). The process of selection of candidates for synthesis was supported by a special molecular modeling workflow including combinatorial library generation, docking, and machine learning-based assessment. Additionally, in silico predictions of selectivity over 5-HT1AR and D2R, as well as functional activity were carried out. The newly synthesized compounds were proved to possess a potent affinity for 5-HT7R, similar to that of the lead structure of 5-(4-fluorophenyl)-3-(3-(4-(2-methoxyphenyl)piperazin-1-yl)-2-hydroxypropyl)-5-methylimidazolidine-2,4-dione (1). For several derivatives, significant selectivity both over 5-HT1AR and D2R was found. DOI : 10.1016/j.ejmech.2016.02.024
KETOCONAZOLE
Inhibition of human recombinant CYP3A4 using BFC as substrate incubated for 30 mins by fluorimetry
—
IC50 = 280.0 nM
Reference
Title : 2-hydroxyisoquinoline-1,3(2H,4H)-diones (HIDs) as human immunodeficiency virus type 1 integrase inhibitors: Influence of the alkylcarboxamide substitution of position 4. Authors : Billamboz M, Suchaud V, Bailly F, Lion C, Andréola ML, Christ F, Debyser Z, Cotelle P. Publication Year :
2016 Journal : Eur. J. Med. Chem. Abstract : Herein, we report further insight into the biological activities displayed by the 2-hydroxyisoquinoline-1,3(2H,4H)-dione (HID) scaffold. Previous studies have evidenced the marked fruitful effect of substitution of this two-metal binding pharmacophore at position 4 by phenyl and benzyl carboxamido chains. Strong human immunodeficiency virus type 1 integrase (HIV-1 IN) inhibitors in the low nanomolar range with micromolar (even down to low nanomolar) anti-HIV activities were obtained. Keeping this essential 4-carboxamido function, we investigated the influence of the replacement of phenyl and benzyl groups by various alkyl chains. This study shows that the recurrent halogenobenzyl pharmacophore found in the INSTIs can be efficiently replaced by an n-alkyl group. With an optimal length of six carbons, we observed a biological profile and a high barrier to resistance equivalent to those of a previously reported hit compound bearing a 4-fluorobenzyl group. DOI : 10.1016/j.ejmech.2016.03.083
KETOCONAZOLE
Inhibition of CYP3A4 in human liver microsomes using midazolam as substrate after 4 mins by LC-MS analysis
Liver
IC50 = 4.0 nM
Reference
Title : Understanding Oxadiazolothiazinone Biological Properties: Negative Inotropic Activity versus Cytochrome P450-Mediated Metabolism. Authors : Carosati E, Cosimelli B, Ioan P, Severi E, Katneni K, Chiu FC, Saponara S, Fusi F, Frosini M, Matucci R, Micucci M, Chiarini A, Spinelli D, Budriesi R. Publication Year :
2016 Journal : J. Med. Chem. Abstract : We present a series of oxadiazolothiazinones, selective inotropic agents on isolated cardiac tissues, devoid of chronotropy and vasorelaxant activity. Functional and binding data for the precursor of the series (compound 1) let us hypothesize LTCC blocking activity and the existence of a recognition site specific for this scaffold. We synthesized and tested 22 new derivatives: introducing a para-methoxyphenyl at C-8 led to compound 12 (EC50 = 0.022 μM), twice as potent as its para-bromo analogue (1). For 10 analogues, we extended the characterization of the biological properties by including the assessment of metabolic stability in human liver microsomes and cytochrome P450 inhibition potential. We observed that the methoxy group led to active compounds with low metabolic stability and high CYP inhibition, whereas the protective effect of bromine resulted in enhanced metabolic stability and reduced CYP inhibition. Thus, we identified two para-bromo benzothiazino-analogues as candidates for further studies. DOI : 10.1021/acs.jmedchem.6b00030
KETOCONAZOLE
Inhibition of CYP3A4 in human liver microsomes using testosterone as substrate after 5 mins by LC-MS analysis
Liver
IC50 = 1.0 nM
Reference
Title : Understanding Oxadiazolothiazinone Biological Properties: Negative Inotropic Activity versus Cytochrome P450-Mediated Metabolism. Authors : Carosati E, Cosimelli B, Ioan P, Severi E, Katneni K, Chiu FC, Saponara S, Fusi F, Frosini M, Matucci R, Micucci M, Chiarini A, Spinelli D, Budriesi R. Publication Year :
2016 Journal : J. Med. Chem. Abstract : We present a series of oxadiazolothiazinones, selective inotropic agents on isolated cardiac tissues, devoid of chronotropy and vasorelaxant activity. Functional and binding data for the precursor of the series (compound 1) let us hypothesize LTCC blocking activity and the existence of a recognition site specific for this scaffold. We synthesized and tested 22 new derivatives: introducing a para-methoxyphenyl at C-8 led to compound 12 (EC50 = 0.022 μM), twice as potent as its para-bromo analogue (1). For 10 analogues, we extended the characterization of the biological properties by including the assessment of metabolic stability in human liver microsomes and cytochrome P450 inhibition potential. We observed that the methoxy group led to active compounds with low metabolic stability and high CYP inhibition, whereas the protective effect of bromine resulted in enhanced metabolic stability and reduced CYP inhibition. Thus, we identified two para-bromo benzothiazino-analogues as candidates for further studies. DOI : 10.1021/acs.jmedchem.6b00030
KETOCONAZOLE
Inhibition of recombinant human CYP3A4 by P450-Glo luminescence assay
—
IC50 = 240.0 nM
Reference
Title : 2-(3-Methoxyphenyl)quinazoline Derivatives: A New Class of Direct Constitutive Androstane Receptor (CAR) Agonists. Authors : Smutny T, Nova A, Drechslerová M, Carazo A, Hyrsova L, Hrušková ZR, Kuneš J, Pour M, Špulák M, Pavek P. Publication Year :
2016 Journal : J. Med. Chem. Abstract : Constitutive androstane receptor (CAR) is a key regulator of xenobiotic and endobiotic metabolism. Together with pregnane X (PXR) and aryl hydrocarbon (AHR) receptors, it is referred to as "xenobiotic receptor". The unique properties of human CAR, such as its high constitutive activity, both direct (ligand-binding domain-dependent) and indirect activation have hindered the discovery of direct selective human CAR ligands. Herein, we report a novel class of direct human CAR agonists in a group of 2-(3-methoxyphenyl)quinazoline derivatives. The compounds are even more potent activators of human CAR than is prototype 6-(4-chlorophenyl)imidazo[2,1-b][1,3]thiazole-5-carbaldehyde O-(3,4-dichlorobenzyl)oxime (CITCO). The three most potent ligands are at the same time extremely potent activators of the other xenobiotic or hormonal receptors, namely PXR, AHR, and vitamin D receptor, which regulate major xenobiotic-metabolizing enzymes and efflux transporters. Thus, the novel CAR ligands can be also considered as constituting the first class of potent pan-xenobiotic receptor ligands that can serve as potential antidotes boosting overall metabolic elimination of xenobiotic or toxic compounds. DOI : 10.1021/acs.jmedchem.5b01891
KETOCONAZOLE
Inhibition of CYP3A4 in human liver microsomes using midazolam as substrate assessed as reduction in 1-OH-midazolam formation by LC/MS/MS analysis
Liver
IC50 = 9.1 nM
Reference
Title : Identification of (R)-(2-Chloro-3-(trifluoromethyl)phenyl)(1-(5-fluoropyridin-2-yl)-4-methyl-6,7-dihydro-1H-imidazo[4,5-c]pyridin-5(4H)-yl)methanone (JNJ 54166060), a Small Molecule Antagonist of the P2X7 receptor. Authors : Swanson DM, Savall BM, Coe KJ, Schoetens F, Koudriakova T, Skaptason J, Wall J, Rech J, Deng X, De Angelis M, Everson A, Lord B, Wang Q, Ao H, Scott B, Sepassi K, Lovenberg TW, Carruthers NI, Bhattacharya A, Letavic MA. Publication Year :
2016 Journal : J Med Chem Abstract : The synthesis and SAR of a series of 4,5,6,7-tetrahydro-imidazo[4,5-c]pyridine P2X7 antagonists are described. Addressing P2X7 affinity and liver microsomal stability issues encountered with this template afforded methyl substituted 4,5,6,7-tetrahydro-imidazo[4,5-c]pyridines ultimately leading to the identification of 1 (JNJ 54166060). 1 is a potent P2X7 antagonist with an ED50 = 2.3 mg/kg in rats, high oral bioavailability and low-moderate clearance in preclinical species, acceptable safety margins in rats, and a predicted human dose of 120 mg of QD. Additionally, 1 possesses a unique CYP profile and was found to be a regioselective inhibitor of midazolam CYP3A metabolism. DOI : 10.1021/acs.jmedchem.6b00989
KETOCONAZOLE
Inhibition of CYP3A4 in human liver microsomes using midazolam as substrate assessed as reduction in 4-OH-midazolam formation by LC/MS/MS analysis
Liver
IC50 = 7.4 nM
Reference
Title : Identification of (R)-(2-Chloro-3-(trifluoromethyl)phenyl)(1-(5-fluoropyridin-2-yl)-4-methyl-6,7-dihydro-1H-imidazo[4,5-c]pyridin-5(4H)-yl)methanone (JNJ 54166060), a Small Molecule Antagonist of the P2X7 receptor. Authors : Swanson DM, Savall BM, Coe KJ, Schoetens F, Koudriakova T, Skaptason J, Wall J, Rech J, Deng X, De Angelis M, Everson A, Lord B, Wang Q, Ao H, Scott B, Sepassi K, Lovenberg TW, Carruthers NI, Bhattacharya A, Letavic MA. Publication Year :
2016 Journal : J Med Chem Abstract : The synthesis and SAR of a series of 4,5,6,7-tetrahydro-imidazo[4,5-c]pyridine P2X7 antagonists are described. Addressing P2X7 affinity and liver microsomal stability issues encountered with this template afforded methyl substituted 4,5,6,7-tetrahydro-imidazo[4,5-c]pyridines ultimately leading to the identification of 1 (JNJ 54166060). 1 is a potent P2X7 antagonist with an ED50 = 2.3 mg/kg in rats, high oral bioavailability and low-moderate clearance in preclinical species, acceptable safety margins in rats, and a predicted human dose of 120 mg of QD. Additionally, 1 possesses a unique CYP profile and was found to be a regioselective inhibitor of midazolam CYP3A metabolism. DOI : 10.1021/acs.jmedchem.6b00989
KETOCONAZOLE
Inhibition of CYP3A4 in pooled human hepatic microsomes using testosterone substrate in presence of NADPH
Liver
IC50 = 28.5 nM
Reference
Title : Discovery and Optimization of 1-Phenoxy-2-aminoindanes as Potent, Selective, and Orally Bioavailable Inhibitors of the Na+/H+ Exchanger Type 3 (NHE3). Authors : Rackelmann N, Matter H, Englert H, Follmann M, Maier T, Weston J, Arndt P, Heyse W, Mertsch K, Wirth K, Bialy L. Publication Year :
2016 Journal : J Med Chem Abstract : The design, synthesis, and structure-activity relationship of 1-phenoxy-2-aminoindanes as inhibitors of the Na+/H+ exchanger type 3 (NHE3) are described based on a hit from high-throughput screening (HTS). The chemical optimization resulted in the discovery of potent, selective, and orally bioavailable NHE3 inhibitors with 13d as best compound, showing high in vitro permeability and lacking CYP2D6 inhibition as main optimization parameters. Aligning 1-phenoxy-2-aminoindanes onto the X-ray structure of 13d then provided 3D-QSAR models for NHE3 inhibition capturing guidelines for optimization. These models showed good correlation coefficients and allowed for activity estimation. In silico ADMET models for Caco-2 permeability and CYP2D6 inhibition were also successfully applied for this series. Moreover, docking into the CYP2D6 X-ray structure provided a reliable alignment for 3D-QSAR models. Finally 13d, renamed as SAR197, was characterized in vitro and by in vivo pharmacokinetic (PK) and pharmacological studies to unveil its potential for reduction of obstructive sleep apneas. DOI : 10.1021/acs.jmedchem.6b00624
KETOCONAZOLE
Inhibition of human Cyp3A4 using testosterone as substrate by LC-MS/MS analysis
—
IC50 = 133.0 nM
Reference
Title : Design, synthesis, and evaluation of (2S,4R)-Ketoconazole sulfonamide analogs as potential treatments for Metabolic Syndrome. Authors : Blass BE, Iyer P, Abou-Gharbia M, Childers WE, Gordon JC, Ramanjulu M, Morton G, Arumugam P, Boruwa J, Ellingboe J, Mitra S, Nimmareddy RR, Paliwal S, Rajasekhar J, Shivakumar S, Srivastava P, Tangirala RS, Venkataramanaiah K, Yanamandra M. Publication Year :
2016 Journal : Bioorg Med Chem Lett Abstract : Metabolic Syndrome, also referred to as 'Syndrome X' or 'Insulin Resistance Syndrome,' remains a major, unmet medical need despite over 30years of intense effort. Recent research suggests that there may be a causal link between this condition and abnormal glucocorticoid processing. Specifically, dysregulation of the hypothalamic-pituitary-adrenocortical (HPA) axis leads to increased systemic cortisol concentrations. Cushing' syndrome, a disorder that is also typified by a marked elevation in levels of cortisol, produces clinical symptomology that is similar to those observed in MetS, and they can be alleviated by decreasing circulating cortisol concentrations. As a result, it has been suggested that decreasing systemic cortisol concentration might have a positive impact on the progression of MetS. This could be accomplished through inhibition of enzymes in the cortisol synthetic pathway, 11β-hydroxylase (Cyp11B1), 17α-hydroxylase-C17,20-lyase (Cyp17), and 21-hydroxylase (Cyp21). We have identified a series of novel sulfonamide analogs of (2S,4R)-Ketoconazole that are potent inhibitors of these enzymes. In addition, selected members of this class of compounds have pharmacokinetic properties consistent with orally delivered drugs, making them well suited to further investigation as potential therapies for MetS. DOI : 10.1016/j.bmcl.2016.10.016
KETOCONAZOLE
Inhibition of human Cyp3A4 using testosterone as substrate by LC-MS/MS analysis
—
IC50 = 146.0 nM
Reference
Title : Design, synthesis, and evaluation of (2S,4R)-Ketoconazole sulfonamide analogs as potential treatments for Metabolic Syndrome. Authors : Blass BE, Iyer P, Abou-Gharbia M, Childers WE, Gordon JC, Ramanjulu M, Morton G, Arumugam P, Boruwa J, Ellingboe J, Mitra S, Nimmareddy RR, Paliwal S, Rajasekhar J, Shivakumar S, Srivastava P, Tangirala RS, Venkataramanaiah K, Yanamandra M. Publication Year :
2016 Journal : Bioorg Med Chem Lett Abstract : Metabolic Syndrome, also referred to as 'Syndrome X' or 'Insulin Resistance Syndrome,' remains a major, unmet medical need despite over 30years of intense effort. Recent research suggests that there may be a causal link between this condition and abnormal glucocorticoid processing. Specifically, dysregulation of the hypothalamic-pituitary-adrenocortical (HPA) axis leads to increased systemic cortisol concentrations. Cushing' syndrome, a disorder that is also typified by a marked elevation in levels of cortisol, produces clinical symptomology that is similar to those observed in MetS, and they can be alleviated by decreasing circulating cortisol concentrations. As a result, it has been suggested that decreasing systemic cortisol concentration might have a positive impact on the progression of MetS. This could be accomplished through inhibition of enzymes in the cortisol synthetic pathway, 11β-hydroxylase (Cyp11B1), 17α-hydroxylase-C17,20-lyase (Cyp17), and 21-hydroxylase (Cyp21). We have identified a series of novel sulfonamide analogs of (2S,4R)-Ketoconazole that are potent inhibitors of these enzymes. In addition, selected members of this class of compounds have pharmacokinetic properties consistent with orally delivered drugs, making them well suited to further investigation as potential therapies for MetS. DOI : 10.1016/j.bmcl.2016.10.016
KETOCONAZOLE
Inhibition of CYP3A4 in human liver microsomes assessed as reduction in nifedipine oxidation incubated for 10 mins
Liver
IC50 = 31.7 nM
Reference
Title : Discovery of biphenyl imidazole derivatives as potent antifungal agents: Design, synthesis, and structure-activity relationship studies. Authors : Zhao D, Zhao S, Zhao L, Zhang X, Wei P, Liu C, Hao C, Sun B, Su X, Cheng M. Publication Year :
2017 Journal : Bioorg Med Chem Abstract : Fungal infections have became a serious medical problem due to their high incidence and mortality. We describe the discovery and structure-activity relationships studies (SARs) of a series of novel biphenyl imidazole derivatives with excellent antifungal activities against Candida albicans and Cryptococcus neoformans. The most promising compounds 12f-g and 19a-b exhibited excellent activity with minimum inhibitory concentration (MIC) values in the range of 0.03125-2μg/mL. Preliminary mechanism studies showed that the potent antifungal activity of compound 12g stemed from inhibition of CYP51 in Candida albicans. Furthermore, compounds 12g and 19b exhibited low inhibition profiles for various human cytochrome P450 isoforms. The SARs and binding mode established in this study will be useful for further lead optimization. DOI : 10.1016/j.bmc.2016.11.051
KETOCONAZOLE
Inhibition of CYP3A4 using in human liver microsomes using midazolam as substrate after 5 to 15 mins
Liver
IC50 = 18.0 nM
Reference
Title : Discovery of GBT440, an Orally Bioavailable R-State Stabilizer of Sickle Cell Hemoglobin. Authors : Metcalf B, Chuang C, Dufu K, Patel MP, Silva-Garcia A, Johnson C, Lu Q, Partridge JR, Patskovska L, Patskovsky Y, Almo SC, Jacobson MP, Hua L, Xu Q, Gwaltney SL, Yee C, Harris J, Morgan BP, James J, Xu D, Hutchaleelaha A, Paulvannan K, Oksenberg D, Li Z. Publication Year :
2017 Journal : ACS Med Chem Lett Abstract : We report the discovery of a new potent allosteric effector of sickle cell hemoglobin, GBT440 (36), that increases the affinity of hemoglobin for oxygen and consequently inhibits its polymerization when subjected to hypoxic conditions. Unlike earlier allosteric activators that bind covalently to hemoglobin in a 2:1 stoichiometry, 36 binds with a 1:1 stoichiometry. Compound 36 is orally bioavailable and partitions highly and favorably into the red blood cell with a RBC/plasma ratio of ∼150. This partitioning onto the target protein is anticipated to allow therapeutic concentrations to be achieved in the red blood cell at low plasma concentrations. GBT440 (36) is in Phase 3 clinical trials for the treatment of sickle cell disease (NCT03036813). DOI : 10.1021/acsmedchemlett.6b00491
KETOCONAZOLE
Inhibition of CYP3A4 using in human liver microsomes using testosterone as substrate after 5 to 15 mins
Liver
IC50 = 15.0 nM
Reference
Title : Discovery of GBT440, an Orally Bioavailable R-State Stabilizer of Sickle Cell Hemoglobin. Authors : Metcalf B, Chuang C, Dufu K, Patel MP, Silva-Garcia A, Johnson C, Lu Q, Partridge JR, Patskovska L, Patskovsky Y, Almo SC, Jacobson MP, Hua L, Xu Q, Gwaltney SL, Yee C, Harris J, Morgan BP, James J, Xu D, Hutchaleelaha A, Paulvannan K, Oksenberg D, Li Z. Publication Year :
2017 Journal : ACS Med Chem Lett Abstract : We report the discovery of a new potent allosteric effector of sickle cell hemoglobin, GBT440 (36), that increases the affinity of hemoglobin for oxygen and consequently inhibits its polymerization when subjected to hypoxic conditions. Unlike earlier allosteric activators that bind covalently to hemoglobin in a 2:1 stoichiometry, 36 binds with a 1:1 stoichiometry. Compound 36 is orally bioavailable and partitions highly and favorably into the red blood cell with a RBC/plasma ratio of ∼150. This partitioning onto the target protein is anticipated to allow therapeutic concentrations to be achieved in the red blood cell at low plasma concentrations. GBT440 (36) is in Phase 3 clinical trials for the treatment of sickle cell disease (NCT03036813). DOI : 10.1021/acsmedchemlett.6b00491
KETOCONAZOLE
Inhibition of recombinant human CYP3A4 expressed in insect cells using DBOMF as substrate pretreated for 10 mins followed by substrate addition measured every minute for 15 mins in presence of NADP+ by fluorescence assay
—
IC50 = 200.0 nM
Reference
Title : Cytochrome P450 binding studies of novel tacrine derivatives: Predicting the risk of hepatotoxicity. Authors : McEneny-King A, Osman W, Edginton AN, Rao PPN. Publication Year :
2017 Journal : Bioorg Med Chem Lett Abstract : The 1,2,3,4-tetrahydroacridine derivative tacrine was the first drug approved to treat Alzheimer's disease (AD). It is known to act as a potent cholinesterase inhibitor. However, tacrine was removed from the market due to its hepatotoxicity concerns as it undergoes metabolism to toxic quinonemethide species through the cytochrome P450 enzyme CYP1A2. Despite these challenges, tacrine serves as a useful template in the development of novel multi-targeting anti-AD agents. In this regard, we sought to evaluate the risk of hepatotoxicity in a series of C9 substituted tacrine derivatives that exhibit cholinesterase inhibition properties. The hepatotoxic potential of tacrine derivatives was evaluated using recombinant cytochrome (CYP) P450 CYP1A2 and CYP3A4 enzymes. Molecular docking studies were conducted to predict their binding modes and potential risk of forming hepatotoxic metabolites. Tacrine derivatives compound 1 (N-(3,4-dimethoxybenzyl)-1,2,3,4-tetrahydroacridin-9-amine) and 2 (6-chloro-N-(3,4-dimethoxybenzyl)-1,2,3,4-tetrahydroacridin-9-amine) which possess a C9 3,4-dimethoxybenzylamino substituent exhibited weak binding to CYP1A2 enzyme (1, IC50=33.0µM; 2, IC50=8.5µM) compared to tacrine (CYP1A2 IC50=1.5µM). Modeling studies show that the presence of a bulky 3,4-dimethoxybenzylamino C9 substituent prevents the orientation of the 1,2,3,4-tetrahydroacridine ring close to the heme-iron center of CYP1A2 thereby reducing the risk of forming hepatotoxic species. DOI : 10.1016/j.bmcl.2017.04.006
KETOCONAZOLE
Inhibition of CYP3A4 (unknown origin) expressed in baculosomes
—
IC50 = 40.0 nM
Reference
Title : Comprehensive Synthesis of Amino Acid-Derived Thiazole Peptidomimetic Analogues to Understand the Enigmatic Drug/Substrate-Binding Site of P-Glycoprotein. Authors : Patel BA, Abel B, Barbuti AM, Velagapudi UK, Chen ZS, Ambudkar SV, Talele TT. Publication Year :
2018 Journal : J Med Chem Abstract : A novel set of 64 analogues based on our lead compound 1 was designed and synthesized with an initial objective of understanding the structural requirements of ligands binding to a highly perplexing substrate-binding site of P-glycoprotein (P-gp) and their effect on modulating the ATPase function of the efflux pump. Compound 1, a stimulator of P-gp ATPase activity, was transformed to ATPase inhibitory compounds 39, 53, and 109. The ATPase inhibition by these compounds was predominantly contributed by the presence of a cyclohexyl group in lieu of the 2-aminobenzophenone moiety of 1. The 4,4-difluorocyclohexyl analogues, 53 and 109, inhibited the photolabeling by [125I]-IAAP, with IC50 values of 0.1 and 0.76 μM, respectively. Selected compounds were shown to reverse paclitaxel resistance in HEK293 cells overexpressing P-gp and were selective toward P-gp over CYP3A4. Induced-fit docking highlighted a plausible binding pattern of inhibitory compounds in the putative-binding pocket of P-gp. The current study underscores the stringent requirement by P-gp to bind to chemically similar molecules. DOI : 10.1021/acs.jmedchem.7b01340
KETOCONAZOLE
Inhibition of CYP3A4 in human liver microsomes using midazolam as substrate preincubated for 10 mins followed by NADPH addition measured after 10 mins by LC/MS/MS analysis
Liver
IC50 = 35.9 nM
Reference
Title : Design, synthesis and evaluation of aromatic heterocyclic derivatives as potent antifungal agents. Authors : Zhao S, Zhang X, Wei P, Su X, Zhao L, Wu M, Hao C, Liu C, Zhao D, Cheng M. Publication Year :
2017 Journal : Eur J Med Chem Abstract : To further enhance the anti-Aspergillus efficacy of our previously discovered antifungal lead compounds (1), a series of aromatic heterocyclic derivatives were designed, synthesized and evaluated for in vitro antifungal activity. Many of the target compounds showed good inhibitory activity against Candida albicans and Cryptococcus neoformans. In particular, the isoxazole nuclei were more suited for improving the activity against Aspergillus spp. Among these compounds, 2-F substituted analogues 23g and 23h displayed the most remarkable in vitro activity against Candida spp., C. neoformans, A. fumigatus and fluconazole-resistant C.alb. strains, which is superior or comparable to the activity of the reference drugs fluconazole and voriconazole. Notably, the compounds 23g and 23h exhibited low inhibition profiles for various isoforms of human cytochrome P450 and excellent blood plasma stability. DOI : 10.1016/j.ejmech.2017.05.043
KETOCONAZOLE
Inhibition of CYP3A4 (unknown origin) by CYP3A4 P450-Glo assay
—
IC50 = 140.0 nM
Reference
Title : Novel naphthyloxy derivatives - Potent histamine H3 receptor ligands. Synthesis and pharmacological evaluation. Authors : Łażewska D, Kaleta M, Hagenow S, Mogilski S, Latacz G, Karcz T, Lubelska A, Honkisz E, Handzlik J, Reiner D, Satała G, Filipek B, Stark H, Kieć-Kononowicz K. Publication Year :
2018 Journal : Bioorg Med Chem Abstract : A series of 1- and 2-naphthyloxy derivatives were synthesized and evaluated for histamine H3 receptor affinity. Most compounds showed high affinities with Ki values below 100 nM. The most potent ligand, 1-(5-(naphthalen-1-yloxy)pentyl)azepane (11) displayed high affinity for the histamine H3 receptor with a Ki value of 21.9 nM. The antagonist behaviour of 11 was confirmed both in vitro in the cAMP assay (IC50 = 312 nM) and in vivo in the rat dipsogenia model (ED50 = 3.68 nM). Moreover, compound 11 showed positive effects on scopolamine induced-memory deficits in mice (at doses of 10 and 15 mg/kg) and an analgesic effect in the formalin test in mice with ED50 = 30.6 mg/kg (early phase) and ED50 = 20.8 mg/kg (late phase). Another interesting compound, 1-(5-(Naphthalen-1-yloxy)pentyl)piperidine (13; H3R Ki = 53.9 nM), was accepted for Anticonvulsant Screening Program at the National Institute of Neurological Disorders and Stroke/National Institute of Health (Rockville, USA). The screening was performed in the maximal electroshock seizure (MES), the subcutaneous pentylenetetrazole (scPTZ) and the 6-Hz psychomotor animal models of epilepsy. Neurologic deficit was evaluated by the rotarod test. Compound 13 inhibited convulsions induced by the MES with ED50 of 19.2 mg/kg (mice, i.p.), 17.8 (rats, i.p.), and 78.1 (rats, p.o.). Moreover, 13 displayed protection against the 6-Hz psychomotor seizures (32 mA) in mice (i.p.) with ED50 of 33.1 mg/kg and (44 mA) ED50 of 57.2 mg/kg. Furthermore, compounds 11 and 13 showed in vitro weak influence on viability of tested cell lines (normal HEK293, neuroblastoma IMR-32, hepatoma HEPG2), weak inhibition of CYP3A4 activity, and no mutagenicity. Thus, these compounds may be used as leads in a further search for histamine H3 receptor ligands with promising in vitro and in vivo activity. DOI : 10.1016/j.bmc.2018.04.023
KETOCONAZOLE
Inhibition of CYP3A4 in human liver microsomes assessed as midazolam 1'-hydroxylation after 4 to 40 mins in presence of NADPH by LCMS analysis
Liver
IC50 = 22.0 nM
Reference
Title : Synthesis and profiling of benzylmorpholine 1,2,4,5-tetraoxane analogue N205: Towards tetraoxane scaffolds with potential for single dose cure of malaria. Authors : O' Neill PM, Stocks PA, Sabbani S, Roberts NL, Amewu RK, Shore ER, Aljayyoussi G, Angulo-Barturén I, Belén M, Jiménez-Díaz, Bazaga SF, Martínez MS, Campo B, Sharma R, Charman SA, Ryan E, Chen G, Shackleford DM, Davies J, Nixon GL, Biagini GA, Ward SA. Publication Year :
2018 Journal : Bioorg Med Chem Abstract : A series of aryl carboxamide and benzylamino dispiro 1,2,4,5-tetraoxane analogues have been designed and synthesized in a short synthetic sequence from readily available starting materials. From this series of endoperoxides, molecules with in vitro IC50s versus Plasmodium falciparum (3D7) as low as 0.84 nM were identified. Based on an assessment of blood stability and in vitro microsomal stability, N205 (10a) was selected for rodent pharmacokinetic and in vivo antimalarial efficacy studies in the mouse Plasmodium berghei and Plasmodium falciparum Pf3D70087/N9 severe combined immunodeficiency (SCID) mouse models. The results indicate that the 4-benzylamino derivatives have excellent profiles with a representative of this series, N205, an excellent starting point for further lead optimization studies. DOI : 10.1016/j.bmc.2018.05.006
KETOCONAZOLE
Inhibition of CYP3A4 in human liver microsomes assessed as testosterone 6beta-hydroxylation after 4 to 40 mins in presence of NADPH by LCMS analysis
Liver
IC50 = 13.0 nM
Reference
Title : Synthesis and profiling of benzylmorpholine 1,2,4,5-tetraoxane analogue N205: Towards tetraoxane scaffolds with potential for single dose cure of malaria. Authors : O' Neill PM, Stocks PA, Sabbani S, Roberts NL, Amewu RK, Shore ER, Aljayyoussi G, Angulo-Barturén I, Belén M, Jiménez-Díaz, Bazaga SF, Martínez MS, Campo B, Sharma R, Charman SA, Ryan E, Chen G, Shackleford DM, Davies J, Nixon GL, Biagini GA, Ward SA. Publication Year :
2018 Journal : Bioorg Med Chem Abstract : A series of aryl carboxamide and benzylamino dispiro 1,2,4,5-tetraoxane analogues have been designed and synthesized in a short synthetic sequence from readily available starting materials. From this series of endoperoxides, molecules with in vitro IC50s versus Plasmodium falciparum (3D7) as low as 0.84 nM were identified. Based on an assessment of blood stability and in vitro microsomal stability, N205 (10a) was selected for rodent pharmacokinetic and in vivo antimalarial efficacy studies in the mouse Plasmodium berghei and Plasmodium falciparum Pf3D70087/N9 severe combined immunodeficiency (SCID) mouse models. The results indicate that the 4-benzylamino derivatives have excellent profiles with a representative of this series, N205, an excellent starting point for further lead optimization studies. DOI : 10.1016/j.bmc.2018.05.006
KETOCONAZOLE
Inhibition of recombinant human CYP3A4 using Luciferin-PPXE as substrate preincubated for 10 mins followed by NADPH addition measured after 15 mins by luminometric method
—
IC50 = 182.6 nM
Reference
Title : Antifungal activity, mode of action variability, and subcellular distribution of coumarin-based antifungal azoles. Authors : Elias R, Benhamou RI, Jaber QZ, Dorot O, Zada SL, Oved K, Pichinuk E, Fridman M. Publication Year :
2019 Journal : Eur J Med Chem Abstract : Azole antifungals inhibit the biosynthesis of ergosterol, the fungal equivalent of cholesterol in mammalian cells. Here we report an investigation of the activity of coumarin-substituted azole antifungals. Screening against a panel of Candida pathogens, including a mutant lacking CYP51, the target of antifungal azoles, revealed that this enzyme is inhibited by triazole-based antifungals, whereas imidazole-based derivatives have more than one mode of action. The imidazole-bearing antifungals more effectively reduced trailing growth associated with persistence and/or recurrence of fungal infections than triazole-based derivatives. The imidazole derivatives were more toxic to mammalian cells and more potently inhibited the activity of CYP3A4, which is one of the main causes of azole toxicity. Using live cell imaging, we showed that regardless of the type of azole ring fluorescent 7-diethylaminocoumarin-based azoles localized to the endoplasmic reticulum, the organelle that harbors CYP51. This study suggests that the coumarin is a promising scaffold for development of novel azole-based antifungals that effectively localize to the fungal cell endoplasmic reticulum. DOI : 10.1016/j.ejmech.2019.07.003
LAPATINIB
Inhibition of CYP3A4-mediated midazolam 1-hydroxylation in human liver microsomes at 50 uM
Liver
Inhibition = 45.0 %
Reference
Title : Metabolism-dependent inhibition of CYP3A4 by lapatinib: evidence for formation of a metabolic intermediate complex with a nitroso/oxime metabolite formed via a nitrone intermediate. Authors : Barbara JE, Kazmi F, Parkinson A, Buckley DB. Publication Year :
2013 Journal : Drug Metab. Dispos. Abstract : Metabolism-dependent inhibition (MDI) of cytochrome P450 (P450) enzymes has the potential to cause clinically relevant drug-drug interactions. In the case of several alkylamine drugs, MDI of P450 involves formation of a metabolite that binds quasi-irreversibly to the ferrous heme iron to form a metabolic intermediate (MI) complex. The specific metabolites coordinately bound to ferrous iron and the pathways leading to MI complex formation are the subject of debate. We describe an approach combining heme iron oxidation with potassium ferricyanide and metabolite profiling to probe the mechanism of MI complex-based CYP3A4 inactivation by the secondary alkylamine drug lapatinib. Ten metabolites formed from lapatinib by CYP3A4-mediated heteroatom dealkylation, C-hydroxylation, N-oxygenation with or without further oxidation, or a combination thereof, were detected by accurate mass spectrometry. The abundance of one metabolite, the N-dealkylated nitroso/oxime lapatinib metabolite (M9), correlated directly with the prevalence or the disruption of the MI complex with CYP3A4. Nitroso/oxime metabolite formation from secondary alkylamines has been proposed to occur through two possible pathways: (1) sequential N-dealkylation, N-hydroxylation, and dehydrogenation (primary hydroxylamine pathway) or (2) N-hydroxylation with dehydrogenation to yield a nitrone followed by N-dealkylation (secondary hydroxylamine pathway). All intermediates for the secondary hydroxylamine pathway were detected but the primary N-hydroxylamine intermediate of the primary hydroxylamine pathway was not. Our findings support the mechanism of lapatinib CYP3A4 inactivation as MI complex formation with the nitroso metabolite formed through the secondary hydroxylamine and nitrone pathway, rather than by N-dealkylation to the primary amine followed by N-hydroxylation and dehydrogenation as is usually assumed. DOI : 10.1124/dmd.113.051151
LAPATINIB
Inhibition of recombinant CYP3A4 (unknown origin)-mediated midazolam 1-hydroxylation at 50 uM
—
Inhibition = 45.0 %
Reference
Title : Metabolism-dependent inhibition of CYP3A4 by lapatinib: evidence for formation of a metabolic intermediate complex with a nitroso/oxime metabolite formed via a nitrone intermediate. Authors : Barbara JE, Kazmi F, Parkinson A, Buckley DB. Publication Year :
2013 Journal : Drug Metab. Dispos. Abstract : Metabolism-dependent inhibition (MDI) of cytochrome P450 (P450) enzymes has the potential to cause clinically relevant drug-drug interactions. In the case of several alkylamine drugs, MDI of P450 involves formation of a metabolite that binds quasi-irreversibly to the ferrous heme iron to form a metabolic intermediate (MI) complex. The specific metabolites coordinately bound to ferrous iron and the pathways leading to MI complex formation are the subject of debate. We describe an approach combining heme iron oxidation with potassium ferricyanide and metabolite profiling to probe the mechanism of MI complex-based CYP3A4 inactivation by the secondary alkylamine drug lapatinib. Ten metabolites formed from lapatinib by CYP3A4-mediated heteroatom dealkylation, C-hydroxylation, N-oxygenation with or without further oxidation, or a combination thereof, were detected by accurate mass spectrometry. The abundance of one metabolite, the N-dealkylated nitroso/oxime lapatinib metabolite (M9), correlated directly with the prevalence or the disruption of the MI complex with CYP3A4. Nitroso/oxime metabolite formation from secondary alkylamines has been proposed to occur through two possible pathways: (1) sequential N-dealkylation, N-hydroxylation, and dehydrogenation (primary hydroxylamine pathway) or (2) N-hydroxylation with dehydrogenation to yield a nitrone followed by N-dealkylation (secondary hydroxylamine pathway). All intermediates for the secondary hydroxylamine pathway were detected but the primary N-hydroxylamine intermediate of the primary hydroxylamine pathway was not. Our findings support the mechanism of lapatinib CYP3A4 inactivation as MI complex formation with the nitroso metabolite formed through the secondary hydroxylamine and nitrone pathway, rather than by N-dealkylation to the primary amine followed by N-hydroxylation and dehydrogenation as is usually assumed. DOI : 10.1124/dmd.113.051151
LEVOFLOXACIN
Time dependent inhibition of CYP3A4 (unknown origin) at 100 uM by LC/MS system
—
Inhibition < 10.0 %
Reference
Title : Combination of GSH trapping and time-dependent inhibition assays as a predictive method of drugs generating highly reactive metabolites. Authors : Nakayama S, Takakusa H, Watanabe A, Miyaji Y, Suzuki W, Sugiyama D, Shiosakai K, Honda K, Okudaira N, Izumi T, Okazaki O. Publication Year :
2011 Journal : Drug Metab. Dispos. Abstract : Covalent binding (CB) of reactive metabolites (RMs) is potentially involved in severe adverse drug reactions. Because the CB assay is of low throughput and costly, a qualitative trapping assay using agents such as [(35)S]GSH is often performed in the early stages of drug discovery. However, trapping methods alone cannot replace the CB assay. We hypothesized that the time-dependent inhibition (TDI) assay might be complementary to the [(35)S]GSH trapping assay in detecting RMs. We performed CB assays, [(35)S]GSH trapping assays, and TDI assays for 42 structurally diverse compounds. First, we showed that the [(35)S]GSH trapping assay alone does not correlate with the extent of CB. Four compounds that the [(35)S]GSH trapping assay failed to detect but that showed high extent of CB were inactivators of the enzyme in the TDI assay. There was a tendency for compounds judged as positive in the TDI assay to show a high degree of CB irrespective of the result of the [(35)S]GSH trapping assay. Finally, to combine parameters from the two assays, we introduced intrinsic clearance to describe the formation of RMs (CL(int, RMs)). The Spearman rank correlation coefficient between the extent of CB and CL(int, RMs) was 0.77 (p < 0.0001), which was better than that for the formation rates of [(35)S]GSH adducts. Therefore, we demonstrated that a combination of the [(35)S]GSH trapping and TDI assays is an effective method for detecting compounds potentially capable of generating highly reactive metabolites in the early stages of drug discovery. DOI : 10.1124/dmd.111.039180
MICONAZOLE
Inhibition of CYP3A4 (unknown origin)
—
IC50 = 850.0 nM
Reference
Title : Strategies for the development of highly selective cytochrome P450 inhibitors: Several CYP targets in current research. Authors : Zhao L, Sun N, Tian L, Zhao S, Sun B, Sun Y, Zhao D. Publication Year :
2019 Journal : Bioorg Med Chem Lett Abstract : Cytochromes P450 (CYPs) play an important role in the metabolism of endogenic and xenobiotic substances, especially drugs. In addition, many CYPs may serve as targets for disease treatment. However, due to the presence of a common heme, the hydrophobicity of the CYP binding cavity, and the high homology within the binding pocket, most CYP inhibitors lack selectivity, which often leads to drug-drug interactions. Therefore, it is meaningful to develop highly selective CYP inhibitors. In this review, we summarize some of the strategies that have been used to develop highly selective CYP inhibitors, such as the weakening of the heme-binding group interaction, reduction of molecular lipophilicity and introduction of small structural changes within compounds. DOI : 10.1016/j.bmcl.2019.06.040
MICONAZOLE
Inhibition of CYP3A4 in human liver microsome using Nifedipine as a substrate incubated for 10 mins in the presence of NADPH by LC-MS/MS analysis
Liver
IC50 = 74.2 nM
Reference
Title : Authors : Publication Year : Journal :
MIDAZOLAM
Inhibition of human CYP3A4 at 10 uM
—
Inhibition = 99.46 %
Reference
Title : Novel Pyrrolopyridone Bromodomain and Extra-Terminal Motif (BET) Inhibitors Effective in Endocrine-Resistant ER+ Breast Cancer with Acquired Resistance to Fulvestrant and Palbociclib. Authors : Li Y, Zhao J, Gutgesell LM, Shen Z, Ratia K, Dye K, Dubrovskyi O, Zhao H, Huang F, Tonetti DA, Thatcher GRJ, Xiong R. Publication Year :
2020 Journal : J Med Chem Abstract : Acquired resistance to fulvestrant and palbociclib is a new challenge to treatment of estrogen receptor positive (ER+) breast cancer. ER is expressed in most resistance settings; thus, bromodomain and extra-terminal protein inhibitors (BETi) that target BET-amplified ER-mediated transcription have therapeutic potential. Novel pyrrolopyridone BETi leveraged novel interactions with L92/L94 confirmed by a cocrystal structure of 27 with BRD4. Optimization of BETi using growth inhibition in fulvestrant-resistant (MCF-7:CFR) cells was confirmed in endocrine-resistant, palbociclib-resistant, and ESR1 mutant cell lines. 27 was more potent in MCF-7:CFR cells than six BET inhibitors in clinical trials. Transcriptomic analysis differentiated 27 from the benchmark BETi, JQ-1, showing downregulation of oncogenes and upregulation of tumor suppressors and apoptosis. The therapeutic approach was validated by oral administration of 27 in orthotopic xenografts of endocrine-resistant breast cancer in monotherapy and in combination with fulvestrant. Importantly, at an equivalent dose in rats, thrombocytopenia was mitigated. DOI : 10.1021/acs.jmedchem.0c00456
MILTEFOSINE
Inhibition of human microsomal CYP3A4 expressed in baculovirus infected insect cells at 10 uM using luciferin-IPA as substrate preincubated with enzyme/substrate mixture for 30 mins followed by further incubation with NADPH for 30 mins by CYP450 luciferase Glo assay relative to control
—
Inhibition = 8.4 %
Reference
Title : Discovery of a benzothiophene-flavonol halting miltefosine and antimonial drug resistance in Leishmania parasites through the application of medicinal chemistry, screening and genomics. Authors : Borsari C, Jiménez-Antón MD, Eick J, Bifeld E, Torrado JJ, Olías-Molero AI, Corral MJ, Santarem N, Baptista C, Severi L, Gul S, Wolf M, Kuzikov M, Ellinger B, Reinshagen J, Witt G, Linciano P, Tait A, Costantino L, Luciani R, Tejera Nevado P, Zander-Dinse D, Franco CH, Ferrari S, Moraes CB, Cordeiro-da-Silva A, Ponterini G, Clos J, Alunda JM, Costi MP. Publication Year :
2019 Journal : Eur J Med Chem Abstract : Leishmaniasis, a major health problem worldwide, has a limited arsenal of drugs for its control. The appearance of resistance to first- and second-line anti-leishmanial drugs confirms the need to develop new and less toxic drugs that overcome spontaneous resistance. In the present study, we report the design and synthesis of a novel library of 38 flavonol-like compounds and their evaluation in a panel of assays encompassing parasite killing, pharmacokinetics, genomics and ADME-Toxicity resulting in the progression of a compound in the drug discovery value chain. Compound 19, 2-(benzo[b]thiophen-3-yl)-3-hydroxy-6-methoxy-4H-chromen-4-one, exhibited a broad-spectrum activity against Leishmania spp. (EC50 1.9 μM for Leishmania infantum, 3.4 μM for L. donovani, 6.7 μM for L. major), Trypanosoma cruzi (EC50 7.5 μM) and T. brucei (EC50 0.8 μM). Focusing on anti-Leishmania activity, compound 19 challenge in vitro did not select for resistance markers in L. donovani, while a Cos-Seq screening for dominant resistance genes identified a gene locus on chromosome 36 that became ineffective at concentrations beyond EC50. Thus, compound 19 is a promising scaffold to tackle drug resistance in Leishmania infection. In vivo pharmacokinetic studies indicated that compound 19 has a long half-life (intravenous (IV): 63.2 h; per os (PO): 46.9 h) with an acceptable ADME-Toxicity profile. When tested in Leishmania infected hamsters, no toxicity and limited efficacy were observed. Low solubility and degradation were investigated spectroscopically as possible causes for the sub-optimal pharmacokinetic properties. Compound 19 resulted a specific compound based on the screening against a protein set, following the intrinsic fluorescence changes. DOI : 10.1016/j.ejmech.2019.111676
OLANZAPINE
Time dependent inhibition of CYP3A4 (unknown origin) at 100 uM by LC/MS system
—
Inhibition < 10.0 %
Reference
Title : Combination of GSH trapping and time-dependent inhibition assays as a predictive method of drugs generating highly reactive metabolites. Authors : Nakayama S, Takakusa H, Watanabe A, Miyaji Y, Suzuki W, Sugiyama D, Shiosakai K, Honda K, Okudaira N, Izumi T, Okazaki O. Publication Year :
2011 Journal : Drug Metab. Dispos. Abstract : Covalent binding (CB) of reactive metabolites (RMs) is potentially involved in severe adverse drug reactions. Because the CB assay is of low throughput and costly, a qualitative trapping assay using agents such as [(35)S]GSH is often performed in the early stages of drug discovery. However, trapping methods alone cannot replace the CB assay. We hypothesized that the time-dependent inhibition (TDI) assay might be complementary to the [(35)S]GSH trapping assay in detecting RMs. We performed CB assays, [(35)S]GSH trapping assays, and TDI assays for 42 structurally diverse compounds. First, we showed that the [(35)S]GSH trapping assay alone does not correlate with the extent of CB. Four compounds that the [(35)S]GSH trapping assay failed to detect but that showed high extent of CB were inactivators of the enzyme in the TDI assay. There was a tendency for compounds judged as positive in the TDI assay to show a high degree of CB irrespective of the result of the [(35)S]GSH trapping assay. Finally, to combine parameters from the two assays, we introduced intrinsic clearance to describe the formation of RMs (CL(int, RMs)). The Spearman rank correlation coefficient between the extent of CB and CL(int, RMs) was 0.77 (p < 0.0001), which was better than that for the formation rates of [(35)S]GSH adducts. Therefore, we demonstrated that a combination of the [(35)S]GSH trapping and TDI assays is an effective method for detecting compounds potentially capable of generating highly reactive metabolites in the early stages of drug discovery. DOI : 10.1124/dmd.111.039180
PHENYTOIN
Time dependent inhibition of CYP3A4 (unknown origin) at 100 uM by LC/MS system
—
Inhibition < 10.0 %
Reference
Title : Combination of GSH trapping and time-dependent inhibition assays as a predictive method of drugs generating highly reactive metabolites. Authors : Nakayama S, Takakusa H, Watanabe A, Miyaji Y, Suzuki W, Sugiyama D, Shiosakai K, Honda K, Okudaira N, Izumi T, Okazaki O. Publication Year :
2011 Journal : Drug Metab. Dispos. Abstract : Covalent binding (CB) of reactive metabolites (RMs) is potentially involved in severe adverse drug reactions. Because the CB assay is of low throughput and costly, a qualitative trapping assay using agents such as [(35)S]GSH is often performed in the early stages of drug discovery. However, trapping methods alone cannot replace the CB assay. We hypothesized that the time-dependent inhibition (TDI) assay might be complementary to the [(35)S]GSH trapping assay in detecting RMs. We performed CB assays, [(35)S]GSH trapping assays, and TDI assays for 42 structurally diverse compounds. First, we showed that the [(35)S]GSH trapping assay alone does not correlate with the extent of CB. Four compounds that the [(35)S]GSH trapping assay failed to detect but that showed high extent of CB were inactivators of the enzyme in the TDI assay. There was a tendency for compounds judged as positive in the TDI assay to show a high degree of CB irrespective of the result of the [(35)S]GSH trapping assay. Finally, to combine parameters from the two assays, we introduced intrinsic clearance to describe the formation of RMs (CL(int, RMs)). The Spearman rank correlation coefficient between the extent of CB and CL(int, RMs) was 0.77 (p < 0.0001), which was better than that for the formation rates of [(35)S]GSH adducts. Therefore, we demonstrated that a combination of the [(35)S]GSH trapping and TDI assays is an effective method for detecting compounds potentially capable of generating highly reactive metabolites in the early stages of drug discovery. DOI : 10.1124/dmd.111.039180
POSACONAZOLE
Inhibition of human recombinant CYP3A4
—
IC50 = 350.0 nM
Reference
Title : Rational modification of a candidate cancer drug for use against Chagas disease. Authors : Kraus JM, Verlinde CL, Karimi M, Lepesheva GI, Gelb MH, Buckner FS. Publication Year :
2009 Journal : J. Med. Chem. Abstract : Chagas disease is one of the major neglected diseases of the world. Existing drug therapies are limited, ineffective, and highly toxic. We describe a novel strategy of drug discovery of adapting an existing clinical compound with excellent pharmaceutical properties to target a pathogenic organism. The protein farnesyltransferase (PFT) inhibitor tipifarnib, now in phase III anticancer clinical trials, was previously found to kill Trypanosoma cruzi by blocking sterol 14 alpha-demethylase (14DM). We rationally developed tipifarnib analogues that display reduced affinity for human PFT to reduce toxicity while increasing affinity for parasite 14DM. The lead compound has picomolar activity against cultured T. cruzi and is efficacious in a mouse model of acute Chagas disease. DOI : 10.1021/jm801313t
POSACONAZOLE
Inhibition of human recombinant CYP3A4
—
IC50 = 82.0 nM
Reference
Title : Second generation analogues of the cancer drug clinical candidate tipifarnib for anti-Chagas disease drug discovery. Authors : Kraus JM, Tatipaka HB, McGuffin SA, Chennamaneni NK, Karimi M, Arif J, Verlinde CL, Buckner FS, Gelb MH. Publication Year :
2010 Journal : J. Med. Chem. Abstract : We previously reported that the cancer drug clinical candidate tipifarnib kills the causative agent of Chagas disease, Trypanosoma cruzi, by blocking ergosterol biosynthesis at the level of inhibition of lanosterol 14alpha-demethylase. Tipifarnib is an inhibitor of human protein farnesyltransferase. We synthesized tipifarnib analogues that no longer bind to protein farnesyltransferase and display increased potency for killing parasites. This was achieved in a structure-guided fashion by changing the substituents attached to the phenyl group at the 4-position of the quinoline ring of tipifarnib and by replacing the amino group by OMe. Several compounds that kill Trypanosoma cruzi at subnanomolar concentrations and are devoid of protein farnesyltransferase inhibition were discovered. The compounds are shown to be advantageous over other lanosterol 14alpha-demethylase inhibitors in that they show only modest potency for inhibition of human cytochrome P450 (3A4). Since tipifarnib displays high oral bioavailability and acceptable pharmacokinetic properties, the newly discovered tipifarnib analogues are ideal leads for the development of drugs to treat Chagas disease. DOI : 10.1021/jm9013136
POSACONAZOLE
Inhibition of CYP3A4 in human hepatocytes using testosterone as substrate by HPLC/MS/MS method
—
IC50 = 50.0 nM
Reference
Title : Design and optimization of highly-selective fungal CYP51 inhibitors. Authors : Hoekstra WJ, Garvey EP, Moore WR, Rafferty SW, Yates CM, Schotzinger RJ. Publication Year :
2014 Journal : Bioorg. Med. Chem. Lett. Abstract : While the orally-active azoles such as voriconazole and itraconazole are effective antifungal agents, they potently inhibit a broad range of off-target human cytochrome P450 enzymes (CYPs) leading to various safety issues (e.g., drug-drug interactions, liver toxicity). Herein, we describe rationally-designed, broad-spectrum antifungal agents that are more selective for the target fungal enzyme, CYP51, than related human CYP enzymes such as CYP3A4. Using proprietary methodology, the triazole metal-binding group found in current clinical agents was replaced with novel, less avid metal-binding groups in concert with potency-enhancing molecular scaffold modifications. This process produced a unique series of fungal CYP51-selective inhibitors that included the oral antifungal 7d (VT-1161), now in Phase 2 clinical trials. This series exhibits excellent potency against key yeast and dermatophyte strains. The chemical methodology described is potentially applicable to the design of new and more effective metalloenzyme inhibitor treatments for a broad array of diseases. DOI : 10.1016/j.bmcl.2014.05.068
POSACONAZOLE
Inhibition of CYP3A4 in human hepatocyte microsomes using testosterone substrate by HPLC/MS/MS method
—
IC50 = 75.0 nM
Reference
Title : Design and optimization of highly-selective, broad spectrum fungal CYP51 inhibitors. Authors : Yates CM, Garvey EP, Shaver SR, Schotzinger RJ, Hoekstra WJ. Publication Year :
2017 Journal : Bioorg Med Chem Lett Abstract : While the orally-active azoles such as fluconazole and posaconazole are effective antifungal agents, they potently inhibit a broad range of off-target human cytochrome P450 enzymes (CYPs) leading to various safety issues (e.g., drug-drug interactions, liver, and reproductive toxicities). Recently we described the rationally-designed, antifungal agent VT-1161 that is more selective for fungal CYP51 than related human CYP enzymes such as CYP3A4. Herein, we describe the use of a homology model of Aspergillus fumigatus to design and optimize a novel series of highly selective, broad spectrum fungal CYP51 inhibitors. This series includes the oral antifungal VT-1598 that exhibits excellent potency against yeast, dermatophyte, and mold fungal pathogens. DOI : 10.1016/j.bmcl.2017.06.037
POSACONAZOLE
Inhibition of CYP3A4 in human liver microsomes by LC-MS/MS analysis
Liver
IC50 = 150.0 nM
Reference
Title : Development of posaconazole-based analogues as hedgehog signaling pathway inhibitors. Authors : Teske KA, Dash RC, Morel SR, Chau LQ, Wechsler-Reya RJ, Hadden MK. Publication Year :
2019 Journal : Eur J Med Chem Abstract : Inhibition of the hedgehog (Hh) signaling pathway has been validated as a therapeutic strategy to treat basal cell carcinoma and holds potential for several other forms of human cancer. Itraconazole and posaconazole are clinically useful triazole anti-fungals that are being repurposed as anti-cancer agents based on their ability to inhibit the Hh pathway. We have previously demonstrated that removal of the triazole from itraconazole does not affect its ability to inhibit the Hh pathway while abolishing its primary side effect, potent inhibition of Cyp3A4. To develop structure-activity relationships for the related posaconazole scaffold, we synthesized and evaluated a series of des-triazole analogues designed through both ligand- and structure-based methods. These compounds demonstrated improved anti-Hh properties compared to posaconazole and enhanced stability without inhibiting Cyp3A4. In addition, we utilized a series of molecular dynamics and binding energy studies to probe specific interactions between the compounds and their proposed binding site on Smoothened. These studies strongly suggest that the tetrahydrofuran region of the scaffold projects out of the binding site and that π-π interactions between the compound and Smoothened play a key role in stabilizing the bound analogues. DOI : 10.1016/j.ejmech.2018.11.056
POSACONAZOLE
Inhibition of recombinant human CYP3A4 using Luciferin-PPXE as substrate preincubated for 10 mins followed by NADPH addition measured after 15 mins by luminometric method
—
IC50 = 672.1 nM
Reference
Title : Antifungal activity, mode of action variability, and subcellular distribution of coumarin-based antifungal azoles. Authors : Elias R, Benhamou RI, Jaber QZ, Dorot O, Zada SL, Oved K, Pichinuk E, Fridman M. Publication Year :
2019 Journal : Eur J Med Chem Abstract : Azole antifungals inhibit the biosynthesis of ergosterol, the fungal equivalent of cholesterol in mammalian cells. Here we report an investigation of the activity of coumarin-substituted azole antifungals. Screening against a panel of Candida pathogens, including a mutant lacking CYP51, the target of antifungal azoles, revealed that this enzyme is inhibited by triazole-based antifungals, whereas imidazole-based derivatives have more than one mode of action. The imidazole-bearing antifungals more effectively reduced trailing growth associated with persistence and/or recurrence of fungal infections than triazole-based derivatives. The imidazole derivatives were more toxic to mammalian cells and more potently inhibited the activity of CYP3A4, which is one of the main causes of azole toxicity. Using live cell imaging, we showed that regardless of the type of azole ring fluorescent 7-diethylaminocoumarin-based azoles localized to the endoplasmic reticulum, the organelle that harbors CYP51. This study suggests that the coumarin is a promising scaffold for development of novel azole-based antifungals that effectively localize to the fungal cell endoplasmic reticulum. DOI : 10.1016/j.ejmech.2019.07.003
PREGABALIN
Inhibition of recombinant CYP3A4 (unknown origin) using BFC as substrate at 1 uM measured after 30 mins in presence of NADPH generating system by fluorescence assay relative to control
—
Inhibition = 43.0 %
Reference
Title : Authors : Publication Year : Journal :
RITONAVIR
Inhibition of CYP3A4-mediated lopinavir bioactivation in human liver microsomes assessed as inhibition of lopinavir-GSH adduct I and II formation at 2 uM after after 50 mins by UPLC-TOFMS analysis
Liver
Inhibition = 80.0 %
Reference
Title : CYP3A4-mediated lopinavir bioactivation and its inhibition by ritonavir. Authors : Li F, Lu J, Ma X. Publication Year :
2012 Journal : Drug Metab. Dispos. Abstract : The combination of lopinavir (LPV) and ritonavir (RTV) is one of the preferred regimens for the treatment of HIV infection with confirmed efficacy and relatively low toxicity. LPV alone suffers the poor bioavailability due to its rapid and extensive metabolism. RTV boosts the plasma concentration of LPV by suppressing its metabolism and thus increasing LPV efficacy. In the current study, we found that RTV also inhibits LPV bioactivation. LPV bioactivation was investigated in human liver microsomes and cDNA-expressed human cytochromes P450. Twelve GSH-trapped reactive metabolites of LPV were identified by using a metabolomic approach. Semicarbazide-trapped reactive metabolites of LPV were also detected. RTV effectively suppressed all pathways of LPV bioactivation via CYP3A4 inhibition. Our data together with previous reports suggest that LPV plus RTV is an ideal combination because RTV not only boosts LPV plasma concentration, but it decreases LPV bioactivation. DOI : 10.1124/dmd.111.041400
RITONAVIR
Time dependent inhibition of CYP3A4 (unknown origin) at 1 uM by LC/MS system
—
Inhibition = 75.0 %
Reference
Title : Combination of GSH trapping and time-dependent inhibition assays as a predictive method of drugs generating highly reactive metabolites. Authors : Nakayama S, Takakusa H, Watanabe A, Miyaji Y, Suzuki W, Sugiyama D, Shiosakai K, Honda K, Okudaira N, Izumi T, Okazaki O. Publication Year :
2011 Journal : Drug Metab. Dispos. Abstract : Covalent binding (CB) of reactive metabolites (RMs) is potentially involved in severe adverse drug reactions. Because the CB assay is of low throughput and costly, a qualitative trapping assay using agents such as [(35)S]GSH is often performed in the early stages of drug discovery. However, trapping methods alone cannot replace the CB assay. We hypothesized that the time-dependent inhibition (TDI) assay might be complementary to the [(35)S]GSH trapping assay in detecting RMs. We performed CB assays, [(35)S]GSH trapping assays, and TDI assays for 42 structurally diverse compounds. First, we showed that the [(35)S]GSH trapping assay alone does not correlate with the extent of CB. Four compounds that the [(35)S]GSH trapping assay failed to detect but that showed high extent of CB were inactivators of the enzyme in the TDI assay. There was a tendency for compounds judged as positive in the TDI assay to show a high degree of CB irrespective of the result of the [(35)S]GSH trapping assay. Finally, to combine parameters from the two assays, we introduced intrinsic clearance to describe the formation of RMs (CL(int, RMs)). The Spearman rank correlation coefficient between the extent of CB and CL(int, RMs) was 0.77 (p < 0.0001), which was better than that for the formation rates of [(35)S]GSH adducts. Therefore, we demonstrated that a combination of the [(35)S]GSH trapping and TDI assays is an effective method for detecting compounds potentially capable of generating highly reactive metabolites in the early stages of drug discovery. DOI : 10.1124/dmd.111.039180
RITONAVIR
Antiviral activity as inhibition of human liver microsomal Cytochrome P450 3A4
—
IC50 = 250.0 nM
Reference
Title : Potent piperazine hydroxyethylamine HIV protease inhibitors containing novel P3 ligands. Authors : Chen X, Kempf DJ, Sham HL, Green BE, Molla A, Korneyeva M, Vasavanonda S, Wideburg NE, Saldivar A, Marsh KC, McDonald E, Norbeck DW. Publication Year :
1998 Journal : Bioorg. Med. Chem. Lett. Abstract : The 2-isopropyl thiazolyl group is a highly optimized P3 ligand for C2 symmetry-based HIV protease inhibitors, as exemplified in the drug ritonavir. Here we report that incorporation of this P3 ligand into a piperazine hydroxyethylamine series also yielded novel, highly potent inhibitors. In tissue culture assays, the presence of human serum was less deleterious to the activity of these inhibitors than to that of ritonavir. Furthermore, potent activity against ritonavir resistant HIV was observed. DOI : 10.1016/s0960-894x(98)00653-2
RITONAVIR
Inhibition of human recombinant CYP3A4-mediated oxidation of 7-benzyloxyquinoline
—
IC50 = 50.0 nM
Reference
Title : Synthesis and evaluation of inhibitors of cytochrome P450 3A (CYP3A) for pharmacokinetic enhancement of drugs. Authors : Flentge CA, Randolph JT, Huang PP, Klein LL, Marsh KC, Harlan JE, Kempf DJ. Publication Year :
2009 Journal : Bioorg. Med. Chem. Lett. Abstract : The HIV protease inhibitor ritonavir (RTV) is also a potent inhibitor of the metabolizing enzyme cytochrome P450 3A (CYP3A) and is clinically useful in HIV therapy in its ability to enhance human plasma levels of other HIV protease inhibitors (PIs). A novel series of CYP3A inhibitors was designed around the structural elements of RTV believed to be important to CYP3A inhibition, with general design features being the attachment of groups that mimic the P2-P3 segment of RTV to a soluble core. Several analogs were found to strongly enhance plasma levels of lopinavir (LPV), including 8, which compares favorably with RTV in the same model. Interestingly, an inverse correlation between in vitro inhibition of CYP3A and elevation of LPV was observed. The compounds described in this study may be useful for enhancing the pharmacokinetics of drugs that are metabolized by CYP3A. DOI : 10.1016/j.bmcl.2009.07.118
RITONAVIR
Inhibition of human CYP3A4
—
Ki < 100.0 nM
Reference
Title : Rapid clinical induction of hepatic cytochrome P4502B6 activity by ritonavir. Authors : Kharasch ED, Mitchell D, Coles R, Blanco R. Publication Year :
2008 Journal : Antimicrob. Agents Chemother. Abstract : Ritonavir is the most potent and efficacious inhibitor of cytochrome P4503A (CYP3A), and it is used accordingly for the pharmacoenhancement of other antiretrovirals. Paradoxically, ritonavir induces the clinical metabolism and clearance of many drugs. The mechanism by which ritonavir inhibits and induces clinical drug metabolism is unknown. Ritonavir induces CYP2B6 in human hepatocytes. This investigation tested the hypothesis that ritonavir induces human CYP2B6 in vivo. Thirteen healthy human immunodeficiency virus-negative volunteers underwent a three-way sequential crossover protocol, receiving racemic bupropion after nothing (control), 3 days of treatment with ritonavir, and 2.5 weeks of treatment with ritonavir (400 mg twice a day). Stereoselective bupropion hydroxylation was used as an in vivo probe for CYP2B6 activity. Plasma and urine (R)- and (S)-bupropion and (R,R)- and (S,S)-hydroxybupropion concentrations were measured by liquid chromatography-mass spectrometry. Racemic, (R)-, and (S)-bupropion plasma ratios of the area under the concentration-time curve from 0 h to infinity (AUC(0-infinity)) (ritonavir/control) were significantly reduced to 0.84, 0.86, and 0.80, respectively, after 3 days of ritonavir treatment and to 0.67, 0.69, and 0.60 after steady-state ritonavir treatment. Apparent oral clearances for racemic, (R)-, and (S)-bupropion all were significantly increased by 1.2-fold after 3 days of ritonavir treatment and by 1.4-, 1.7-, and 1.5-fold after steady-state ritonavir treatment. The plasma (S,S)-hydroxybupropion/(S)-bupropion AUC(0-72) ratio was significantly increased by ritonavir. Formation clearances of both (R,R)- and (S,S)-hydroxybupropion were increased 1.8-fold after 3 days of ritonavir treatment and 2.1-fold after steady-state ritonavir treatment. These results show that ritonavir induces human CYP2B6 activity. Induction is rapid, occurring after only 3 days of ritonavir, and is sustained for at least 2 weeks. The ritonavir induction of CYP2B6 activity may have significant implications for drug interactions and clarify previously unexplained interactions. DOI : 10.1128/aac.01600-07
RITONAVIR
Mechanism based inhibition of human cytochrome P450 3A4
—
Ki = 100.0 nM
Reference
Title : Cytochrome p450 enzymes mechanism based inhibitors: common sub-structures and reactivity. Authors : Fontana E, Dansette PM, Poli SM. Publication Year :
2005 Journal : Curr. Drug Metab. Abstract : The inhibition of human cytochrome P450s (CYPs) is one of the most common mechanisms which can lead to drug-drug interactions. The inhibition of CYPs can be reversible (competitive or non-competitive) or irreversible. Irreversible inhibition usually derives from activation of a drug by CYPs into a reactive metabolite, which tightly binds to the enzyme active site, leading to a long lasting inactivation. This process is called "mechanism based inhibition" or "suicide inhibition". The irreversible inactivation usually implies the formation of a covalent bond between the metabolite and the enzyme, which can lead to hapten formation and can in some cases trigger an autoimmune-response. For these reasons it is of utmost importance to study the mechanism of the CYP inhibition of new potential drugs as early as possible during the drug discovery process. The literature on CYPs is vast and covers numerous aspects of their biology and biochemistry, however to our knowledge there is no general and systematic review focusing on mechanism-based inhibitors; we have reviewed the literature and compiled all the available data on chemical entities, which are known to be CYP suicide inhibitors. Each compound is reported together with its chemical structure, the CYP isoform and the parameters describing the inactivation. Literature references are reported together with their PMID (PubMed ID number) to allow a fast retrieval of the papers. This review offers a quick reference to help predict liabilities of new chemical entities without carrying out extensive in vitro work, and will hopefully help in designing safer drugs. DOI : 10.2174/138920005774330639
RITONAVIR
Inhibition of human CYP3A4 in liver microsomes
Liver
IC50 = 100.0 nM
Reference
Title : Preclinical metabolism of LB42908, a novel farnesyl transferase inhibitor, and its effects on the cytochrome P450 isozyme activities. Authors : Kim HJ, Lee SH, Koh JS, Lee SH. Publication Year :
2012 Journal : Bioorg. Med. Chem. Lett. Abstract : Metabolism of LB42908, a novel farnesyl transferase inhibitor, was investigated for preclinical development. In vitro hepatic metabolism of LB42908 gave rise to at least 9 metabolites via phase I biotransformation pathways, which were characterized by HPLC-UV, LC-MS, and LC-MS/MS analyses. N-Dealkylation was shown to be a major phase I metabolic pathway. Species-specific in vitro metabolism of LB42908 was studied in liver fractions of rat, dog, monkey, and human. Order of metabolic stability is human≈dog>rat≈monkey in both S9 and microsomal fractions. Tissue-specific metabolism of LB42908 in various tissue homogenates of rats demonstrated that the liver was the major organ responsible for phase I metabolism of LB42908. The results from both qualitative and quantitative metabolism studies such as metabolic profiling and metabolic clearance indicated that dog would be the animal model of choice for preclinical toxicology studies. In addition, LB42908 was a potent CYP3A4 inhibitor in human liver microsomes and induced the activities of several CYP isozymes, implying that it has the potential for drug-drug interactions. Repeated dosing of LB42908 in rats did not significantly affect its own metabolism, indicating that long-term administration of LB42908 would not alter its pharmacokinetic profiles. DOI : 10.1016/j.bmcl.2012.03.070
RITONAVIR
Time dependent inhibition of CYP3A4-mediated testosterone-6-beta hydroxylation in human liver microsome
Liver
Ki = 170.0 nM
Reference
Title : Mechanism-based inactivation (MBI) of cytochrome P450 enzymes: structure-activity relationships and discovery strategies to mitigate drug-drug interaction risks. Authors : Orr ST, Ripp SL, Ballard TE, Henderson JL, Scott DO, Obach RS, Sun H, Kalgutkar AS. Publication Year :
2012 Journal : J. Med. Chem. DOI : 10.1021/jm300065h
RITONAVIR
Inhibition of C-terminal His-tagged wild type CYP3A4 (unknown origin)-mediated hydroxylation of 7-benzyloxy-4-trifluoromethylcoumarin expressed in Escherichia coli after 5 mins by fluorimetric analysis
—
IC50 = 550.0 nM
Reference
Title : Pyridine-substituted desoxyritonavir is a more potent inhibitor of cytochrome P450 3A4 than ritonavir. Authors : Sevrioukova IF, Poulos TL. Publication Year :
2013 Journal : J. Med. Chem. Abstract : Utilization of the cytochrome P450 3A4 (CYP3A4) inhibitor ritonavir as a pharmacoenhancer for anti-HIV drugs revolutionized the treatment of HIV infection. However, owing to ritonavir-related complications, there is a need for development of new CYP3A4 inhibitors with improved pharmacochemical properties, which requires a full understanding of the CYP3A4 inactivation mechanisms and the unraveling of possible inhibitor binding modes. We investigated the mechanism of CYP3A4 interaction with three desoxyritonavir analogues, containing the heme-ligating imidazole, oxazole, or pyridine group instead of the thiazole moiety (compounds 1, 2, and 3, respectively). Our data show that compound 3 is superior to ritonavir in terms of binding affinity and inhibitory potency owing to greater flexibility and the ability to adopt a conformation that minimizes steric clashing and optimizes protein-ligand interactions. Additionally, Ser119 was identified as a key residue assisting binding of ritonavir-like inhibitors, which emphasizes the importance of polar interactions in the CYP3A4-ligand association. DOI : 10.1021/jm400288z
RITONAVIR
Inhibition of human CYP3A4-mediated midazolam 1'-hydroxylase activity
—
IC50 = 110.0 nM
Reference
Title : Structure-activity relationships of diamine inhibitors of cytochrome P450 (CYP) 3A as novel pharmacoenhancers, part I: core region. Authors : Liu H, Xu L, Hui H, Vivian R, Callebaut C, Murray BP, Hong A, Lee MS, Tsai LK, Chau JK, Stray KM, Cannizzaro C, Choi YC, Rhodes GR, Desai MC. Publication Year :
2014 Journal : Bioorg. Med. Chem. Lett. Abstract : Ritonavir (RTV), an HIV-1 protease inhibitor (PI), is also a potent mechanism-based inhibitor of human cytochrome P450 3A (CYP3A) and has been widely prescribed as a pharmacoenhancer. As a boosting agent for marketed PIs, it reduces pill burden, and improves compliance. Removal of the hydroxyl group from RTV reduces, but does not eliminate HIV PI activity and does not affect CYP3A inhibition. Herein we report the discovery of a novel series of CYP3A inhibitors that are devoid of antiviral activity. The synthesis and evaluation of analogs with extensive modifications of the 1,4-diamine core along with the structure activity relationships with respect to anti-HIV activity, CYP3A inhibitory activity, selectivity against other CYP enzymes and the human pregnane X receptor (PXR) will be discussed. DOI : 10.1016/j.bmcl.2013.12.058
RITONAVIR
Inhibition of human full length CYP3A4 assessed using 7-benzyloxy-4 (trifluoromethyl)coumarin as substrate preincubated for 20 mins in presence of NADPH followed by substrate addition and measured for 2 mins by fluorometric analysis
—
IC50 = 130.0 nM
Reference
Title : An increase in side-group hydrophobicity largely improves the potency of ritonavir-like inhibitors of CYP3A4. Authors : Samuels ER, Sevrioukova IF. Publication Year :
2020 Journal : Bioorg Med Chem Abstract : Identification of structural determinants required for potent inhibition of drug-metabolizing cytochrome P450 3A4 (CYP3A4) could help develop safer drugs and more effective pharmacoenhancers. We utilize a rational inhibitor design to decipher structure-activity relationships in analogues of ritonavir, a highly potent CYP3A4 inhibitor marketed as pharmacoenhancer. Analysis of compounds with the R<sub>1</sub> side-group as phenyl or naphthalene and R<sub>2</sub> as indole or naphthalene in different stereo configuration showed that (i) analogues with the R<sub>2</sub>-naphthalene tend to bind tighter and inhibit CYP3A4 more potently than the R<sub>2</sub>-phenyl/indole containing counterparts; (ii) stereochemistry becomes a more important contributing factor, as the bulky side-groups limit the ability to optimize protein-ligand interactions; (iii) the relationship between the R<sub>1</sub>/R<sub>2</sub> configuration and preferential binding to CYP3A4 is complex and depends on the side-group functionality/interplay and backbone spacing; and (iv) three inhibitors, 5a-b and 7d, were superior to ritonavir (IC<sub>50</sub> of 0.055-0.085 μM vs. 0.130 μM, respectively). DOI : 10.1016/j.bmc.2020.115349
SAQUINAVIR
Mechanism based inhibition of human cytochrome P450 3A4 measured by testosterone hydroxylation
—
Ki = 170.0 nM
Reference
Title : Cytochrome p450 enzymes mechanism based inhibitors: common sub-structures and reactivity. Authors : Fontana E, Dansette PM, Poli SM. Publication Year :
2005 Journal : Curr. Drug Metab. Abstract : The inhibition of human cytochrome P450s (CYPs) is one of the most common mechanisms which can lead to drug-drug interactions. The inhibition of CYPs can be reversible (competitive or non-competitive) or irreversible. Irreversible inhibition usually derives from activation of a drug by CYPs into a reactive metabolite, which tightly binds to the enzyme active site, leading to a long lasting inactivation. This process is called "mechanism based inhibition" or "suicide inhibition". The irreversible inactivation usually implies the formation of a covalent bond between the metabolite and the enzyme, which can lead to hapten formation and can in some cases trigger an autoimmune-response. For these reasons it is of utmost importance to study the mechanism of the CYP inhibition of new potential drugs as early as possible during the drug discovery process. The literature on CYPs is vast and covers numerous aspects of their biology and biochemistry, however to our knowledge there is no general and systematic review focusing on mechanism-based inhibitors; we have reviewed the literature and compiled all the available data on chemical entities, which are known to be CYP suicide inhibitors. Each compound is reported together with its chemical structure, the CYP isoform and the parameters describing the inactivation. Literature references are reported together with their PMID (PubMed ID number) to allow a fast retrieval of the papers. This review offers a quick reference to help predict liabilities of new chemical entities without carrying out extensive in vitro work, and will hopefully help in designing safer drugs. DOI : 10.2174/138920005774330639
SAQUINAVIR
Time dependent inhibition of CYP3A4-mediated testosterone-6-beta hydroxylation in human liver microsome
Liver
Ki = 650.0 nM
Reference
Title : Mechanism-based inactivation (MBI) of cytochrome P450 enzymes: structure-activity relationships and discovery strategies to mitigate drug-drug interaction risks. Authors : Orr ST, Ripp SL, Ballard TE, Henderson JL, Scott DO, Obach RS, Sun H, Kalgutkar AS. Publication Year :
2012 Journal : J. Med. Chem. DOI : 10.1021/jm300065h
SULFAMETHOXAZOLE
Time dependent inhibition of CYP3A4 (unknown origin) at 100 uM by LC/MS system
—
Inhibition < 10.0 %
Reference
Title : Combination of GSH trapping and time-dependent inhibition assays as a predictive method of drugs generating highly reactive metabolites. Authors : Nakayama S, Takakusa H, Watanabe A, Miyaji Y, Suzuki W, Sugiyama D, Shiosakai K, Honda K, Okudaira N, Izumi T, Okazaki O. Publication Year :
2011 Journal : Drug Metab. Dispos. Abstract : Covalent binding (CB) of reactive metabolites (RMs) is potentially involved in severe adverse drug reactions. Because the CB assay is of low throughput and costly, a qualitative trapping assay using agents such as [(35)S]GSH is often performed in the early stages of drug discovery. However, trapping methods alone cannot replace the CB assay. We hypothesized that the time-dependent inhibition (TDI) assay might be complementary to the [(35)S]GSH trapping assay in detecting RMs. We performed CB assays, [(35)S]GSH trapping assays, and TDI assays for 42 structurally diverse compounds. First, we showed that the [(35)S]GSH trapping assay alone does not correlate with the extent of CB. Four compounds that the [(35)S]GSH trapping assay failed to detect but that showed high extent of CB were inactivators of the enzyme in the TDI assay. There was a tendency for compounds judged as positive in the TDI assay to show a high degree of CB irrespective of the result of the [(35)S]GSH trapping assay. Finally, to combine parameters from the two assays, we introduced intrinsic clearance to describe the formation of RMs (CL(int, RMs)). The Spearman rank correlation coefficient between the extent of CB and CL(int, RMs) was 0.77 (p < 0.0001), which was better than that for the formation rates of [(35)S]GSH adducts. Therefore, we demonstrated that a combination of the [(35)S]GSH trapping and TDI assays is an effective method for detecting compounds potentially capable of generating highly reactive metabolites in the early stages of drug discovery. DOI : 10.1124/dmd.111.039180
SULFAPHENAZOLE
Inhibition of CYP3A4 in human liver microsomes assessed as midazolam hydroxylation to 1'-hydroxymidazolam at 10 uM after 10 mins relative to control
Liver
Inhibition = 21.0 %
Reference
Title : Rational development of 4-aminopyridyl-based inhibitors targeting Trypanosoma cruzi CYP51 as anti-chagas agents. Authors : Choi JY, Calvet CM, Gunatilleke SS, Ruiz C, Cameron MD, McKerrow JH, Podust LM, Roush WR. Publication Year :
2013 Journal : J. Med. Chem. Abstract : A new series of 4-aminopyridyl-based lead inhibitors targeting Trypanosoma cruzi CYP51 (TcCYP51) has been developed using structure-based drug design as well as structure-property relationship (SPR) analyses. The screening hit starting point, LP10 (KD ≤ 42 nM; EC50 = 0.65 μM), has been optimized to give the potential leads 14t, 27i, 27q, 27r, and 27t, which have low-nanomolar binding affinity to TcCYP51 and significant activity against T. cruzi amastigotes cultured in human myoblasts (EC50 = 14-18 nM for 27i and 27r). Many of the optimized compounds have improved microsome stability, and most are selective against human CYPs 1A2, 2D6, and 3A4 (<50% inhibition at 1 μM). A rationale for the improvement in microsome stability and selectivity of inhibitors against human metabolic CYP enzymes is presented. In addition, the binding mode of 14t with the Trypanosoma brucei CYP51 (TbCYP51) orthologue has been characterized by X-ray structure analysis. DOI : 10.1021/jm401067s
TACROLIMUS
Inhibition of human CYP3A4 expressed in supersomes assessed inhibition of 1'-OH midazolam formation preincubated at 2 uM followed by addition of 20-fold diluted buffer with 20 uM MDZ by LC-MS method in presence of NADPH
—
Inhibition = 42.0 %
Reference
Title : Cyclosporine A- and tacrolimus-mediated inhibition of CYP3A4 and CYP3A5 in vitro. Authors : Amundsen R, Åsberg A, Ohm IK, Christensen H. Publication Year :
2012 Journal : Drug Metab. Dispos. Abstract : Cyclosporine A (CsA) and tacrolimus (Tac) are immunosuppressive drugs used in the majority of patients with solid organ transplants, generally in combination with a wide range of drugs. CsA and Tac seem not only to be substrates of CYP3A but have also been described as inhibitors of CYP3A. For CsA, in particular, inhibition of CYP3A has been suggested as the main mechanism of interactions seen clinically with various drugs. The aim of this study was to investigate the inhibitory effect and inhibition characteristics of CsA and Tac on CYP3A4 and CYP3A5 in vitro and to evaluate its clinical relevance. Inhibition by CsA and Tac was studied using midazolam as the probe substrate in coincubation and preincubation investigations using human liver microsomes (HLMs) as well as specific CYP3A4- and CYP3A5-expressing insect microsomes (Supersomes). In vitro-in vivo extrapolations (IVIVEs) were performed to evaluate the clinical relevance of the inhibition. Both CsA and Tac competitively inhibited CYP3A in HLMs, showing inhibition constants (K(i)) of 0.98 and 0.61 μM, respectively. Experiments in Supersomes revealed that Tac inhibited both CYP3A4 and CYP3A5, whereas CsA only inhibited CYP3A4. In contrast to the HLM experiments, studies in Supersomes showed inhibition by Tac to be NADPH- and time-dependent, with a 5-fold reduction in IC(50) after preincubation, supporting a time-dependent inhibition mechanism in recombinant microsomes. By application of HLM data, IVIVE estimated the area under the concentration versus time curve of midazolam to increase by 73 and 27% with CsA and Tac, respectively. The inhibitory effect was predominantly on the intestinal level, whereas hepatic intrinsic clearance seemed unaffected. DOI : 10.1124/dmd.111.043018
TACROLIMUS
Inhibition of human CYP3A4 expressed in supersomes assessed inhibition of 1'-OH midazolam formation by LC-MS method
—
IC50 = 620.0 nM
Reference
Title : Cyclosporine A- and tacrolimus-mediated inhibition of CYP3A4 and CYP3A5 in vitro. Authors : Amundsen R, Åsberg A, Ohm IK, Christensen H. Publication Year :
2012 Journal : Drug Metab. Dispos. Abstract : Cyclosporine A (CsA) and tacrolimus (Tac) are immunosuppressive drugs used in the majority of patients with solid organ transplants, generally in combination with a wide range of drugs. CsA and Tac seem not only to be substrates of CYP3A but have also been described as inhibitors of CYP3A. For CsA, in particular, inhibition of CYP3A has been suggested as the main mechanism of interactions seen clinically with various drugs. The aim of this study was to investigate the inhibitory effect and inhibition characteristics of CsA and Tac on CYP3A4 and CYP3A5 in vitro and to evaluate its clinical relevance. Inhibition by CsA and Tac was studied using midazolam as the probe substrate in coincubation and preincubation investigations using human liver microsomes (HLMs) as well as specific CYP3A4- and CYP3A5-expressing insect microsomes (Supersomes). In vitro-in vivo extrapolations (IVIVEs) were performed to evaluate the clinical relevance of the inhibition. Both CsA and Tac competitively inhibited CYP3A in HLMs, showing inhibition constants (K(i)) of 0.98 and 0.61 μM, respectively. Experiments in Supersomes revealed that Tac inhibited both CYP3A4 and CYP3A5, whereas CsA only inhibited CYP3A4. In contrast to the HLM experiments, studies in Supersomes showed inhibition by Tac to be NADPH- and time-dependent, with a 5-fold reduction in IC(50) after preincubation, supporting a time-dependent inhibition mechanism in recombinant microsomes. By application of HLM data, IVIVE estimated the area under the concentration versus time curve of midazolam to increase by 73 and 27% with CsA and Tac, respectively. The inhibitory effect was predominantly on the intestinal level, whereas hepatic intrinsic clearance seemed unaffected. DOI : 10.1124/dmd.111.043018
TACROLIMUS
Inhibition of human CYP3A4 expressed in supersomes assessed inhibition of 1'-OH midazolam formation preincubated with compound before substrate addition by LC-MS method
—
IC50 = 120.0 nM
Reference
Title : Cyclosporine A- and tacrolimus-mediated inhibition of CYP3A4 and CYP3A5 in vitro. Authors : Amundsen R, Åsberg A, Ohm IK, Christensen H. Publication Year :
2012 Journal : Drug Metab. Dispos. Abstract : Cyclosporine A (CsA) and tacrolimus (Tac) are immunosuppressive drugs used in the majority of patients with solid organ transplants, generally in combination with a wide range of drugs. CsA and Tac seem not only to be substrates of CYP3A but have also been described as inhibitors of CYP3A. For CsA, in particular, inhibition of CYP3A has been suggested as the main mechanism of interactions seen clinically with various drugs. The aim of this study was to investigate the inhibitory effect and inhibition characteristics of CsA and Tac on CYP3A4 and CYP3A5 in vitro and to evaluate its clinical relevance. Inhibition by CsA and Tac was studied using midazolam as the probe substrate in coincubation and preincubation investigations using human liver microsomes (HLMs) as well as specific CYP3A4- and CYP3A5-expressing insect microsomes (Supersomes). In vitro-in vivo extrapolations (IVIVEs) were performed to evaluate the clinical relevance of the inhibition. Both CsA and Tac competitively inhibited CYP3A in HLMs, showing inhibition constants (K(i)) of 0.98 and 0.61 μM, respectively. Experiments in Supersomes revealed that Tac inhibited both CYP3A4 and CYP3A5, whereas CsA only inhibited CYP3A4. In contrast to the HLM experiments, studies in Supersomes showed inhibition by Tac to be NADPH- and time-dependent, with a 5-fold reduction in IC(50) after preincubation, supporting a time-dependent inhibition mechanism in recombinant microsomes. By application of HLM data, IVIVE estimated the area under the concentration versus time curve of midazolam to increase by 73 and 27% with CsA and Tac, respectively. The inhibitory effect was predominantly on the intestinal level, whereas hepatic intrinsic clearance seemed unaffected. DOI : 10.1124/dmd.111.043018
TESTOSTERONE
Activity of human recombinant CYP3A4 assessed as conversion of testosterone to 6-beta-hydroxytestesterone at 25 uM after 60 mins in presence of 1 uM Miconazole
—
Inhibition = 41.1 %
Reference
Title : Metabolic stereoselectivity of cytochrome P450 3A4 towards deoxypodophyllotoxin: In silico predictions and experimental validation. Authors : Julsing MK, Vasilev NP, Schneidman-Duhovny D, Muntendam R, Woerdenbag HJ, Quax WJ, Wolfson HJ, Ionkova I, Kayser O. Publication Year :
2008 Journal : Eur. J. Med. Chem. Abstract : Deoxypodophyllotoxin is stereoselectively converted into epipodophyllotoxin by recombinant human cytochrome P450 3A4 (CYP3A4). Further kinetic analysis revealed that the Michaelis-Menten K(m) and V(max) for hydroxylation of deoxypodophyllotoxin by CYP3A4 at C7 position were 1.93 microM and 1.48 nmol/min/nmol, respectively. Deoxypodophyllotoxin was subjected to automated docking analysis in order to get better knowledge of the interaction between the CYP3A4 enzyme and the substrate, using the PatchDock algorithm with distance constraints. Automated docking showed that the beta-hydrogen atom at C7 position is in the most appropriate binding orientation at the site of oxidation. The docking results are consistent with the experimental data for the bioconversion of deoxypodophyllotoxin into epipodophyllotoxin by CYP3A4. In addition, the effects of five lignans, deoxypodophyllotoxin, epipodophyllotoxin, podophyllotoxin, demethylenedeoxypodophyllotoxin, and demethylenepodophyllotoxin, on CYP3A4 were compared in order to investigate the influence of the methylenedioxy group on the biotransformation process, to give insight into the mode of metabolization and to explain inhibitory activity of lignans. DOI : 10.1016/j.ejmech.2007.09.005
TESTOSTERONE
Activity of human recombinant CYP3A4 assessed as conversion of testosterone to 6-beta-hydroxytestesterone at 25 uM after 60 mins in presence of 10 uM Miconazole
—
Inhibition = 10.5 %
Reference
Title : Metabolic stereoselectivity of cytochrome P450 3A4 towards deoxypodophyllotoxin: In silico predictions and experimental validation. Authors : Julsing MK, Vasilev NP, Schneidman-Duhovny D, Muntendam R, Woerdenbag HJ, Quax WJ, Wolfson HJ, Ionkova I, Kayser O. Publication Year :
2008 Journal : Eur. J. Med. Chem. Abstract : Deoxypodophyllotoxin is stereoselectively converted into epipodophyllotoxin by recombinant human cytochrome P450 3A4 (CYP3A4). Further kinetic analysis revealed that the Michaelis-Menten K(m) and V(max) for hydroxylation of deoxypodophyllotoxin by CYP3A4 at C7 position were 1.93 microM and 1.48 nmol/min/nmol, respectively. Deoxypodophyllotoxin was subjected to automated docking analysis in order to get better knowledge of the interaction between the CYP3A4 enzyme and the substrate, using the PatchDock algorithm with distance constraints. Automated docking showed that the beta-hydrogen atom at C7 position is in the most appropriate binding orientation at the site of oxidation. The docking results are consistent with the experimental data for the bioconversion of deoxypodophyllotoxin into epipodophyllotoxin by CYP3A4. In addition, the effects of five lignans, deoxypodophyllotoxin, epipodophyllotoxin, podophyllotoxin, demethylenedeoxypodophyllotoxin, and demethylenepodophyllotoxin, on CYP3A4 were compared in order to investigate the influence of the methylenedioxy group on the biotransformation process, to give insight into the mode of metabolization and to explain inhibitory activity of lignans. DOI : 10.1016/j.ejmech.2007.09.005
TESTOSTERONE
Activity of human recombinant CYP3A4 assessed as conversion of testosterone to 6-beta-hydroxytestesterone at 25 uM after 60 mins in presence of 25 uM Miconazole
—
Inhibition = 4.38 %
Reference
Title : Metabolic stereoselectivity of cytochrome P450 3A4 towards deoxypodophyllotoxin: In silico predictions and experimental validation. Authors : Julsing MK, Vasilev NP, Schneidman-Duhovny D, Muntendam R, Woerdenbag HJ, Quax WJ, Wolfson HJ, Ionkova I, Kayser O. Publication Year :
2008 Journal : Eur. J. Med. Chem. Abstract : Deoxypodophyllotoxin is stereoselectively converted into epipodophyllotoxin by recombinant human cytochrome P450 3A4 (CYP3A4). Further kinetic analysis revealed that the Michaelis-Menten K(m) and V(max) for hydroxylation of deoxypodophyllotoxin by CYP3A4 at C7 position were 1.93 microM and 1.48 nmol/min/nmol, respectively. Deoxypodophyllotoxin was subjected to automated docking analysis in order to get better knowledge of the interaction between the CYP3A4 enzyme and the substrate, using the PatchDock algorithm with distance constraints. Automated docking showed that the beta-hydrogen atom at C7 position is in the most appropriate binding orientation at the site of oxidation. The docking results are consistent with the experimental data for the bioconversion of deoxypodophyllotoxin into epipodophyllotoxin by CYP3A4. In addition, the effects of five lignans, deoxypodophyllotoxin, epipodophyllotoxin, podophyllotoxin, demethylenedeoxypodophyllotoxin, and demethylenepodophyllotoxin, on CYP3A4 were compared in order to investigate the influence of the methylenedioxy group on the biotransformation process, to give insight into the mode of metabolization and to explain inhibitory activity of lignans. DOI : 10.1016/j.ejmech.2007.09.005
TESTOSTERONE
Activity of human recombinant CYP3A4 assessed as conversion of testosterone to 6-beta-hydroxytestesterone at 25 uM after 60 mins in presence of 1 uM deoxypodophyllotoxin
—
Inhibition = 61.0 %
Reference
Title : Metabolic stereoselectivity of cytochrome P450 3A4 towards deoxypodophyllotoxin: In silico predictions and experimental validation. Authors : Julsing MK, Vasilev NP, Schneidman-Duhovny D, Muntendam R, Woerdenbag HJ, Quax WJ, Wolfson HJ, Ionkova I, Kayser O. Publication Year :
2008 Journal : Eur. J. Med. Chem. Abstract : Deoxypodophyllotoxin is stereoselectively converted into epipodophyllotoxin by recombinant human cytochrome P450 3A4 (CYP3A4). Further kinetic analysis revealed that the Michaelis-Menten K(m) and V(max) for hydroxylation of deoxypodophyllotoxin by CYP3A4 at C7 position were 1.93 microM and 1.48 nmol/min/nmol, respectively. Deoxypodophyllotoxin was subjected to automated docking analysis in order to get better knowledge of the interaction between the CYP3A4 enzyme and the substrate, using the PatchDock algorithm with distance constraints. Automated docking showed that the beta-hydrogen atom at C7 position is in the most appropriate binding orientation at the site of oxidation. The docking results are consistent with the experimental data for the bioconversion of deoxypodophyllotoxin into epipodophyllotoxin by CYP3A4. In addition, the effects of five lignans, deoxypodophyllotoxin, epipodophyllotoxin, podophyllotoxin, demethylenedeoxypodophyllotoxin, and demethylenepodophyllotoxin, on CYP3A4 were compared in order to investigate the influence of the methylenedioxy group on the biotransformation process, to give insight into the mode of metabolization and to explain inhibitory activity of lignans. DOI : 10.1016/j.ejmech.2007.09.005
TESTOSTERONE
Activity of human recombinant CYP3A4 assessed as conversion of testosterone to 6-beta-hydroxytestesterone at 25 uM after 60 mins in presence of 10 uM deoxypodophyllotoxin
—
Inhibition = 10.4 %
Reference
Title : Metabolic stereoselectivity of cytochrome P450 3A4 towards deoxypodophyllotoxin: In silico predictions and experimental validation. Authors : Julsing MK, Vasilev NP, Schneidman-Duhovny D, Muntendam R, Woerdenbag HJ, Quax WJ, Wolfson HJ, Ionkova I, Kayser O. Publication Year :
2008 Journal : Eur. J. Med. Chem. Abstract : Deoxypodophyllotoxin is stereoselectively converted into epipodophyllotoxin by recombinant human cytochrome P450 3A4 (CYP3A4). Further kinetic analysis revealed that the Michaelis-Menten K(m) and V(max) for hydroxylation of deoxypodophyllotoxin by CYP3A4 at C7 position were 1.93 microM and 1.48 nmol/min/nmol, respectively. Deoxypodophyllotoxin was subjected to automated docking analysis in order to get better knowledge of the interaction between the CYP3A4 enzyme and the substrate, using the PatchDock algorithm with distance constraints. Automated docking showed that the beta-hydrogen atom at C7 position is in the most appropriate binding orientation at the site of oxidation. The docking results are consistent with the experimental data for the bioconversion of deoxypodophyllotoxin into epipodophyllotoxin by CYP3A4. In addition, the effects of five lignans, deoxypodophyllotoxin, epipodophyllotoxin, podophyllotoxin, demethylenedeoxypodophyllotoxin, and demethylenepodophyllotoxin, on CYP3A4 were compared in order to investigate the influence of the methylenedioxy group on the biotransformation process, to give insight into the mode of metabolization and to explain inhibitory activity of lignans. DOI : 10.1016/j.ejmech.2007.09.005
TESTOSTERONE
Activity of human recombinant CYP3A4 assessed as conversion of testosterone to 6-beta-hydroxytestesterone at 25 uM after 60 mins in presence of 25 uM deoxypodophyllotoxin
—
Inhibition = 4.85 %
Reference
Title : Metabolic stereoselectivity of cytochrome P450 3A4 towards deoxypodophyllotoxin: In silico predictions and experimental validation. Authors : Julsing MK, Vasilev NP, Schneidman-Duhovny D, Muntendam R, Woerdenbag HJ, Quax WJ, Wolfson HJ, Ionkova I, Kayser O. Publication Year :
2008 Journal : Eur. J. Med. Chem. Abstract : Deoxypodophyllotoxin is stereoselectively converted into epipodophyllotoxin by recombinant human cytochrome P450 3A4 (CYP3A4). Further kinetic analysis revealed that the Michaelis-Menten K(m) and V(max) for hydroxylation of deoxypodophyllotoxin by CYP3A4 at C7 position were 1.93 microM and 1.48 nmol/min/nmol, respectively. Deoxypodophyllotoxin was subjected to automated docking analysis in order to get better knowledge of the interaction between the CYP3A4 enzyme and the substrate, using the PatchDock algorithm with distance constraints. Automated docking showed that the beta-hydrogen atom at C7 position is in the most appropriate binding orientation at the site of oxidation. The docking results are consistent with the experimental data for the bioconversion of deoxypodophyllotoxin into epipodophyllotoxin by CYP3A4. In addition, the effects of five lignans, deoxypodophyllotoxin, epipodophyllotoxin, podophyllotoxin, demethylenedeoxypodophyllotoxin, and demethylenepodophyllotoxin, on CYP3A4 were compared in order to investigate the influence of the methylenedioxy group on the biotransformation process, to give insight into the mode of metabolization and to explain inhibitory activity of lignans. DOI : 10.1016/j.ejmech.2007.09.005
TESTOSTERONE
Activity of human recombinant CYP3A4 assessed as conversion of testosterone to 6-beta-hydroxytestesterone at 25 uM after 60 mins in presence of 1 uM epipodophyllotoxin
—
Inhibition = 69.4 %
Reference
Title : Metabolic stereoselectivity of cytochrome P450 3A4 towards deoxypodophyllotoxin: In silico predictions and experimental validation. Authors : Julsing MK, Vasilev NP, Schneidman-Duhovny D, Muntendam R, Woerdenbag HJ, Quax WJ, Wolfson HJ, Ionkova I, Kayser O. Publication Year :
2008 Journal : Eur. J. Med. Chem. Abstract : Deoxypodophyllotoxin is stereoselectively converted into epipodophyllotoxin by recombinant human cytochrome P450 3A4 (CYP3A4). Further kinetic analysis revealed that the Michaelis-Menten K(m) and V(max) for hydroxylation of deoxypodophyllotoxin by CYP3A4 at C7 position were 1.93 microM and 1.48 nmol/min/nmol, respectively. Deoxypodophyllotoxin was subjected to automated docking analysis in order to get better knowledge of the interaction between the CYP3A4 enzyme and the substrate, using the PatchDock algorithm with distance constraints. Automated docking showed that the beta-hydrogen atom at C7 position is in the most appropriate binding orientation at the site of oxidation. The docking results are consistent with the experimental data for the bioconversion of deoxypodophyllotoxin into epipodophyllotoxin by CYP3A4. In addition, the effects of five lignans, deoxypodophyllotoxin, epipodophyllotoxin, podophyllotoxin, demethylenedeoxypodophyllotoxin, and demethylenepodophyllotoxin, on CYP3A4 were compared in order to investigate the influence of the methylenedioxy group on the biotransformation process, to give insight into the mode of metabolization and to explain inhibitory activity of lignans. DOI : 10.1016/j.ejmech.2007.09.005
TESTOSTERONE
Activity of human recombinant CYP3A4 assessed as conversion of testosterone to 6-beta-hydroxytestesterone at 25 uM after 60 mins in presence of 10 uM epipodophyllotoxin
—
Inhibition = 7.96 %
Reference
Title : Metabolic stereoselectivity of cytochrome P450 3A4 towards deoxypodophyllotoxin: In silico predictions and experimental validation. Authors : Julsing MK, Vasilev NP, Schneidman-Duhovny D, Muntendam R, Woerdenbag HJ, Quax WJ, Wolfson HJ, Ionkova I, Kayser O. Publication Year :
2008 Journal : Eur. J. Med. Chem. Abstract : Deoxypodophyllotoxin is stereoselectively converted into epipodophyllotoxin by recombinant human cytochrome P450 3A4 (CYP3A4). Further kinetic analysis revealed that the Michaelis-Menten K(m) and V(max) for hydroxylation of deoxypodophyllotoxin by CYP3A4 at C7 position were 1.93 microM and 1.48 nmol/min/nmol, respectively. Deoxypodophyllotoxin was subjected to automated docking analysis in order to get better knowledge of the interaction between the CYP3A4 enzyme and the substrate, using the PatchDock algorithm with distance constraints. Automated docking showed that the beta-hydrogen atom at C7 position is in the most appropriate binding orientation at the site of oxidation. The docking results are consistent with the experimental data for the bioconversion of deoxypodophyllotoxin into epipodophyllotoxin by CYP3A4. In addition, the effects of five lignans, deoxypodophyllotoxin, epipodophyllotoxin, podophyllotoxin, demethylenedeoxypodophyllotoxin, and demethylenepodophyllotoxin, on CYP3A4 were compared in order to investigate the influence of the methylenedioxy group on the biotransformation process, to give insight into the mode of metabolization and to explain inhibitory activity of lignans. DOI : 10.1016/j.ejmech.2007.09.005
TESTOSTERONE
Activity of human recombinant CYP3A4 assessed as conversion of testosterone to 6-beta-hydroxytestesterone at 25 uM after 60 mins in presence of 25 uM epipodophyllotoxin
—
Inhibition = 11.46 %
Reference
Title : Metabolic stereoselectivity of cytochrome P450 3A4 towards deoxypodophyllotoxin: In silico predictions and experimental validation. Authors : Julsing MK, Vasilev NP, Schneidman-Duhovny D, Muntendam R, Woerdenbag HJ, Quax WJ, Wolfson HJ, Ionkova I, Kayser O. Publication Year :
2008 Journal : Eur. J. Med. Chem. Abstract : Deoxypodophyllotoxin is stereoselectively converted into epipodophyllotoxin by recombinant human cytochrome P450 3A4 (CYP3A4). Further kinetic analysis revealed that the Michaelis-Menten K(m) and V(max) for hydroxylation of deoxypodophyllotoxin by CYP3A4 at C7 position were 1.93 microM and 1.48 nmol/min/nmol, respectively. Deoxypodophyllotoxin was subjected to automated docking analysis in order to get better knowledge of the interaction between the CYP3A4 enzyme and the substrate, using the PatchDock algorithm with distance constraints. Automated docking showed that the beta-hydrogen atom at C7 position is in the most appropriate binding orientation at the site of oxidation. The docking results are consistent with the experimental data for the bioconversion of deoxypodophyllotoxin into epipodophyllotoxin by CYP3A4. In addition, the effects of five lignans, deoxypodophyllotoxin, epipodophyllotoxin, podophyllotoxin, demethylenedeoxypodophyllotoxin, and demethylenepodophyllotoxin, on CYP3A4 were compared in order to investigate the influence of the methylenedioxy group on the biotransformation process, to give insight into the mode of metabolization and to explain inhibitory activity of lignans. DOI : 10.1016/j.ejmech.2007.09.005
TESTOSTERONE
Activity of human recombinant CYP3A4 assessed as conversion of testosterone to 6-beta-hydroxytestesterone at 25 uM after 60 mins in presence of 1 uM podophyllotoxin
—
Inhibition = 44.1 %
Reference
Title : Metabolic stereoselectivity of cytochrome P450 3A4 towards deoxypodophyllotoxin: In silico predictions and experimental validation. Authors : Julsing MK, Vasilev NP, Schneidman-Duhovny D, Muntendam R, Woerdenbag HJ, Quax WJ, Wolfson HJ, Ionkova I, Kayser O. Publication Year :
2008 Journal : Eur. J. Med. Chem. Abstract : Deoxypodophyllotoxin is stereoselectively converted into epipodophyllotoxin by recombinant human cytochrome P450 3A4 (CYP3A4). Further kinetic analysis revealed that the Michaelis-Menten K(m) and V(max) for hydroxylation of deoxypodophyllotoxin by CYP3A4 at C7 position were 1.93 microM and 1.48 nmol/min/nmol, respectively. Deoxypodophyllotoxin was subjected to automated docking analysis in order to get better knowledge of the interaction between the CYP3A4 enzyme and the substrate, using the PatchDock algorithm with distance constraints. Automated docking showed that the beta-hydrogen atom at C7 position is in the most appropriate binding orientation at the site of oxidation. The docking results are consistent with the experimental data for the bioconversion of deoxypodophyllotoxin into epipodophyllotoxin by CYP3A4. In addition, the effects of five lignans, deoxypodophyllotoxin, epipodophyllotoxin, podophyllotoxin, demethylenedeoxypodophyllotoxin, and demethylenepodophyllotoxin, on CYP3A4 were compared in order to investigate the influence of the methylenedioxy group on the biotransformation process, to give insight into the mode of metabolization and to explain inhibitory activity of lignans. DOI : 10.1016/j.ejmech.2007.09.005
TESTOSTERONE
Activity of human recombinant CYP3A4 assessed as conversion of testosterone to 6-beta-hydroxytestesterone at 25 uM after 60 mins in presence of 10 uM podophyllotoxin
—
Inhibition = 7.82 %
Reference
Title : Metabolic stereoselectivity of cytochrome P450 3A4 towards deoxypodophyllotoxin: In silico predictions and experimental validation. Authors : Julsing MK, Vasilev NP, Schneidman-Duhovny D, Muntendam R, Woerdenbag HJ, Quax WJ, Wolfson HJ, Ionkova I, Kayser O. Publication Year :
2008 Journal : Eur. J. Med. Chem. Abstract : Deoxypodophyllotoxin is stereoselectively converted into epipodophyllotoxin by recombinant human cytochrome P450 3A4 (CYP3A4). Further kinetic analysis revealed that the Michaelis-Menten K(m) and V(max) for hydroxylation of deoxypodophyllotoxin by CYP3A4 at C7 position were 1.93 microM and 1.48 nmol/min/nmol, respectively. Deoxypodophyllotoxin was subjected to automated docking analysis in order to get better knowledge of the interaction between the CYP3A4 enzyme and the substrate, using the PatchDock algorithm with distance constraints. Automated docking showed that the beta-hydrogen atom at C7 position is in the most appropriate binding orientation at the site of oxidation. The docking results are consistent with the experimental data for the bioconversion of deoxypodophyllotoxin into epipodophyllotoxin by CYP3A4. In addition, the effects of five lignans, deoxypodophyllotoxin, epipodophyllotoxin, podophyllotoxin, demethylenedeoxypodophyllotoxin, and demethylenepodophyllotoxin, on CYP3A4 were compared in order to investigate the influence of the methylenedioxy group on the biotransformation process, to give insight into the mode of metabolization and to explain inhibitory activity of lignans. DOI : 10.1016/j.ejmech.2007.09.005
TESTOSTERONE
Activity of human recombinant CYP3A4 assessed as conversion of testosterone to 6-beta-hydroxytestesterone at 25 uM after 60 mins in presence of 25 uM podophyllotoxin
—
Inhibition = 7.04 %
Reference
Title : Metabolic stereoselectivity of cytochrome P450 3A4 towards deoxypodophyllotoxin: In silico predictions and experimental validation. Authors : Julsing MK, Vasilev NP, Schneidman-Duhovny D, Muntendam R, Woerdenbag HJ, Quax WJ, Wolfson HJ, Ionkova I, Kayser O. Publication Year :
2008 Journal : Eur. J. Med. Chem. Abstract : Deoxypodophyllotoxin is stereoselectively converted into epipodophyllotoxin by recombinant human cytochrome P450 3A4 (CYP3A4). Further kinetic analysis revealed that the Michaelis-Menten K(m) and V(max) for hydroxylation of deoxypodophyllotoxin by CYP3A4 at C7 position were 1.93 microM and 1.48 nmol/min/nmol, respectively. Deoxypodophyllotoxin was subjected to automated docking analysis in order to get better knowledge of the interaction between the CYP3A4 enzyme and the substrate, using the PatchDock algorithm with distance constraints. Automated docking showed that the beta-hydrogen atom at C7 position is in the most appropriate binding orientation at the site of oxidation. The docking results are consistent with the experimental data for the bioconversion of deoxypodophyllotoxin into epipodophyllotoxin by CYP3A4. In addition, the effects of five lignans, deoxypodophyllotoxin, epipodophyllotoxin, podophyllotoxin, demethylenedeoxypodophyllotoxin, and demethylenepodophyllotoxin, on CYP3A4 were compared in order to investigate the influence of the methylenedioxy group on the biotransformation process, to give insight into the mode of metabolization and to explain inhibitory activity of lignans. DOI : 10.1016/j.ejmech.2007.09.005
TESTOSTERONE
Activity of human recombinant CYP3A4 assessed as conversion of testosterone to 6-beta-hydroxytestesterone at 25 uM after 60 mins in presence of 1 uM demethylene-deoxy-podophyllotoxin
—
Inhibition = 103.0 %
Reference
Title : Metabolic stereoselectivity of cytochrome P450 3A4 towards deoxypodophyllotoxin: In silico predictions and experimental validation. Authors : Julsing MK, Vasilev NP, Schneidman-Duhovny D, Muntendam R, Woerdenbag HJ, Quax WJ, Wolfson HJ, Ionkova I, Kayser O. Publication Year :
2008 Journal : Eur. J. Med. Chem. Abstract : Deoxypodophyllotoxin is stereoselectively converted into epipodophyllotoxin by recombinant human cytochrome P450 3A4 (CYP3A4). Further kinetic analysis revealed that the Michaelis-Menten K(m) and V(max) for hydroxylation of deoxypodophyllotoxin by CYP3A4 at C7 position were 1.93 microM and 1.48 nmol/min/nmol, respectively. Deoxypodophyllotoxin was subjected to automated docking analysis in order to get better knowledge of the interaction between the CYP3A4 enzyme and the substrate, using the PatchDock algorithm with distance constraints. Automated docking showed that the beta-hydrogen atom at C7 position is in the most appropriate binding orientation at the site of oxidation. The docking results are consistent with the experimental data for the bioconversion of deoxypodophyllotoxin into epipodophyllotoxin by CYP3A4. In addition, the effects of five lignans, deoxypodophyllotoxin, epipodophyllotoxin, podophyllotoxin, demethylenedeoxypodophyllotoxin, and demethylenepodophyllotoxin, on CYP3A4 were compared in order to investigate the influence of the methylenedioxy group on the biotransformation process, to give insight into the mode of metabolization and to explain inhibitory activity of lignans. DOI : 10.1016/j.ejmech.2007.09.005
TESTOSTERONE
Activity of human recombinant CYP3A4 assessed as conversion of testosterone to 6-beta-hydroxytestesterone at 25 uM after 60 mins in presence of 10 uM demethylene-deoxy-podophyllotoxin
—
Inhibition = 69.6 %
Reference
Title : Metabolic stereoselectivity of cytochrome P450 3A4 towards deoxypodophyllotoxin: In silico predictions and experimental validation. Authors : Julsing MK, Vasilev NP, Schneidman-Duhovny D, Muntendam R, Woerdenbag HJ, Quax WJ, Wolfson HJ, Ionkova I, Kayser O. Publication Year :
2008 Journal : Eur. J. Med. Chem. Abstract : Deoxypodophyllotoxin is stereoselectively converted into epipodophyllotoxin by recombinant human cytochrome P450 3A4 (CYP3A4). Further kinetic analysis revealed that the Michaelis-Menten K(m) and V(max) for hydroxylation of deoxypodophyllotoxin by CYP3A4 at C7 position were 1.93 microM and 1.48 nmol/min/nmol, respectively. Deoxypodophyllotoxin was subjected to automated docking analysis in order to get better knowledge of the interaction between the CYP3A4 enzyme and the substrate, using the PatchDock algorithm with distance constraints. Automated docking showed that the beta-hydrogen atom at C7 position is in the most appropriate binding orientation at the site of oxidation. The docking results are consistent with the experimental data for the bioconversion of deoxypodophyllotoxin into epipodophyllotoxin by CYP3A4. In addition, the effects of five lignans, deoxypodophyllotoxin, epipodophyllotoxin, podophyllotoxin, demethylenedeoxypodophyllotoxin, and demethylenepodophyllotoxin, on CYP3A4 were compared in order to investigate the influence of the methylenedioxy group on the biotransformation process, to give insight into the mode of metabolization and to explain inhibitory activity of lignans. DOI : 10.1016/j.ejmech.2007.09.005
TESTOSTERONE
Activity of human recombinant CYP3A4 assessed as conversion of testosterone to 6-beta-hydroxytestesterone at 25 uM after 60 mins in presence of 25 uM demethylene-deoxy-podophyllotoxin
—
Inhibition = 69.7 %
Reference
Title : Metabolic stereoselectivity of cytochrome P450 3A4 towards deoxypodophyllotoxin: In silico predictions and experimental validation. Authors : Julsing MK, Vasilev NP, Schneidman-Duhovny D, Muntendam R, Woerdenbag HJ, Quax WJ, Wolfson HJ, Ionkova I, Kayser O. Publication Year :
2008 Journal : Eur. J. Med. Chem. Abstract : Deoxypodophyllotoxin is stereoselectively converted into epipodophyllotoxin by recombinant human cytochrome P450 3A4 (CYP3A4). Further kinetic analysis revealed that the Michaelis-Menten K(m) and V(max) for hydroxylation of deoxypodophyllotoxin by CYP3A4 at C7 position were 1.93 microM and 1.48 nmol/min/nmol, respectively. Deoxypodophyllotoxin was subjected to automated docking analysis in order to get better knowledge of the interaction between the CYP3A4 enzyme and the substrate, using the PatchDock algorithm with distance constraints. Automated docking showed that the beta-hydrogen atom at C7 position is in the most appropriate binding orientation at the site of oxidation. The docking results are consistent with the experimental data for the bioconversion of deoxypodophyllotoxin into epipodophyllotoxin by CYP3A4. In addition, the effects of five lignans, deoxypodophyllotoxin, epipodophyllotoxin, podophyllotoxin, demethylenedeoxypodophyllotoxin, and demethylenepodophyllotoxin, on CYP3A4 were compared in order to investigate the influence of the methylenedioxy group on the biotransformation process, to give insight into the mode of metabolization and to explain inhibitory activity of lignans. DOI : 10.1016/j.ejmech.2007.09.005
TESTOSTERONE
Activity of human recombinant CYP3A4 assessed as conversion of testosterone to 6-beta-hydroxytestesterone at 25 uM after 60 mins in presence of 1 uM demethylenepodophyllotoxin
—
Inhibition = 95.0 %
Reference
Title : Metabolic stereoselectivity of cytochrome P450 3A4 towards deoxypodophyllotoxin: In silico predictions and experimental validation. Authors : Julsing MK, Vasilev NP, Schneidman-Duhovny D, Muntendam R, Woerdenbag HJ, Quax WJ, Wolfson HJ, Ionkova I, Kayser O. Publication Year :
2008 Journal : Eur. J. Med. Chem. Abstract : Deoxypodophyllotoxin is stereoselectively converted into epipodophyllotoxin by recombinant human cytochrome P450 3A4 (CYP3A4). Further kinetic analysis revealed that the Michaelis-Menten K(m) and V(max) for hydroxylation of deoxypodophyllotoxin by CYP3A4 at C7 position were 1.93 microM and 1.48 nmol/min/nmol, respectively. Deoxypodophyllotoxin was subjected to automated docking analysis in order to get better knowledge of the interaction between the CYP3A4 enzyme and the substrate, using the PatchDock algorithm with distance constraints. Automated docking showed that the beta-hydrogen atom at C7 position is in the most appropriate binding orientation at the site of oxidation. The docking results are consistent with the experimental data for the bioconversion of deoxypodophyllotoxin into epipodophyllotoxin by CYP3A4. In addition, the effects of five lignans, deoxypodophyllotoxin, epipodophyllotoxin, podophyllotoxin, demethylenedeoxypodophyllotoxin, and demethylenepodophyllotoxin, on CYP3A4 were compared in order to investigate the influence of the methylenedioxy group on the biotransformation process, to give insight into the mode of metabolization and to explain inhibitory activity of lignans. DOI : 10.1016/j.ejmech.2007.09.005
TESTOSTERONE
Activity of human recombinant CYP3A4 assessed as conversion of testosterone to 6-beta-hydroxytestesterone at 25 uM after 60 mins in presence of 10 uM demethylenepodophyllotoxin
—
Inhibition = 98.7 %
Reference
Title : Metabolic stereoselectivity of cytochrome P450 3A4 towards deoxypodophyllotoxin: In silico predictions and experimental validation. Authors : Julsing MK, Vasilev NP, Schneidman-Duhovny D, Muntendam R, Woerdenbag HJ, Quax WJ, Wolfson HJ, Ionkova I, Kayser O. Publication Year :
2008 Journal : Eur. J. Med. Chem. Abstract : Deoxypodophyllotoxin is stereoselectively converted into epipodophyllotoxin by recombinant human cytochrome P450 3A4 (CYP3A4). Further kinetic analysis revealed that the Michaelis-Menten K(m) and V(max) for hydroxylation of deoxypodophyllotoxin by CYP3A4 at C7 position were 1.93 microM and 1.48 nmol/min/nmol, respectively. Deoxypodophyllotoxin was subjected to automated docking analysis in order to get better knowledge of the interaction between the CYP3A4 enzyme and the substrate, using the PatchDock algorithm with distance constraints. Automated docking showed that the beta-hydrogen atom at C7 position is in the most appropriate binding orientation at the site of oxidation. The docking results are consistent with the experimental data for the bioconversion of deoxypodophyllotoxin into epipodophyllotoxin by CYP3A4. In addition, the effects of five lignans, deoxypodophyllotoxin, epipodophyllotoxin, podophyllotoxin, demethylenedeoxypodophyllotoxin, and demethylenepodophyllotoxin, on CYP3A4 were compared in order to investigate the influence of the methylenedioxy group on the biotransformation process, to give insight into the mode of metabolization and to explain inhibitory activity of lignans. DOI : 10.1016/j.ejmech.2007.09.005
TESTOSTERONE
Activity of human recombinant CYP3A4 assessed as conversion of testosterone to 6-beta-hydroxytestesterone at 25 uM after 60 mins in presence of 25 uM demethylenepodophyllotoxin
—
Inhibition = 97.1 %
Reference
Title : Metabolic stereoselectivity of cytochrome P450 3A4 towards deoxypodophyllotoxin: In silico predictions and experimental validation. Authors : Julsing MK, Vasilev NP, Schneidman-Duhovny D, Muntendam R, Woerdenbag HJ, Quax WJ, Wolfson HJ, Ionkova I, Kayser O. Publication Year :
2008 Journal : Eur. J. Med. Chem. Abstract : Deoxypodophyllotoxin is stereoselectively converted into epipodophyllotoxin by recombinant human cytochrome P450 3A4 (CYP3A4). Further kinetic analysis revealed that the Michaelis-Menten K(m) and V(max) for hydroxylation of deoxypodophyllotoxin by CYP3A4 at C7 position were 1.93 microM and 1.48 nmol/min/nmol, respectively. Deoxypodophyllotoxin was subjected to automated docking analysis in order to get better knowledge of the interaction between the CYP3A4 enzyme and the substrate, using the PatchDock algorithm with distance constraints. Automated docking showed that the beta-hydrogen atom at C7 position is in the most appropriate binding orientation at the site of oxidation. The docking results are consistent with the experimental data for the bioconversion of deoxypodophyllotoxin into epipodophyllotoxin by CYP3A4. In addition, the effects of five lignans, deoxypodophyllotoxin, epipodophyllotoxin, podophyllotoxin, demethylenedeoxypodophyllotoxin, and demethylenepodophyllotoxin, on CYP3A4 were compared in order to investigate the influence of the methylenedioxy group on the biotransformation process, to give insight into the mode of metabolization and to explain inhibitory activity of lignans. DOI : 10.1016/j.ejmech.2007.09.005
VALPROIC ACID
Time dependent inhibition of CYP3A4 (unknown origin) at 100 uM by LC/MS system
—
Inhibition < 10.0 %
Reference
Title : Combination of GSH trapping and time-dependent inhibition assays as a predictive method of drugs generating highly reactive metabolites. Authors : Nakayama S, Takakusa H, Watanabe A, Miyaji Y, Suzuki W, Sugiyama D, Shiosakai K, Honda K, Okudaira N, Izumi T, Okazaki O. Publication Year :
2011 Journal : Drug Metab. Dispos. Abstract : Covalent binding (CB) of reactive metabolites (RMs) is potentially involved in severe adverse drug reactions. Because the CB assay is of low throughput and costly, a qualitative trapping assay using agents such as [(35)S]GSH is often performed in the early stages of drug discovery. However, trapping methods alone cannot replace the CB assay. We hypothesized that the time-dependent inhibition (TDI) assay might be complementary to the [(35)S]GSH trapping assay in detecting RMs. We performed CB assays, [(35)S]GSH trapping assays, and TDI assays for 42 structurally diverse compounds. First, we showed that the [(35)S]GSH trapping assay alone does not correlate with the extent of CB. Four compounds that the [(35)S]GSH trapping assay failed to detect but that showed high extent of CB were inactivators of the enzyme in the TDI assay. There was a tendency for compounds judged as positive in the TDI assay to show a high degree of CB irrespective of the result of the [(35)S]GSH trapping assay. Finally, to combine parameters from the two assays, we introduced intrinsic clearance to describe the formation of RMs (CL(int, RMs)). The Spearman rank correlation coefficient between the extent of CB and CL(int, RMs) was 0.77 (p < 0.0001), which was better than that for the formation rates of [(35)S]GSH adducts. Therefore, we demonstrated that a combination of the [(35)S]GSH trapping and TDI assays is an effective method for detecting compounds potentially capable of generating highly reactive metabolites in the early stages of drug discovery. DOI : 10.1124/dmd.111.039180
VALSARTAN
Time dependent inhibition of CYP3A4 (unknown origin) at 100 uM by LC/MS system
—
Inhibition < 10.0 %
Reference
Title : Combination of GSH trapping and time-dependent inhibition assays as a predictive method of drugs generating highly reactive metabolites. Authors : Nakayama S, Takakusa H, Watanabe A, Miyaji Y, Suzuki W, Sugiyama D, Shiosakai K, Honda K, Okudaira N, Izumi T, Okazaki O. Publication Year :
2011 Journal : Drug Metab. Dispos. Abstract : Covalent binding (CB) of reactive metabolites (RMs) is potentially involved in severe adverse drug reactions. Because the CB assay is of low throughput and costly, a qualitative trapping assay using agents such as [(35)S]GSH is often performed in the early stages of drug discovery. However, trapping methods alone cannot replace the CB assay. We hypothesized that the time-dependent inhibition (TDI) assay might be complementary to the [(35)S]GSH trapping assay in detecting RMs. We performed CB assays, [(35)S]GSH trapping assays, and TDI assays for 42 structurally diverse compounds. First, we showed that the [(35)S]GSH trapping assay alone does not correlate with the extent of CB. Four compounds that the [(35)S]GSH trapping assay failed to detect but that showed high extent of CB were inactivators of the enzyme in the TDI assay. There was a tendency for compounds judged as positive in the TDI assay to show a high degree of CB irrespective of the result of the [(35)S]GSH trapping assay. Finally, to combine parameters from the two assays, we introduced intrinsic clearance to describe the formation of RMs (CL(int, RMs)). The Spearman rank correlation coefficient between the extent of CB and CL(int, RMs) was 0.77 (p < 0.0001), which was better than that for the formation rates of [(35)S]GSH adducts. Therefore, we demonstrated that a combination of the [(35)S]GSH trapping and TDI assays is an effective method for detecting compounds potentially capable of generating highly reactive metabolites in the early stages of drug discovery. DOI : 10.1124/dmd.111.039180
VORINOSTAT
Inhibition of human CYP3A4 using 7-benzyloxy-4-(trifluoromethyl)-coumarin as substrate at 10 uM after 30 mins by fluorimetric analysis
—
Inhibition = 35.0 %
Reference
Title : Synthesis, biological characterization and molecular modeling insights of spirochromanes as potent HDAC inhibitors. Authors : Thaler F, Moretti L, Amici R, Abate A, Colombo A, Carenzi G, Fulco MC, Boggio R, Dondio G, Gagliardi S, Minucci S, Sartori L, Varasi M, Mercurio C. Publication Year :
2016 Journal : Eur. J. Med. Chem. Abstract : In the last decades, inhibitors of histone deacetylases (HDAC) have become an important class of anti-cancer agents. In a previous study we described the synthesis of spiro[chromane-2,4'-piperidine]hydroxamic acid derivatives able to inhibit histone deacetylase enzymes. Herein, we present our exploration for new derivatives by replacing the piperidine moiety with various cycloamines. The goal was to obtain highly potent compounds with a good in vitro ADME profile. In addition, molecular modeling studies unravelled the binding mode of these inhibitors. DOI : 10.1016/j.ejmech.2015.11.010
ZAFIRLUKAST
Time dependent inhibition of CYP3A4 (unknown origin) at 10 uM by LC/MS system
—
Inhibition = 43.0 %
Reference
Title : Combination of GSH trapping and time-dependent inhibition assays as a predictive method of drugs generating highly reactive metabolites. Authors : Nakayama S, Takakusa H, Watanabe A, Miyaji Y, Suzuki W, Sugiyama D, Shiosakai K, Honda K, Okudaira N, Izumi T, Okazaki O. Publication Year :
2011 Journal : Drug Metab. Dispos. Abstract : Covalent binding (CB) of reactive metabolites (RMs) is potentially involved in severe adverse drug reactions. Because the CB assay is of low throughput and costly, a qualitative trapping assay using agents such as [(35)S]GSH is often performed in the early stages of drug discovery. However, trapping methods alone cannot replace the CB assay. We hypothesized that the time-dependent inhibition (TDI) assay might be complementary to the [(35)S]GSH trapping assay in detecting RMs. We performed CB assays, [(35)S]GSH trapping assays, and TDI assays for 42 structurally diverse compounds. First, we showed that the [(35)S]GSH trapping assay alone does not correlate with the extent of CB. Four compounds that the [(35)S]GSH trapping assay failed to detect but that showed high extent of CB were inactivators of the enzyme in the TDI assay. There was a tendency for compounds judged as positive in the TDI assay to show a high degree of CB irrespective of the result of the [(35)S]GSH trapping assay. Finally, to combine parameters from the two assays, we introduced intrinsic clearance to describe the formation of RMs (CL(int, RMs)). The Spearman rank correlation coefficient between the extent of CB and CL(int, RMs) was 0.77 (p < 0.0001), which was better than that for the formation rates of [(35)S]GSH adducts. Therefore, we demonstrated that a combination of the [(35)S]GSH trapping and TDI assays is an effective method for detecting compounds potentially capable of generating highly reactive metabolites in the early stages of drug discovery. DOI : 10.1124/dmd.111.039180
ALISKIREN
Inhibitory activity of 200 gM SPP100 on cDNA-derived CYP3A4 enzyme in microsome prepared from human lymphoblastoid cell line
—
Inhibition = 11.0 %
Reference
Title : FDA drug approval package for Aliskiren hemifumarate Authors : None Publication Year :
None Journal : None Abstract : Drug Approval Package documents downloaded from Drugs@FDA. Data extracted from various pdfs.
- hERG study and start of ADME section: http://www.accessdata.fda.gov/drugsatfda_docs/nda/2007/021985s000_PharmR_P1.pdf
- ADME/PK studies, metabolic schemes: http://www.accessdata.fda.gov/drugsatfda_docs/nda/2007/021985s000_PharmR_P2.pdf
- Marmoset toxicity study including PK parameters: http://www.accessdata.fda.gov/drugsatfda_docs/nda/2007/021985s000_PharmR_P3.pdf
- Metabolism studies and human PK: http://www.accessdata.fda.gov/drugsatfda_docs/nda/2007/021985s000_ClinPharmR_P1.pdf
- human metabolic scheme: http://www.accessdata.fda.gov/drugsatfda_docs/nda/2007/021985s000_ClinPharmR_P2.pdf
- human metabolic scheme: http://www.accessdata.fda.gov/drugsatfda_docs/nda/2007/021985s000_ClinPharmR_P3.pdf
ALOSETRON
Inhibition of human CYP3A4
—
IC50 = 600.0 nM
Reference
Title : Discovery of 2-substituted benzoxazole carboxamides as 5-HT3 receptor antagonists. Authors : Yang Z, Fairfax DJ, Maeng JH, Masih L, Usyatinsky A, Hassler C, Isaacson S, Fitzpatrick K, DeOrazio RJ, Chen J, Harding JP, Isherwood M, Dobritsa S, Christensen KL, Wierschke JD, Bliss BI, Peterson LH, Beer CM, Cioffi C, Lynch M, Rennells WM, Richards JJ, Rust T, Khmelnitsky YL, Cohen ML, Manning DD. Publication Year :
2010 Journal : Bioorg. Med. Chem. Lett. Abstract : A new class of 2-substituted benzoxazole carboxamides are presented as potent functional 5-HT(3) receptor antagonists. The chemical series possesses nanomolar in vitro activity against human 5-HT(3)A receptors. A chemistry optimization program was conducted and identified 2-aminobenzoxazoles as orally active 5-HT(3) receptor antagonists with good metabolic stability. These novel analogues possess drug-like characteristics and have potential utility for the treatment of diseases attributable to improper 5-HT(3) receptor function, especially diarrhea predominant irritable bowel syndrome (IBS-D). DOI : 10.1016/j.bmcl.2010.09.038
ALOSETRON
Inhibition of human CYP3A4
—
IC50 = 600.0 nM
Reference
Title : Novel serotonin type 3 receptor partial agonists for the potential treatment of irritable bowel syndrome. Authors : Manning DD, Cioffi CL, Usyatinsky A, Fitzpatrick K, Masih L, Guo C, Zhang Z, Choo SH, Sikkander MI, Ryan KN, Naginskaya J, Hassler C, Dobritsa S, Wierschke JD, Earley WG, Butler AS, Brady CA, Barnes NM, Cohen ML, Guzzo PR. Publication Year :
2011 Journal : Bioorg. Med. Chem. Lett. Abstract : Serotonin type 3 (5-HT(3)) receptor partial agonists are being targeted as potential new drugs for the treatment of irritable bowel syndrome (IBS). Two new chemical series bearing indazole and indole cores have exhibited nanomolar binding affinity for the h5-HT(3)A receptor. A range of partial agonist activities in HEK cells heterologously expressing the h5-HT(3)A receptor were measured for the indazole series. Excellent 5-HT(3) receptor selectivity, favorable in vitro metabolic stability and CYP inhibition properties, and good oral in vivo potency in the murine von Bezold-Jarisch reflex model is exemplified thereby indicating the series to have potential utility as improved IBS agents. DOI : 10.1016/j.bmcl.2010.11.080
ALOSETRON
Inhibition of CYP3A4 (unknown origin)
—
IC50 = 600.0 nM
Reference
Title : The discovery of diazepinone-based 5-HT3 receptor partial agonists. Authors : Manning DD, Guo C, Zhang Z, Ryan KN, Naginskaya J, Choo SH, Masih L, Earley WG, Wierschke JD, Newman AS, Brady CA, Barnes NM, Guzzo PR. Publication Year :
2014 Journal : Bioorg. Med. Chem. Lett. Abstract : Serotonin type 3 (5-HT3) receptor partial agonists have been targeted as potential new drugs for the symptomatic relief of irritable bowel syndrome (IBS). Multiple diazepinone-based compounds have been discovered, which exhibit nanomolar binding affinity for the h5-HT3A receptor and display a range of intrinsic activities (IA=7-87% of 5-HT Emax) in HEK cells heterologously expressing the h5-HT3A receptor. Favorable physicochemical properties and in vitro ADME profile coupled with oral activity in the murine von Bezold-Jarisch reflex model demonstrates the series has promise for producing low to moderate IA partial agonists suitable for an IBS indication. DOI : 10.1016/j.bmcl.2014.03.074
AMIODARONE
Inhibition of CYP3A4-mediated midazolam metabolite formation in human liver microsomes at 5 uM after 8 mins by LC-MS/MS analysis relative to control
Liver
Inhibition = -18.4 %
Reference
Title : Identifying a selective substrate and inhibitor pair for the evaluation of CYP2J2 activity. Authors : Lee CA, Jones JP, Katayama J, Kaspera R, Jiang Y, Freiwald S, Smith E, Walker GS, Totah RA. Publication Year :
2012 Journal : Drug Metab. Dispos. Abstract : CYP2J2, an arachidonic acid epoxygenase, is recognized for its role in the first-pass metabolism of astemizole and ebastine. To fully assess the role of CYP2J2 in drug metabolism, a selective substrate and potent specific chemical inhibitor are essential. In this study, we report amiodarone 4-hydoxylation as a specific CYP2J2-catalyzed reaction with no CYP3A4, or other drug-metabolizing enzyme, involvement. Amiodarone 4-hydroxylation enabled the determination of liver relative activity factor and intersystem extrapolation factor for CYP2J2. Amiodarone 4-hydroxylation correlated with astemizole O-demethylation but not with CYP2J2 protein content in a sample of human liver microsomes. To identify a specific CYP2J2 inhibitor, 138 drugs were screened using terfenadine and astemizole as probe substrates with recombinant CYP2J2. Forty-two drugs inhibited CYP2J2 activity by ≥50% at 30 μM, but inhibition was substrate-dependent. Of these, danazol was a potent inhibitor of both hydroxylation of terfenadine (IC(50) = 77 nM) and O-demethylation of astemizole (K(i) = 20 nM), and inhibition was mostly competitive. Danazol inhibited CYP2C9, CYP2C8, and CYP2D6 with IC(50) values of 1.44, 1.95, and 2.74 μM, respectively. Amiodarone or astemizole were included in a seven-probe cocktail for cytochrome P450 (P450) drug-interaction screening potential, and astemizole demonstrated a better profile because it did not appreciably interact with other P450 probes. Thus, danazol, amiodarone, and astemizole will facilitate the ability to determine the metabolic role of CYP2J2 in hepatic and extrahepatic tissues. DOI : 10.1124/dmd.111.043505
AMLODIPINE
Time dependent inhibition of CYP3A4 (unknown origin) at 100 uM by LC/MS system
—
Inhibition = 42.0 %
Reference
Title : Combination of GSH trapping and time-dependent inhibition assays as a predictive method of drugs generating highly reactive metabolites. Authors : Nakayama S, Takakusa H, Watanabe A, Miyaji Y, Suzuki W, Sugiyama D, Shiosakai K, Honda K, Okudaira N, Izumi T, Okazaki O. Publication Year :
2011 Journal : Drug Metab. Dispos. Abstract : Covalent binding (CB) of reactive metabolites (RMs) is potentially involved in severe adverse drug reactions. Because the CB assay is of low throughput and costly, a qualitative trapping assay using agents such as [(35)S]GSH is often performed in the early stages of drug discovery. However, trapping methods alone cannot replace the CB assay. We hypothesized that the time-dependent inhibition (TDI) assay might be complementary to the [(35)S]GSH trapping assay in detecting RMs. We performed CB assays, [(35)S]GSH trapping assays, and TDI assays for 42 structurally diverse compounds. First, we showed that the [(35)S]GSH trapping assay alone does not correlate with the extent of CB. Four compounds that the [(35)S]GSH trapping assay failed to detect but that showed high extent of CB were inactivators of the enzyme in the TDI assay. There was a tendency for compounds judged as positive in the TDI assay to show a high degree of CB irrespective of the result of the [(35)S]GSH trapping assay. Finally, to combine parameters from the two assays, we introduced intrinsic clearance to describe the formation of RMs (CL(int, RMs)). The Spearman rank correlation coefficient between the extent of CB and CL(int, RMs) was 0.77 (p < 0.0001), which was better than that for the formation rates of [(35)S]GSH adducts. Therefore, we demonstrated that a combination of the [(35)S]GSH trapping and TDI assays is an effective method for detecting compounds potentially capable of generating highly reactive metabolites in the early stages of drug discovery. DOI : 10.1124/dmd.111.039180
AMLODIPINE
Time dependent inhibition of CYP3A4 (unknown origin) at 10 uM by LC/MS system
—
Inhibition < 10.0 %
Reference
Title : Combination of GSH trapping and time-dependent inhibition assays as a predictive method of drugs generating highly reactive metabolites. Authors : Nakayama S, Takakusa H, Watanabe A, Miyaji Y, Suzuki W, Sugiyama D, Shiosakai K, Honda K, Okudaira N, Izumi T, Okazaki O. Publication Year :
2011 Journal : Drug Metab. Dispos. Abstract : Covalent binding (CB) of reactive metabolites (RMs) is potentially involved in severe adverse drug reactions. Because the CB assay is of low throughput and costly, a qualitative trapping assay using agents such as [(35)S]GSH is often performed in the early stages of drug discovery. However, trapping methods alone cannot replace the CB assay. We hypothesized that the time-dependent inhibition (TDI) assay might be complementary to the [(35)S]GSH trapping assay in detecting RMs. We performed CB assays, [(35)S]GSH trapping assays, and TDI assays for 42 structurally diverse compounds. First, we showed that the [(35)S]GSH trapping assay alone does not correlate with the extent of CB. Four compounds that the [(35)S]GSH trapping assay failed to detect but that showed high extent of CB were inactivators of the enzyme in the TDI assay. There was a tendency for compounds judged as positive in the TDI assay to show a high degree of CB irrespective of the result of the [(35)S]GSH trapping assay. Finally, to combine parameters from the two assays, we introduced intrinsic clearance to describe the formation of RMs (CL(int, RMs)). The Spearman rank correlation coefficient between the extent of CB and CL(int, RMs) was 0.77 (p < 0.0001), which was better than that for the formation rates of [(35)S]GSH adducts. Therefore, we demonstrated that a combination of the [(35)S]GSH trapping and TDI assays is an effective method for detecting compounds potentially capable of generating highly reactive metabolites in the early stages of drug discovery. DOI : 10.1124/dmd.111.039180
AMODIAQUINE
Time dependent inhibition of CYP3A4 (unknown origin) at 100 uM by LC/MS system
—
Inhibition < 10.0 %
Reference
Title : Combination of GSH trapping and time-dependent inhibition assays as a predictive method of drugs generating highly reactive metabolites. Authors : Nakayama S, Takakusa H, Watanabe A, Miyaji Y, Suzuki W, Sugiyama D, Shiosakai K, Honda K, Okudaira N, Izumi T, Okazaki O. Publication Year :
2011 Journal : Drug Metab. Dispos. Abstract : Covalent binding (CB) of reactive metabolites (RMs) is potentially involved in severe adverse drug reactions. Because the CB assay is of low throughput and costly, a qualitative trapping assay using agents such as [(35)S]GSH is often performed in the early stages of drug discovery. However, trapping methods alone cannot replace the CB assay. We hypothesized that the time-dependent inhibition (TDI) assay might be complementary to the [(35)S]GSH trapping assay in detecting RMs. We performed CB assays, [(35)S]GSH trapping assays, and TDI assays for 42 structurally diverse compounds. First, we showed that the [(35)S]GSH trapping assay alone does not correlate with the extent of CB. Four compounds that the [(35)S]GSH trapping assay failed to detect but that showed high extent of CB were inactivators of the enzyme in the TDI assay. There was a tendency for compounds judged as positive in the TDI assay to show a high degree of CB irrespective of the result of the [(35)S]GSH trapping assay. Finally, to combine parameters from the two assays, we introduced intrinsic clearance to describe the formation of RMs (CL(int, RMs)). The Spearman rank correlation coefficient between the extent of CB and CL(int, RMs) was 0.77 (p < 0.0001), which was better than that for the formation rates of [(35)S]GSH adducts. Therefore, we demonstrated that a combination of the [(35)S]GSH trapping and TDI assays is an effective method for detecting compounds potentially capable of generating highly reactive metabolites in the early stages of drug discovery. DOI : 10.1124/dmd.111.039180
ATORVASTATIN
Time dependent inhibition of CYP3A4 (unknown origin) at 100 uM by LC/MS system
—
Inhibition < 10.0 %
Reference
Title : Combination of GSH trapping and time-dependent inhibition assays as a predictive method of drugs generating highly reactive metabolites. Authors : Nakayama S, Takakusa H, Watanabe A, Miyaji Y, Suzuki W, Sugiyama D, Shiosakai K, Honda K, Okudaira N, Izumi T, Okazaki O. Publication Year :
2011 Journal : Drug Metab. Dispos. Abstract : Covalent binding (CB) of reactive metabolites (RMs) is potentially involved in severe adverse drug reactions. Because the CB assay is of low throughput and costly, a qualitative trapping assay using agents such as [(35)S]GSH is often performed in the early stages of drug discovery. However, trapping methods alone cannot replace the CB assay. We hypothesized that the time-dependent inhibition (TDI) assay might be complementary to the [(35)S]GSH trapping assay in detecting RMs. We performed CB assays, [(35)S]GSH trapping assays, and TDI assays for 42 structurally diverse compounds. First, we showed that the [(35)S]GSH trapping assay alone does not correlate with the extent of CB. Four compounds that the [(35)S]GSH trapping assay failed to detect but that showed high extent of CB were inactivators of the enzyme in the TDI assay. There was a tendency for compounds judged as positive in the TDI assay to show a high degree of CB irrespective of the result of the [(35)S]GSH trapping assay. Finally, to combine parameters from the two assays, we introduced intrinsic clearance to describe the formation of RMs (CL(int, RMs)). The Spearman rank correlation coefficient between the extent of CB and CL(int, RMs) was 0.77 (p < 0.0001), which was better than that for the formation rates of [(35)S]GSH adducts. Therefore, we demonstrated that a combination of the [(35)S]GSH trapping and TDI assays is an effective method for detecting compounds potentially capable of generating highly reactive metabolites in the early stages of drug discovery. DOI : 10.1124/dmd.111.039180
BATEFENTEROL
Inhibition of CYP3A4 (unknown origin) at 600 nM
—
Inhibition < 20.0 %
Reference
Title : Discovery of (R)-1-(3-((2-chloro-4-(((2-hydroxy-2-(8-hydroxy-2-oxo-1,2-dihydroquinolin-5-yl)ethyl)amino)methyl)-5-methoxyphenyl)amino)-3-oxopropyl)piperidin-4-yl [1,1'-biphenyl]-2-ylcarbamate (TD-5959, GSK961081, batefenterol): first-in-class dual pharmacology multivalent muscarinic antagonist and β₂ agonist (MABA) for the treatment of chronic obstructive pulmonary disease (COPD). Authors : Hughes AD, Chen Y, Hegde SS, Jasper JR, Jaw-Tsai S, Lee TW, McNamara A, Pulido-Rios MT, Steinfeld T, Mammen M. Publication Year :
2015 Journal : J. Med. Chem. Abstract : Through application of our multivalent approach to drug discovery we previously reported the first discovery of dual pharmacology MABA bronchodilators, exemplified by 1. Herein we describe the subsequent lead optimization of both muscarinic antagonist and β2 agonist activities, through modification of the linker motif, to achieve 24 h duration of action in a guinea pig bronchoprotection model. Concomitantly we targeted high lung selectivities, low systemic exposures and identified crystalline forms suitable for inhalation devices. This article culminates with the discovery of our first clinical candidate 12f (TD-5959, GSK961081, batefenterol). In a phase 2b trial, batefenterol produced statistical and clinically significant differences compared to placebo and numerically greater improvements in the primary end point of trough FEV1 compared to salmeterol after 4 weeks of dosing in patients with moderate to severe chronic obstructive pulmonary disease (COPD). DOI : 10.1021/jm501915g
BENZBROMARONE
Time dependent inhibition of CYP3A4 (unknown origin) at 10 uM by LC/MS system
—
Inhibition < 10.0 %
Reference
Title : Combination of GSH trapping and time-dependent inhibition assays as a predictive method of drugs generating highly reactive metabolites. Authors : Nakayama S, Takakusa H, Watanabe A, Miyaji Y, Suzuki W, Sugiyama D, Shiosakai K, Honda K, Okudaira N, Izumi T, Okazaki O. Publication Year :
2011 Journal : Drug Metab. Dispos. Abstract : Covalent binding (CB) of reactive metabolites (RMs) is potentially involved in severe adverse drug reactions. Because the CB assay is of low throughput and costly, a qualitative trapping assay using agents such as [(35)S]GSH is often performed in the early stages of drug discovery. However, trapping methods alone cannot replace the CB assay. We hypothesized that the time-dependent inhibition (TDI) assay might be complementary to the [(35)S]GSH trapping assay in detecting RMs. We performed CB assays, [(35)S]GSH trapping assays, and TDI assays for 42 structurally diverse compounds. First, we showed that the [(35)S]GSH trapping assay alone does not correlate with the extent of CB. Four compounds that the [(35)S]GSH trapping assay failed to detect but that showed high extent of CB were inactivators of the enzyme in the TDI assay. There was a tendency for compounds judged as positive in the TDI assay to show a high degree of CB irrespective of the result of the [(35)S]GSH trapping assay. Finally, to combine parameters from the two assays, we introduced intrinsic clearance to describe the formation of RMs (CL(int, RMs)). The Spearman rank correlation coefficient between the extent of CB and CL(int, RMs) was 0.77 (p < 0.0001), which was better than that for the formation rates of [(35)S]GSH adducts. Therefore, we demonstrated that a combination of the [(35)S]GSH trapping and TDI assays is an effective method for detecting compounds potentially capable of generating highly reactive metabolites in the early stages of drug discovery. DOI : 10.1124/dmd.111.039180
BRIVANIB
Drug metabolism in human liver microsomes assessed as monoclonal antibody-mediated inhibition of CYP3A4-mediated (R)-1-(4-(4-fluoro-2-(hydroxymethyl)-1H-indol-5-yloxy)-5-methylpyrrolo[1,2-f][1,2,4]triazin-6-yloxy)propan-2-ol formation at 2 and 20 uM for 30 mins by LC/MS analysis
Liver
Inhibition = 50.0 %
Reference
Title : Identification of the oxidative and conjugative enzymes involved in the biotransformation of brivanib. Authors : Gong J, Gan J, Iyer RA. Publication Year :
2012 Journal : Drug Metab. Dispos. Abstract : Brivanib alaninate, the L-alanine ester prodrug of brivanib, is currently being developed as an anticancer agent. In humans, brivanib alaninate is rapidly hydrolyzed to brivanib. Prominent biotransformation pathways of brivanib included oxidation and direct sulfate conjugation. A series of in vitro studies were conducted to identify the human esterases involved in the prodrug hydrolysis and to identify the primary human cytochrome P450 and sulfotransferase (SULT) enzymes involved in the metabolism of brivanib. Brivanib alaninate was efficiently converted to brivanib in the presence of either human carboxylesterase 1 or carboxylesterase 2. Because esterases are ubiquitous, it is likely that multiple esterases are involved in the hydrolysis. Oxidation of brivanib in human liver microsomes (HLM) primarily formed a hydroxylated metabolite (M7). Incubation of brivanib with human cDNA-expressed P450 enzymes and with HLM in the presence of selective chemical inhibitors and monoclonal P450 antibodies demonstrated that CYP1A2 and CYP3A4 were the major contributors for the formation of M7. Direct sulfation of brivanib was catalyzed by multiple SULT enzymes, including SULT1A1, SULT1B1, SULT2A1, SULT1A3, and SULT1E1. Because the primary in vitro oxidative metabolite (M7) was not detected in humans after oral doses of brivanib alaninate, further metabolism studies of M7 in HLM and human liver cytosol were performed. The data demonstrated that M7 was metabolized to the prominent metabolites observed in humans. Overall, multiple enzymes are involved in the metabolism of brivanib, suggesting a low potential for drug-drug interactions either through polymorphism or through inhibition of a particular drug-metabolizing enzyme. DOI : 10.1124/dmd.111.042457
BRIVANIB
Drug metabolism in human liver microsomes assessed as 1 uM ketaconazole-mediated inhibition of CYP3A4-mediated (R)-1-(4-(4-fluoro-2-(hydroxymethyl)-1H-indol-5-yloxy)-5-methylpyrrolo[1,2-f][1,2,4]triazin-6-yloxy)propan-2-ol formation at 2 and 20 uM for 30 mins by LC/MS analysis
Liver
Inhibition = 50.0 %
Reference
Title : Identification of the oxidative and conjugative enzymes involved in the biotransformation of brivanib. Authors : Gong J, Gan J, Iyer RA. Publication Year :
2012 Journal : Drug Metab. Dispos. Abstract : Brivanib alaninate, the L-alanine ester prodrug of brivanib, is currently being developed as an anticancer agent. In humans, brivanib alaninate is rapidly hydrolyzed to brivanib. Prominent biotransformation pathways of brivanib included oxidation and direct sulfate conjugation. A series of in vitro studies were conducted to identify the human esterases involved in the prodrug hydrolysis and to identify the primary human cytochrome P450 and sulfotransferase (SULT) enzymes involved in the metabolism of brivanib. Brivanib alaninate was efficiently converted to brivanib in the presence of either human carboxylesterase 1 or carboxylesterase 2. Because esterases are ubiquitous, it is likely that multiple esterases are involved in the hydrolysis. Oxidation of brivanib in human liver microsomes (HLM) primarily formed a hydroxylated metabolite (M7). Incubation of brivanib with human cDNA-expressed P450 enzymes and with HLM in the presence of selective chemical inhibitors and monoclonal P450 antibodies demonstrated that CYP1A2 and CYP3A4 were the major contributors for the formation of M7. Direct sulfation of brivanib was catalyzed by multiple SULT enzymes, including SULT1A1, SULT1B1, SULT2A1, SULT1A3, and SULT1E1. Because the primary in vitro oxidative metabolite (M7) was not detected in humans after oral doses of brivanib alaninate, further metabolism studies of M7 in HLM and human liver cytosol were performed. The data demonstrated that M7 was metabolized to the prominent metabolites observed in humans. Overall, multiple enzymes are involved in the metabolism of brivanib, suggesting a low potential for drug-drug interactions either through polymorphism or through inhibition of a particular drug-metabolizing enzyme. DOI : 10.1124/dmd.111.042457
CAFFEINE
Time dependent inhibition of CYP3A4 (unknown origin) at 100 uM by LC/MS system
—
Inhibition < 10.0 %
Reference
Title : Combination of GSH trapping and time-dependent inhibition assays as a predictive method of drugs generating highly reactive metabolites. Authors : Nakayama S, Takakusa H, Watanabe A, Miyaji Y, Suzuki W, Sugiyama D, Shiosakai K, Honda K, Okudaira N, Izumi T, Okazaki O. Publication Year :
2011 Journal : Drug Metab. Dispos. Abstract : Covalent binding (CB) of reactive metabolites (RMs) is potentially involved in severe adverse drug reactions. Because the CB assay is of low throughput and costly, a qualitative trapping assay using agents such as [(35)S]GSH is often performed in the early stages of drug discovery. However, trapping methods alone cannot replace the CB assay. We hypothesized that the time-dependent inhibition (TDI) assay might be complementary to the [(35)S]GSH trapping assay in detecting RMs. We performed CB assays, [(35)S]GSH trapping assays, and TDI assays for 42 structurally diverse compounds. First, we showed that the [(35)S]GSH trapping assay alone does not correlate with the extent of CB. Four compounds that the [(35)S]GSH trapping assay failed to detect but that showed high extent of CB were inactivators of the enzyme in the TDI assay. There was a tendency for compounds judged as positive in the TDI assay to show a high degree of CB irrespective of the result of the [(35)S]GSH trapping assay. Finally, to combine parameters from the two assays, we introduced intrinsic clearance to describe the formation of RMs (CL(int, RMs)). The Spearman rank correlation coefficient between the extent of CB and CL(int, RMs) was 0.77 (p < 0.0001), which was better than that for the formation rates of [(35)S]GSH adducts. Therefore, we demonstrated that a combination of the [(35)S]GSH trapping and TDI assays is an effective method for detecting compounds potentially capable of generating highly reactive metabolites in the early stages of drug discovery. DOI : 10.1124/dmd.111.039180
CETIRIZINE
Inhibition of CYP3A4 in human microsomes at 25 uM
—
Inhibition = 13.0 %
Reference
Title : Synthesis and structure-activity relationships of novel histamine H1 antagonists: indolylpiperidinyl benzoic acid derivatives. Authors : Fonquerna S, Miralpeix M, Pagès L, Puig C, Cardús A, Antón F, Cárdenas A, Vilella D, Aparici M, Calaf E, Prieto J, Gras J, Huerta JM, Warrellow G, Beleta J, Ryder H. Publication Year :
2004 Journal : J. Med. Chem. Abstract : A series of indolylpiperidinyl derivatives were prepared and evaluated for their activity as histamine H(1) antagonists. Structure-activity relationship studies were directed toward improving in vivo activity and pharmacokinetic profile of our first lead (1). Substitution of fluorine in position 6 on the indolyl ring led to higher in vivo activity in the inhibition of histamine-induced cutaneous vascular permeability assay but lower selectivity toward 5HT(2) receptor. Extensive optimization was carried out within this series and a number of histamine H(1) antagonists showing potency and long duration of action in vivo and low brain penetration or cardiotoxic potential were identified. Within this novel series, indolylpiperidines 15, 20, 48,51 and 52 exhibited a long half-life in rat and have been selected for further preclinical evaluation. DOI : 10.1021/jm0498203
CPI-1205
Inhibition of CYP3A4 (unknown origin) at 10 uM relative to control
—
Inhibition = 0.0 %
Reference
Title : Identification of (R)-N-((4-Methoxy-6-methyl-2-oxo-1,2-dihydropyridin-3-yl)methyl)-2-methyl-1-(1-(1-(2,2,2-trifluoroethyl)piperidin-4-yl)ethyl)-1H-indole-3-carboxamide (CPI-1205), a Potent and Selective Inhibitor of Histone Methyltransferase EZH2, Suitable for Phase I Clinical Trials for B-Cell Lymphomas. Authors : Vaswani RG,Gehling VS,Dakin LA,Cook AS,Nasveschuk CG,Duplessis M,Iyer P,Balasubramanian S,Zhao F,Good AC,Campbell R,Lee C,Cantone N,Cummings RT,Normant E,Bellon SF,Albrecht BK,Harmange JC,Trojer P,Audia JE,Zhang Y,Justin N,Chen S,Wilson JR,Gamblin SJ Publication Year :
2016 Journal : J Med Chem Abstract : Polycomb repressive complex 2 (PRC2) has been shown to play a major role in transcriptional silencing in part by installing methylation marks on lysine 27 of histone 3. Dysregulation of PRC2 function correlates with certain malignancies and poor prognosis. EZH2 is the catalytic engine of the PRC2 complex and thus represents a key candidate oncology target for pharmacological intervention. Here we report the optimization of our indole-based EZH2 inhibitor series that led to the identification of CPI-1205, a highly potent (biochemical IC = 0.002 μM, cellular EC = 0.032 μM) and selective inhibitor of EZH2. This compound demonstrates robust antitumor effects in a Karpas-422 xenograft model when dosed at 160 mg/kg BID and is currently in Phase I clinical trials. Additionally, we disclose the co-crystal structure of our inhibitor series bound to the human PRC2 complex. DOI : 10.1021/acs.jmedchem.6b01315
DONEPEZIL
Inhibition of human recombinant CYP3A4 expressed in insect cell microsomes preincubated for 10 mins at 0.1 uM by fluorescence assay
—
Inhibition = 27.0 %
Reference
Title : Investigating the binding interactions of the anti-Alzheimer's drug donepezil with CYP3A4 and P-glycoprotein. Authors : McEneny-King A, Edginton AN, Rao PP. Publication Year :
2015 Journal : Bioorg. Med. Chem. Lett. Abstract : The anti-Alzheimer's agent donepezil is known to bind to the hepatic enzyme CYP3A4, but its relationship with the efflux transporter P-glycoprotein (P-gp) is not as well elucidated. We conducted in vitro inhibition studies of donepezil using human recombinant CYP3A4 and P-gp. These studies show that donepezil is a weak inhibitor of CYP3A4 (IC50=54.68±1.00μM) whereas the reference agent ketoconazole exhibited potent inhibition (CYP3A4 IC50=0.20±0.01μM). P-gp inhibition studies indicate that donepezil exhibits better inhibition relative to CYP3A4 (P-gp EC50=34.85±4.63μM) although it was less potent compared to ketoconazole (P-gp EC50=9.74±1.23μM). At higher concentrations, donepezil exhibited significant inhibition of CYP3A4 (69%, 84% and 87% inhibition at 100, 250 and 500μM, respectively). This indicates its potential to cause drug-drug interactions with other CYP3A4 substrates upon co-administration; however, this scenario is unlikely in vivo due to the low therapeutic concentrations of donepezil. Similarly, donepezil co-administration with P-gp substrates or inhibitors is unlikely to result in beneficial or adverse drug interactions. The molecular docking studies show that the 5,6-dimethoxyindan-1-one moiety of donepezil was oriented closer to the heme center in CYP3A4 whereas in the P-gp binding site, the protonated benzylpiperidine pharmacophore of donepezil played a major role in its binding ability. Energy parameters indicate that donepezil complex with both CYP3A4 and P-gp was less stable (CDOCKER energies=-15.05 and -4.91kcal/mol, respectively) compared to the ketoconazole-CYP3A4 and P-gp complex (CDOCKER energies=-41.89 and -20.03kcal/mol, respectively). DOI : 10.1016/j.bmcl.2014.11.046
DONEPEZIL
Inhibition of human recombinant CYP3A4 expressed in insect cell microsomes preincubated for 10 mins at 1 uM by fluorescence assay
—
Inhibition = 8.1 %
Reference
Title : Investigating the binding interactions of the anti-Alzheimer's drug donepezil with CYP3A4 and P-glycoprotein. Authors : McEneny-King A, Edginton AN, Rao PP. Publication Year :
2015 Journal : Bioorg. Med. Chem. Lett. Abstract : The anti-Alzheimer's agent donepezil is known to bind to the hepatic enzyme CYP3A4, but its relationship with the efflux transporter P-glycoprotein (P-gp) is not as well elucidated. We conducted in vitro inhibition studies of donepezil using human recombinant CYP3A4 and P-gp. These studies show that donepezil is a weak inhibitor of CYP3A4 (IC50=54.68±1.00μM) whereas the reference agent ketoconazole exhibited potent inhibition (CYP3A4 IC50=0.20±0.01μM). P-gp inhibition studies indicate that donepezil exhibits better inhibition relative to CYP3A4 (P-gp EC50=34.85±4.63μM) although it was less potent compared to ketoconazole (P-gp EC50=9.74±1.23μM). At higher concentrations, donepezil exhibited significant inhibition of CYP3A4 (69%, 84% and 87% inhibition at 100, 250 and 500μM, respectively). This indicates its potential to cause drug-drug interactions with other CYP3A4 substrates upon co-administration; however, this scenario is unlikely in vivo due to the low therapeutic concentrations of donepezil. Similarly, donepezil co-administration with P-gp substrates or inhibitors is unlikely to result in beneficial or adverse drug interactions. The molecular docking studies show that the 5,6-dimethoxyindan-1-one moiety of donepezil was oriented closer to the heme center in CYP3A4 whereas in the P-gp binding site, the protonated benzylpiperidine pharmacophore of donepezil played a major role in its binding ability. Energy parameters indicate that donepezil complex with both CYP3A4 and P-gp was less stable (CDOCKER energies=-15.05 and -4.91kcal/mol, respectively) compared to the ketoconazole-CYP3A4 and P-gp complex (CDOCKER energies=-41.89 and -20.03kcal/mol, respectively). DOI : 10.1016/j.bmcl.2014.11.046
DONEPEZIL
Inhibition of human recombinant CYP3A4 expressed in insect cell microsomes preincubated for 10 mins at 10 uM by fluorescence assay
—
Inhibition = 22.2 %
Reference
Title : Investigating the binding interactions of the anti-Alzheimer's drug donepezil with CYP3A4 and P-glycoprotein. Authors : McEneny-King A, Edginton AN, Rao PP. Publication Year :
2015 Journal : Bioorg. Med. Chem. Lett. Abstract : The anti-Alzheimer's agent donepezil is known to bind to the hepatic enzyme CYP3A4, but its relationship with the efflux transporter P-glycoprotein (P-gp) is not as well elucidated. We conducted in vitro inhibition studies of donepezil using human recombinant CYP3A4 and P-gp. These studies show that donepezil is a weak inhibitor of CYP3A4 (IC50=54.68±1.00μM) whereas the reference agent ketoconazole exhibited potent inhibition (CYP3A4 IC50=0.20±0.01μM). P-gp inhibition studies indicate that donepezil exhibits better inhibition relative to CYP3A4 (P-gp EC50=34.85±4.63μM) although it was less potent compared to ketoconazole (P-gp EC50=9.74±1.23μM). At higher concentrations, donepezil exhibited significant inhibition of CYP3A4 (69%, 84% and 87% inhibition at 100, 250 and 500μM, respectively). This indicates its potential to cause drug-drug interactions with other CYP3A4 substrates upon co-administration; however, this scenario is unlikely in vivo due to the low therapeutic concentrations of donepezil. Similarly, donepezil co-administration with P-gp substrates or inhibitors is unlikely to result in beneficial or adverse drug interactions. The molecular docking studies show that the 5,6-dimethoxyindan-1-one moiety of donepezil was oriented closer to the heme center in CYP3A4 whereas in the P-gp binding site, the protonated benzylpiperidine pharmacophore of donepezil played a major role in its binding ability. Energy parameters indicate that donepezil complex with both CYP3A4 and P-gp was less stable (CDOCKER energies=-15.05 and -4.91kcal/mol, respectively) compared to the ketoconazole-CYP3A4 and P-gp complex (CDOCKER energies=-41.89 and -20.03kcal/mol, respectively). DOI : 10.1016/j.bmcl.2014.11.046
DONEPEZIL
Inhibition of human recombinant CYP3A4 expressed in insect cell microsomes preincubated for 10 mins at 100 uM by fluorescence assay
—
Inhibition = 68.4 %
Reference
Title : Investigating the binding interactions of the anti-Alzheimer's drug donepezil with CYP3A4 and P-glycoprotein. Authors : McEneny-King A, Edginton AN, Rao PP. Publication Year :
2015 Journal : Bioorg. Med. Chem. Lett. Abstract : The anti-Alzheimer's agent donepezil is known to bind to the hepatic enzyme CYP3A4, but its relationship with the efflux transporter P-glycoprotein (P-gp) is not as well elucidated. We conducted in vitro inhibition studies of donepezil using human recombinant CYP3A4 and P-gp. These studies show that donepezil is a weak inhibitor of CYP3A4 (IC50=54.68±1.00μM) whereas the reference agent ketoconazole exhibited potent inhibition (CYP3A4 IC50=0.20±0.01μM). P-gp inhibition studies indicate that donepezil exhibits better inhibition relative to CYP3A4 (P-gp EC50=34.85±4.63μM) although it was less potent compared to ketoconazole (P-gp EC50=9.74±1.23μM). At higher concentrations, donepezil exhibited significant inhibition of CYP3A4 (69%, 84% and 87% inhibition at 100, 250 and 500μM, respectively). This indicates its potential to cause drug-drug interactions with other CYP3A4 substrates upon co-administration; however, this scenario is unlikely in vivo due to the low therapeutic concentrations of donepezil. Similarly, donepezil co-administration with P-gp substrates or inhibitors is unlikely to result in beneficial or adverse drug interactions. The molecular docking studies show that the 5,6-dimethoxyindan-1-one moiety of donepezil was oriented closer to the heme center in CYP3A4 whereas in the P-gp binding site, the protonated benzylpiperidine pharmacophore of donepezil played a major role in its binding ability. Energy parameters indicate that donepezil complex with both CYP3A4 and P-gp was less stable (CDOCKER energies=-15.05 and -4.91kcal/mol, respectively) compared to the ketoconazole-CYP3A4 and P-gp complex (CDOCKER energies=-41.89 and -20.03kcal/mol, respectively). DOI : 10.1016/j.bmcl.2014.11.046
DONEPEZIL
Inhibition of human recombinant CYP3A4 expressed in insect cell microsomes preincubated for 10 mins at 250 uM by fluorescence assay
—
Inhibition = 83.9 %
Reference
Title : Investigating the binding interactions of the anti-Alzheimer's drug donepezil with CYP3A4 and P-glycoprotein. Authors : McEneny-King A, Edginton AN, Rao PP. Publication Year :
2015 Journal : Bioorg. Med. Chem. Lett. Abstract : The anti-Alzheimer's agent donepezil is known to bind to the hepatic enzyme CYP3A4, but its relationship with the efflux transporter P-glycoprotein (P-gp) is not as well elucidated. We conducted in vitro inhibition studies of donepezil using human recombinant CYP3A4 and P-gp. These studies show that donepezil is a weak inhibitor of CYP3A4 (IC50=54.68±1.00μM) whereas the reference agent ketoconazole exhibited potent inhibition (CYP3A4 IC50=0.20±0.01μM). P-gp inhibition studies indicate that donepezil exhibits better inhibition relative to CYP3A4 (P-gp EC50=34.85±4.63μM) although it was less potent compared to ketoconazole (P-gp EC50=9.74±1.23μM). At higher concentrations, donepezil exhibited significant inhibition of CYP3A4 (69%, 84% and 87% inhibition at 100, 250 and 500μM, respectively). This indicates its potential to cause drug-drug interactions with other CYP3A4 substrates upon co-administration; however, this scenario is unlikely in vivo due to the low therapeutic concentrations of donepezil. Similarly, donepezil co-administration with P-gp substrates or inhibitors is unlikely to result in beneficial or adverse drug interactions. The molecular docking studies show that the 5,6-dimethoxyindan-1-one moiety of donepezil was oriented closer to the heme center in CYP3A4 whereas in the P-gp binding site, the protonated benzylpiperidine pharmacophore of donepezil played a major role in its binding ability. Energy parameters indicate that donepezil complex with both CYP3A4 and P-gp was less stable (CDOCKER energies=-15.05 and -4.91kcal/mol, respectively) compared to the ketoconazole-CYP3A4 and P-gp complex (CDOCKER energies=-41.89 and -20.03kcal/mol, respectively). DOI : 10.1016/j.bmcl.2014.11.046
DONEPEZIL
Inhibition of human recombinant CYP3A4 expressed in insect cell microsomes preincubated for 10 mins at 500 uM by fluorescence assay
—
Inhibition = 87.3 %
Reference
Title : Investigating the binding interactions of the anti-Alzheimer's drug donepezil with CYP3A4 and P-glycoprotein. Authors : McEneny-King A, Edginton AN, Rao PP. Publication Year :
2015 Journal : Bioorg. Med. Chem. Lett. Abstract : The anti-Alzheimer's agent donepezil is known to bind to the hepatic enzyme CYP3A4, but its relationship with the efflux transporter P-glycoprotein (P-gp) is not as well elucidated. We conducted in vitro inhibition studies of donepezil using human recombinant CYP3A4 and P-gp. These studies show that donepezil is a weak inhibitor of CYP3A4 (IC50=54.68±1.00μM) whereas the reference agent ketoconazole exhibited potent inhibition (CYP3A4 IC50=0.20±0.01μM). P-gp inhibition studies indicate that donepezil exhibits better inhibition relative to CYP3A4 (P-gp EC50=34.85±4.63μM) although it was less potent compared to ketoconazole (P-gp EC50=9.74±1.23μM). At higher concentrations, donepezil exhibited significant inhibition of CYP3A4 (69%, 84% and 87% inhibition at 100, 250 and 500μM, respectively). This indicates its potential to cause drug-drug interactions with other CYP3A4 substrates upon co-administration; however, this scenario is unlikely in vivo due to the low therapeutic concentrations of donepezil. Similarly, donepezil co-administration with P-gp substrates or inhibitors is unlikely to result in beneficial or adverse drug interactions. The molecular docking studies show that the 5,6-dimethoxyindan-1-one moiety of donepezil was oriented closer to the heme center in CYP3A4 whereas in the P-gp binding site, the protonated benzylpiperidine pharmacophore of donepezil played a major role in its binding ability. Energy parameters indicate that donepezil complex with both CYP3A4 and P-gp was less stable (CDOCKER energies=-15.05 and -4.91kcal/mol, respectively) compared to the ketoconazole-CYP3A4 and P-gp complex (CDOCKER energies=-41.89 and -20.03kcal/mol, respectively). DOI : 10.1016/j.bmcl.2014.11.046
DONEPEZIL
Time dependent inhibition of CYP3A4 (unknown origin) at 100 uM by LC/MS system
—
Inhibition < 10.0 %
Reference
Title : Combination of GSH trapping and time-dependent inhibition assays as a predictive method of drugs generating highly reactive metabolites. Authors : Nakayama S, Takakusa H, Watanabe A, Miyaji Y, Suzuki W, Sugiyama D, Shiosakai K, Honda K, Okudaira N, Izumi T, Okazaki O. Publication Year :
2011 Journal : Drug Metab. Dispos. Abstract : Covalent binding (CB) of reactive metabolites (RMs) is potentially involved in severe adverse drug reactions. Because the CB assay is of low throughput and costly, a qualitative trapping assay using agents such as [(35)S]GSH is often performed in the early stages of drug discovery. However, trapping methods alone cannot replace the CB assay. We hypothesized that the time-dependent inhibition (TDI) assay might be complementary to the [(35)S]GSH trapping assay in detecting RMs. We performed CB assays, [(35)S]GSH trapping assays, and TDI assays for 42 structurally diverse compounds. First, we showed that the [(35)S]GSH trapping assay alone does not correlate with the extent of CB. Four compounds that the [(35)S]GSH trapping assay failed to detect but that showed high extent of CB were inactivators of the enzyme in the TDI assay. There was a tendency for compounds judged as positive in the TDI assay to show a high degree of CB irrespective of the result of the [(35)S]GSH trapping assay. Finally, to combine parameters from the two assays, we introduced intrinsic clearance to describe the formation of RMs (CL(int, RMs)). The Spearman rank correlation coefficient between the extent of CB and CL(int, RMs) was 0.77 (p < 0.0001), which was better than that for the formation rates of [(35)S]GSH adducts. Therefore, we demonstrated that a combination of the [(35)S]GSH trapping and TDI assays is an effective method for detecting compounds potentially capable of generating highly reactive metabolites in the early stages of drug discovery. DOI : 10.1124/dmd.111.039180
DULOXETINE
Inhibition of human recombinant CYP3A4 incubated for 5 mins by fluorescence assay
—
IC50 = 440.0 nM
Reference
Title : Exploration of 3-Aminoazetidines as Triple Reuptake Inhibitors by Bioisosteric Modification of 3-α-Oxyazetidine. Authors : Han M, Song C, Jeong N, Hahn HG. Publication Year :
2014 Journal : ACS Med. Chem. Lett. Abstract : For a development of broad spectrum antidepressant 3-aminoazetidine derivatives, two series of compounds were explored by bioisosteric modification of 3-α-oxyazetidine. We synthesized 166 novel 3-aminoazetidine derivatives in series A and B, starting from Boc-protected 3-azetidinone (3) and Boc-protected 3-azetidinal (9) respectively, through parallel syntheses. The inhibitory reuptake activities against serotonin (5-HT), norepinephrine (NE), and dopamine (DA) neurotransmitters were measured by the Neurotransmitter Transporter Uptake Assay Kit using the human embryonic kidney 293 (HEK293) cells stably transfected with the respective three kinds of human transporters (hSERT, hNET, and hDAT). Our study aimed to identify compounds having relative inhibitory activities against hSERT > hNET > hDAT. Lead optimization including microsomal stability, CYP, hERG assay, Ames test, BBB, and PK study resulted in the identification of compound 10dl as a candidate for further studies. DOI : 10.1021/ml500187a
DULOXETINE
Inhibition of recombinant human CYP3A4 preincubated for 5 mins before fluorescent substrate addition by fluorescence assay
—
IC50 = 440.0 nM
Reference
Title : Synthesis and biological evaluation of 3-phenethylazetidine derivatives as triple reuptake inhibitors. Authors : Yun J, Han M, Song C, Cheon SH, Choi K, Hahn HG. Publication Year :
2014 Journal : Bioorg. Med. Chem. Lett. Abstract : We report the synthesis of 3-phenethylazetidine derivatives 2 and their biological activities against 5-HT, NE and DA transporters as well as microsomal stability, CYP inhibition, and hERG inhibition profiles. Compound 2at showed most potent triple reuptake inhibitor with good selectivity as a candidate for depression. DOI : 10.1016/j.bmcl.2014.06.026
FASIGLIFAM
Inhibition of CYP3A4 in human liver microsomes at 5 uM using midazolam as substrate by LC-MS/MS analysis relative to control
Liver
Inhibition = 33.96 %
Reference
Title : Design, synthesis and biological evaluation of a series of novel GPR40 agonists containing nitrogen heterocyclic rings. Authors : Sun Z, Zhou T, Pan X, Yang Y, Huan Y, Xiao Z, Shen Z, Liu Z. Publication Year :
2018 Journal : Bioorg Med Chem Lett Abstract : A novel series of GPR40 agonists is designed by introducing nitrogen-containing heterocyclic ring at the terminal phenyl ring of TAK-875 with the aim of decreasing its lipophilicity. Three different β-substituted phenylpropionic acids were investigated as the acidic components. A total of 34 compounds have been synthesized, among which, compound 30 exhibited comparable GPR40 agonistic activity in vitro with TAK-875 and relatively lower lipophilicity through calculation (30, EC50 = 1.2 μM, cLogP = 1.3; TAK-875: EC50 = 5.1 μM, cLogP = 3.4). Moreover, compound 30 was able to enhance the insulin secretion of primary islets isolated from normal ICR mice and showed no obvious inhibition against cytochromes P450 in vitro. In vivo, compound 30 exhibited efficacy in oral glucose tolerance test (oGTT) in normal ICR mice. DOI : 10.1016/j.bmcl.2018.07.048
FLUOXETINE
Time dependent inhibition of CYP3A4 (unknown origin) at 100 uM by LC/MS system
—
Inhibition < 10.0 %
Reference
Title : Combination of GSH trapping and time-dependent inhibition assays as a predictive method of drugs generating highly reactive metabolites. Authors : Nakayama S, Takakusa H, Watanabe A, Miyaji Y, Suzuki W, Sugiyama D, Shiosakai K, Honda K, Okudaira N, Izumi T, Okazaki O. Publication Year :
2011 Journal : Drug Metab. Dispos. Abstract : Covalent binding (CB) of reactive metabolites (RMs) is potentially involved in severe adverse drug reactions. Because the CB assay is of low throughput and costly, a qualitative trapping assay using agents such as [(35)S]GSH is often performed in the early stages of drug discovery. However, trapping methods alone cannot replace the CB assay. We hypothesized that the time-dependent inhibition (TDI) assay might be complementary to the [(35)S]GSH trapping assay in detecting RMs. We performed CB assays, [(35)S]GSH trapping assays, and TDI assays for 42 structurally diverse compounds. First, we showed that the [(35)S]GSH trapping assay alone does not correlate with the extent of CB. Four compounds that the [(35)S]GSH trapping assay failed to detect but that showed high extent of CB were inactivators of the enzyme in the TDI assay. There was a tendency for compounds judged as positive in the TDI assay to show a high degree of CB irrespective of the result of the [(35)S]GSH trapping assay. Finally, to combine parameters from the two assays, we introduced intrinsic clearance to describe the formation of RMs (CL(int, RMs)). The Spearman rank correlation coefficient between the extent of CB and CL(int, RMs) was 0.77 (p < 0.0001), which was better than that for the formation rates of [(35)S]GSH adducts. Therefore, we demonstrated that a combination of the [(35)S]GSH trapping and TDI assays is an effective method for detecting compounds potentially capable of generating highly reactive metabolites in the early stages of drug discovery. DOI : 10.1124/dmd.111.039180
INDINAVIR
In vitro inhibition of human liver microsome Cytochrome P450 3A4
Liver
IC50 = 150.0 nM
Reference
Title : HIV protease inhibitors with picomolar potency against PI-Resistant HIV-1 by extension of the P3 substituent. Authors : Duffy JL, Rano TA, Kevin NJ, Chapman KT, Schleif WA, Olsen DB, Stahlhut M, Rutkowski CA, Kuo LC, Jin L, Lin JH, Emini EA, Tata JR. Publication Year :
2003 Journal : Bioorg. Med. Chem. Lett. Abstract : A biaryl pyridylfuran P(3) substituent on the hydroxyethylene isostere scaffold affords HIV protease inhibitors (PI's) with picomolar (IC(50)) potency against the protease enzymes from PI-resistant HIV-1 strains. Inclusion of a gem-dimethyl substituent afforded compound 3 with 100% oral bioavailability (dogs) and more than double the t(1/2) of indinavir. Inhibition of multiple P450 isoforms is dependent on the regiochemistry of the pyridyl nitrogen in these compounds. DOI : 10.1016/s0960-894x(03)00475-x
INDINAVIR
In vitro inhibition of Cytochrome P450 3A4
—
IC50 = 150.0 nM
Reference
Title : Novel HIV-1 protease inhibitors active against multiple PI-resistant viral strains: coadministration with indinavir. Authors : Kevin NJ, Duffy JL, Kirk BA, Chapman KT, Schleif WA, Olsen DB, Stahlhut M, Rutkowski CA, Kuo LC, Jin L, Lin JH, Emini EA, Tata JR. Publication Year :
2003 Journal : Bioorg. Med. Chem. Lett. Abstract : HIV-1 protease inhibitors (PI) with an N-arylpyrrole moiety in the P(3) position afforded excellent antiviral potency and substantially improved aqueous solubility over previously reported variants. The rapid in vitro clearance of these compounds in human liver microsomes prompted oral coadministration with indinavir to hinder their metabolism by the cyctochrome P450 3A4 isozyme and allow for in vivo PK assessment. DOI : 10.1016/j.bmcl.2003.08.049
INDINAVIR
Inhibition of human liver microsome Cytochrome P450 3A4
Liver
IC50 = 150.0 nM
Reference
Title : The design, synthesis and evaluation of novel HIV-1 protease inhibitors with high potency against PI-resistant viral strains. Authors : Zhang F, Chapman KT, Schleif WA, Olsen DB, Stahlhut M, Rutkowski CA, Kuo LC, Jin L, Lin JH, Emini EA, Tata JR. Publication Year :
2003 Journal : Bioorg. Med. Chem. Lett. Abstract : Replacement of the pyridylmethyl moiety in indinavir with a pyridyl oxazole yielded HIV-1 protease inhibitors (PI) with greatly improved potency against PI-resistant HIV-1 strains. A meta-methoxy group on the pyridyl ring and a gem-dimethyl methyl linkage afforded compound 10 with notable in vitro antiviral activity against HIV-1 viral strains with reduced susceptibility to the clinically available PIs. Compound 10 also demonstrated favorable in vivo pharmacokinetics in animal models. DOI : 10.1016/s0960-894x(03)00474-8
INDINAVIR
Mechanism based inhibition of human cytochrome P450 3A4 measured by testosterone hydroxylation
—
Ki = 480.0 nM
Reference
Title : Cytochrome p450 enzymes mechanism based inhibitors: common sub-structures and reactivity. Authors : Fontana E, Dansette PM, Poli SM. Publication Year :
2005 Journal : Curr. Drug Metab. Abstract : The inhibition of human cytochrome P450s (CYPs) is one of the most common mechanisms which can lead to drug-drug interactions. The inhibition of CYPs can be reversible (competitive or non-competitive) or irreversible. Irreversible inhibition usually derives from activation of a drug by CYPs into a reactive metabolite, which tightly binds to the enzyme active site, leading to a long lasting inactivation. This process is called "mechanism based inhibition" or "suicide inhibition". The irreversible inactivation usually implies the formation of a covalent bond between the metabolite and the enzyme, which can lead to hapten formation and can in some cases trigger an autoimmune-response. For these reasons it is of utmost importance to study the mechanism of the CYP inhibition of new potential drugs as early as possible during the drug discovery process. The literature on CYPs is vast and covers numerous aspects of their biology and biochemistry, however to our knowledge there is no general and systematic review focusing on mechanism-based inhibitors; we have reviewed the literature and compiled all the available data on chemical entities, which are known to be CYP suicide inhibitors. Each compound is reported together with its chemical structure, the CYP isoform and the parameters describing the inactivation. Literature references are reported together with their PMID (PubMed ID number) to allow a fast retrieval of the papers. This review offers a quick reference to help predict liabilities of new chemical entities without carrying out extensive in vitro work, and will hopefully help in designing safer drugs. DOI : 10.2174/138920005774330639
LEVOKETOCONAZOLE
Inhibition of CYP3A4M in human liver microsomes using midazolam as substrate in presence of NADPH incubated for 10 mins by LC-MS/MS analysis
Liver
IC50 = 26.9 nM
Reference
Title : Authors : Publication Year : Journal :
LIAROZOLE
Inhibition of CYP3A4 (unknown origin) incubated for 45 mins using NADPH by fluorescence assay
—
IC50 = 690.0 nM
Reference
Title : Design, synthesis, and biological evaluation of amide imidazole derivatives as novel metabolic enzyme CYP26A1 inhibitors. Authors : Sun B, Liu K, Han J, Zhao LY, Su X, Lin B, Zhao DM, Cheng MS. Publication Year :
2015 Journal : Bioorg. Med. Chem. Abstract : All-trans-retinoic acid (ATRA) as a physiological metabolite of vitamin A is widely applied in the treatment of cancer, skin, neurodegenerative and autoimmune diseases. CYP26A1 enzyme, induced by ATRA in liver and target tissues, metabolizes ATRA into 4-hydroxyl-RA. Inhibition of CYP26A1 metabolic enzyme represents a promising strategy for discovery of new specific anticancer agents. Herein, we describe the design, synthesis and biological evaluation of a series of new amide imidazole derivatives as retinoic acid metabolism blocking agents (RAMBAs) toward CYP26A1 enzyme. First, based on the recent theoretical models (Sun et al., J. Mol. Graph. Model., 2015, 56, 10-19) a series of RAMBAs with novel scaffolds were designed using fragment-based drug discovery approach. Subsequently, the new RAMBAs were synthesized and evaluated for their biological activities. All the compounds demonstrated appropriate enzyme activities and cell activities. The promising inhibitors 20 and 23 with IC50 value of 0.22 μM and 0.46 μM toward CYP26A1, respectively, were further evaluated for CYP selectivity and the metabolic profile of ATRA. Both compounds 20 and 23 showed higher selectivity for CYP26A1 over other CYPs (CYP2D6, CYP3A4) when compared to liarozole. They also showed better inhibitory activities for the metabolism of ATRA when also compared to liarozole. These studies further validated the pharmacophore and structure-activity relationship models obtained about CYP26A1 inhibitors and highlighted the promising activities of the new series of CYP26A1 inhibitors designed from such models. They also paved the way for future development of those candidates as potential drugs. DOI : 10.1016/j.bmc.2015.08.019
LOPINAVIR
Mechanism based inhibition of human cytochrome P450 3A4 measured by testosterone hydroxylation
—
Ki = 410.0 nM
Reference
Title : Cytochrome p450 enzymes mechanism based inhibitors: common sub-structures and reactivity. Authors : Fontana E, Dansette PM, Poli SM. Publication Year :
2005 Journal : Curr. Drug Metab. Abstract : The inhibition of human cytochrome P450s (CYPs) is one of the most common mechanisms which can lead to drug-drug interactions. The inhibition of CYPs can be reversible (competitive or non-competitive) or irreversible. Irreversible inhibition usually derives from activation of a drug by CYPs into a reactive metabolite, which tightly binds to the enzyme active site, leading to a long lasting inactivation. This process is called "mechanism based inhibition" or "suicide inhibition". The irreversible inactivation usually implies the formation of a covalent bond between the metabolite and the enzyme, which can lead to hapten formation and can in some cases trigger an autoimmune-response. For these reasons it is of utmost importance to study the mechanism of the CYP inhibition of new potential drugs as early as possible during the drug discovery process. The literature on CYPs is vast and covers numerous aspects of their biology and biochemistry, however to our knowledge there is no general and systematic review focusing on mechanism-based inhibitors; we have reviewed the literature and compiled all the available data on chemical entities, which are known to be CYP suicide inhibitors. Each compound is reported together with its chemical structure, the CYP isoform and the parameters describing the inactivation. Literature references are reported together with their PMID (PubMed ID number) to allow a fast retrieval of the papers. This review offers a quick reference to help predict liabilities of new chemical entities without carrying out extensive in vitro work, and will hopefully help in designing safer drugs. DOI : 10.2174/138920005774330639
MIBEFRADIL
Inhibition of recombinant CYP3A4 (unknown origin) expressed in baculosomes at 10 uM measured after 1.5 min in presence of 1 mM NADPH
—
Inhibition < 20.0 %
Reference
Title : Mechanism-based inactivation of cytochrome P450 3A4 by mibefradil through heme destruction. Authors : Foti RS, Rock DA, Pearson JT, Wahlstrom JL, Wienkers LC. Publication Year :
2011 Journal : Drug Metab. Dispos. Abstract : Mibefradil (Posicor) was developed as a calcium channel blocker for the treatment of chronic hypertension. The compound was withdrawn from the market in 1998 because of the potential for rhabdomyolysis, renal failure, or bradycardia when it was coadministered with other drugs. Mibefradil has previously been shown to be a potent reversible (IC(50) = 0.3-2 μM) and mechanism-based (K(i) = 2.3 μM; k(inact) = 0.4 min(-1)) inhibitor of CYP3A4-catalyzed statin metabolism. At present, the mechanism of CYP3A4 inactivation by mibefradil is not known. Mechanism-based inactivation experiments and spectral studies were used to examine the mechanism of CYP3A4 inactivation by mibefradil and its major metabolite, des-methoxyacetyl mibefradil (Ro 40-5966), in vitro. Both mibefradil and Ro 40-5966 were shown to exhibit type I binding characteristics (K(s) = 0.69 ± 0.06 and 1.39 ± 0.04 μM, respectively) toward CYP3A4. Complete K(i)/k(inact) experiments were performed, revealing a rapid and irreversible decrease in CYP3A4-catalyzed 1'-hydroxymidazolam formation. Approximately 70% of CYP3A4 activity was lost in the first minute of incubation with mibefradil, and inactivation was nonlinear after 2 min. Ro 40-5966 also resulted in time-dependent inhibition of CYP3A4, albeit to a lesser extent than mibefradil. The decrease in CYP3A4 activity in the presence of mibefradil and NADPH was subsequently shown to have a good correlation with the time-dependent loss of CO binding, which, coupled with the lack of stable heme and/or apoprotein adducts, suggests heme destruction as the mechanism of inactivation of CYP3A4 by mibefradil. DOI : 10.1124/dmd.111.038505
MIBEFRADIL
Inhibition of recombinant CYP3A4 (unknown origin) expressed in baculosomes assessed as decrease in heme level at 10 uM preincubation for 5 mins by LC/UV/MS analysis in presence of 1 mM NADPH
—
Inhibition = 80.0 %
Reference
Title : Mechanism-based inactivation of cytochrome P450 3A4 by mibefradil through heme destruction. Authors : Foti RS, Rock DA, Pearson JT, Wahlstrom JL, Wienkers LC. Publication Year :
2011 Journal : Drug Metab. Dispos. Abstract : Mibefradil (Posicor) was developed as a calcium channel blocker for the treatment of chronic hypertension. The compound was withdrawn from the market in 1998 because of the potential for rhabdomyolysis, renal failure, or bradycardia when it was coadministered with other drugs. Mibefradil has previously been shown to be a potent reversible (IC(50) = 0.3-2 μM) and mechanism-based (K(i) = 2.3 μM; k(inact) = 0.4 min(-1)) inhibitor of CYP3A4-catalyzed statin metabolism. At present, the mechanism of CYP3A4 inactivation by mibefradil is not known. Mechanism-based inactivation experiments and spectral studies were used to examine the mechanism of CYP3A4 inactivation by mibefradil and its major metabolite, des-methoxyacetyl mibefradil (Ro 40-5966), in vitro. Both mibefradil and Ro 40-5966 were shown to exhibit type I binding characteristics (K(s) = 0.69 ± 0.06 and 1.39 ± 0.04 μM, respectively) toward CYP3A4. Complete K(i)/k(inact) experiments were performed, revealing a rapid and irreversible decrease in CYP3A4-catalyzed 1'-hydroxymidazolam formation. Approximately 70% of CYP3A4 activity was lost in the first minute of incubation with mibefradil, and inactivation was nonlinear after 2 min. Ro 40-5966 also resulted in time-dependent inhibition of CYP3A4, albeit to a lesser extent than mibefradil. The decrease in CYP3A4 activity in the presence of mibefradil and NADPH was subsequently shown to have a good correlation with the time-dependent loss of CO binding, which, coupled with the lack of stable heme and/or apoprotein adducts, suggests heme destruction as the mechanism of inactivation of CYP3A4 by mibefradil. DOI : 10.1124/dmd.111.038505
MIBEFRADIL
Inhibition of recombinant CYP3A4 (unknown origin) expressed in baculosomes in presence of 1 mM NADPH
—
Ki = 280.0 nM
Reference
Title : Mechanism-based inactivation of cytochrome P450 3A4 by mibefradil through heme destruction. Authors : Foti RS, Rock DA, Pearson JT, Wahlstrom JL, Wienkers LC. Publication Year :
2011 Journal : Drug Metab. Dispos. Abstract : Mibefradil (Posicor) was developed as a calcium channel blocker for the treatment of chronic hypertension. The compound was withdrawn from the market in 1998 because of the potential for rhabdomyolysis, renal failure, or bradycardia when it was coadministered with other drugs. Mibefradil has previously been shown to be a potent reversible (IC(50) = 0.3-2 μM) and mechanism-based (K(i) = 2.3 μM; k(inact) = 0.4 min(-1)) inhibitor of CYP3A4-catalyzed statin metabolism. At present, the mechanism of CYP3A4 inactivation by mibefradil is not known. Mechanism-based inactivation experiments and spectral studies were used to examine the mechanism of CYP3A4 inactivation by mibefradil and its major metabolite, des-methoxyacetyl mibefradil (Ro 40-5966), in vitro. Both mibefradil and Ro 40-5966 were shown to exhibit type I binding characteristics (K(s) = 0.69 ± 0.06 and 1.39 ± 0.04 μM, respectively) toward CYP3A4. Complete K(i)/k(inact) experiments were performed, revealing a rapid and irreversible decrease in CYP3A4-catalyzed 1'-hydroxymidazolam formation. Approximately 70% of CYP3A4 activity was lost in the first minute of incubation with mibefradil, and inactivation was nonlinear after 2 min. Ro 40-5966 also resulted in time-dependent inhibition of CYP3A4, albeit to a lesser extent than mibefradil. The decrease in CYP3A4 activity in the presence of mibefradil and NADPH was subsequently shown to have a good correlation with the time-dependent loss of CO binding, which, coupled with the lack of stable heme and/or apoprotein adducts, suggests heme destruction as the mechanism of inactivation of CYP3A4 by mibefradil. DOI : 10.1124/dmd.111.038505
MIBEFRADIL
Inhibition of CYP3A4 in human liver microsomes using testosterone substrate by LC-MS/MS method
Liver
IC50 = 470.0 nM
Reference
Title : Discovery and characterization of novel, potent, and selective cytochrome P450 2J2 inhibitors. Authors : Ren S, Zeng J, Mei Y, Zhang JZ, Yan SF, Fei J, Chen L. Publication Year :
2013 Journal : Drug Metab. Dispos. Abstract : Cytochrome P450 (CYP) 2J2 is one of the human CYPs involved in phase I xenobiotics metabolism. It is mainly expressed in extrahepatic tissues, including intestine and cardiovascular systems. The general role of CYP2J2 in drug metabolism is not yet fully understood, and the recent discovery that CYP2J2 can metabolize a wide range of structurally diverse drugs and its primary distribution in the intestine suggest its potentially indispensable role in first-pass intestinal metabolism and involvement in drug-drug interaction. To fully characterize its role in drug metabolism, selective and potent inhibitors of CYP2J2 are necessary tools. In the current study, 69 known drugs were screened for the inhibition of CYP2J2, and we discovered a number of marketed drugs as potent and selective CYP2J2 inhibitors. In particular, telmisartan and flunarizine have CYP2J2 inhibition IC(50) values of 0.42 μM and 0.94 μM, respectively, which are at least 10-fold more selective against all other major metabolizing CYPs; moreover, they are not substrates of CYP2J2 and show no time-dependent inhibition toward this CYP. The results of enzyme kinetics studies, supported by molecular modeling, have also elucidated that telmisartan is a mixed-type inhibitor, and flunarizine competitively inhibits CYP2J2. The K(i) for telmisartan is 0.19 μM, with an α value, an indicator of the type of inhibition mechanism, of 2.80, and flunarizine has a K(i) value of 0.13 μM. These newly discovered CYP2J2 inhibitors can be potentially used as a tool to study CYP2J2 in drug metabolism and interaction in a clinical setting. DOI : 10.1124/dmd.112.048264
NALMEFENE
Inhibition of human CYP3A4 at 10 uM
—
Inhibition = 60.0 %
Reference
Title : Synthesis and pharmacological evaluation of 6-naltrexamine analogs for alcohol cessation. Authors : Ghirmai S, Azar MR, Cashman JR. Publication Year :
2009 Journal : Bioorg. Med. Chem. Abstract : A series of substituted aryl amide derivatives of 6-naltrexamine, 3 designed to be metabolically stable were synthesized and used to characterize the structural requirements for their potency to binding and functional activity of human mu (mu), delta (delta) and kappa (kappa) opioid and nociceptin (NOP) receptors. Binding assays showed that 4-10 had subnanomolar K(i) values for mu and kappa opioid receptors. Functional assays for stimulation of [(35)S]GTPgammaS binding showed that several compounds acted as partial or inverse agonists and antagonists of the mu and delta, kappa opioid or NOP receptors. The compounds showed considerable stability in the presence of rat, mouse or human liver preparations and NADPH. The inhibitory activity on the functional activity of human cytochrome P450s was examined to determine any potential inhibition by 4-9. Only modest inhibition of CYP3A4, CYP2C9 and CYP2C19 was observed for a few of the analogs. As a representative example, radiolabeled 6 was examined in vivo and showed reasonable brain penetration. The inhibition of ethanol self-administration in rats trained to self-administer a 10% (w/v) ethanol solution, utilizing operant techniques showed 5-8 to have very potent efficacy (ED(50) values 19-50 microg/kg). DOI : 10.1016/j.bmc.2009.07.069
NEFAZODONE
Time dependent inhibition of CYP3A4 in human liver microsomes
Liver
IC50 = 15.0 nM
Reference
Title : Mechanism-based inactivation (MBI) of cytochrome P450 enzymes: structure-activity relationships and discovery strategies to mitigate drug-drug interaction risks. Authors : Orr ST, Ripp SL, Ballard TE, Henderson JL, Scott DO, Obach RS, Sun H, Kalgutkar AS. Publication Year :
2012 Journal : J. Med. Chem. DOI : 10.1021/jm300065h
NELFINAVIR
Mechanism based inhibition of human cytochrome P450 3A4 measured by testosterone hydroxylation
—
Ki = 480.0 nM
Reference
Title : Cytochrome p450 enzymes mechanism based inhibitors: common sub-structures and reactivity. Authors : Fontana E, Dansette PM, Poli SM. Publication Year :
2005 Journal : Curr. Drug Metab. Abstract : The inhibition of human cytochrome P450s (CYPs) is one of the most common mechanisms which can lead to drug-drug interactions. The inhibition of CYPs can be reversible (competitive or non-competitive) or irreversible. Irreversible inhibition usually derives from activation of a drug by CYPs into a reactive metabolite, which tightly binds to the enzyme active site, leading to a long lasting inactivation. This process is called "mechanism based inhibition" or "suicide inhibition". The irreversible inactivation usually implies the formation of a covalent bond between the metabolite and the enzyme, which can lead to hapten formation and can in some cases trigger an autoimmune-response. For these reasons it is of utmost importance to study the mechanism of the CYP inhibition of new potential drugs as early as possible during the drug discovery process. The literature on CYPs is vast and covers numerous aspects of their biology and biochemistry, however to our knowledge there is no general and systematic review focusing on mechanism-based inhibitors; we have reviewed the literature and compiled all the available data on chemical entities, which are known to be CYP suicide inhibitors. Each compound is reported together with its chemical structure, the CYP isoform and the parameters describing the inactivation. Literature references are reported together with their PMID (PubMed ID number) to allow a fast retrieval of the papers. This review offers a quick reference to help predict liabilities of new chemical entities without carrying out extensive in vitro work, and will hopefully help in designing safer drugs. DOI : 10.2174/138920005774330639
NICARDIPINE
Inhibition of CYP3A4 in human liver microsomes using testosterone substrate by LC-MS/MS method
Liver
IC50 = 380.0 nM
Reference
Title : Discovery and characterization of novel, potent, and selective cytochrome P450 2J2 inhibitors. Authors : Ren S, Zeng J, Mei Y, Zhang JZ, Yan SF, Fei J, Chen L. Publication Year :
2013 Journal : Drug Metab. Dispos. Abstract : Cytochrome P450 (CYP) 2J2 is one of the human CYPs involved in phase I xenobiotics metabolism. It is mainly expressed in extrahepatic tissues, including intestine and cardiovascular systems. The general role of CYP2J2 in drug metabolism is not yet fully understood, and the recent discovery that CYP2J2 can metabolize a wide range of structurally diverse drugs and its primary distribution in the intestine suggest its potentially indispensable role in first-pass intestinal metabolism and involvement in drug-drug interaction. To fully characterize its role in drug metabolism, selective and potent inhibitors of CYP2J2 are necessary tools. In the current study, 69 known drugs were screened for the inhibition of CYP2J2, and we discovered a number of marketed drugs as potent and selective CYP2J2 inhibitors. In particular, telmisartan and flunarizine have CYP2J2 inhibition IC(50) values of 0.42 μM and 0.94 μM, respectively, which are at least 10-fold more selective against all other major metabolizing CYPs; moreover, they are not substrates of CYP2J2 and show no time-dependent inhibition toward this CYP. The results of enzyme kinetics studies, supported by molecular modeling, have also elucidated that telmisartan is a mixed-type inhibitor, and flunarizine competitively inhibits CYP2J2. The K(i) for telmisartan is 0.19 μM, with an α value, an indicator of the type of inhibition mechanism, of 2.80, and flunarizine has a K(i) value of 0.13 μM. These newly discovered CYP2J2 inhibitors can be potentially used as a tool to study CYP2J2 in drug metabolism and interaction in a clinical setting. DOI : 10.1124/dmd.112.048264
NILOTINIB
Inhibition of human recombinant CYP3A4 expressed in insect cell microsomes in presence of NADPH by fluorescence assay
—
IC50 = 580.0 nM
Reference
Title : Accelerated Discovery of Novel Ponatinib Analogs with Improved Properties for the Treatment of Parkinson's Disease. Authors : Kaiser TM, Dentmon ZW, Dalloul CE, Sharma SK, Liotta DC. Publication Year :
2020 Journal : ACS Med Chem Lett Abstract : Parkinson's disease (PD) is a debilitating and common neurodegenerative disease. New insights implicating c-Abl activation as a driving force in PD have opened a new drug development avenue for PD treatment beyond the symptomatic relief by L-DOPA. BCR-Abl inhibitors, which include nilotinib and ponatinib, have been found to inhibit this process, and nilotinib has shown improvement in outcomes in a 12-patient, nonrandomized trial. However, nilotinib is a potent inhibitor of hERG, a cardiac K<sup>+</sup> channel whose inhibition increases risk of sudden death. We used our machine learning approach to predict novel molecules that would inhibit c-Abl while also having minimal liability against hERG. Of our six novel compounds tested, we identified two that had c-Abl potencies comparable to nilotinib, but with significantly improved profiles regarding the hERG channel. Our best compound exhibited a hERG IC<sub>50</sub> of 12.1 μM (compared to nilotinib with an IC<sub>50</sub> of 0.45 μM and ponatinib with IC<sub>50</sub> of 0.767 μM). This work is a step forward for a machine learning enabled, multiparameter optimization of a chemical space and represents a significant advance in the development of novel Parkinson's therapies. DOI : 10.1021/acsmedchemlett.9b00612
OLMESARTAN
Time dependent inhibition of CYP3A4 (unknown origin) at 100 uM by LC/MS system
—
Inhibition < 10.0 %
Reference
Title : Combination of GSH trapping and time-dependent inhibition assays as a predictive method of drugs generating highly reactive metabolites. Authors : Nakayama S, Takakusa H, Watanabe A, Miyaji Y, Suzuki W, Sugiyama D, Shiosakai K, Honda K, Okudaira N, Izumi T, Okazaki O. Publication Year :
2011 Journal : Drug Metab. Dispos. Abstract : Covalent binding (CB) of reactive metabolites (RMs) is potentially involved in severe adverse drug reactions. Because the CB assay is of low throughput and costly, a qualitative trapping assay using agents such as [(35)S]GSH is often performed in the early stages of drug discovery. However, trapping methods alone cannot replace the CB assay. We hypothesized that the time-dependent inhibition (TDI) assay might be complementary to the [(35)S]GSH trapping assay in detecting RMs. We performed CB assays, [(35)S]GSH trapping assays, and TDI assays for 42 structurally diverse compounds. First, we showed that the [(35)S]GSH trapping assay alone does not correlate with the extent of CB. Four compounds that the [(35)S]GSH trapping assay failed to detect but that showed high extent of CB were inactivators of the enzyme in the TDI assay. There was a tendency for compounds judged as positive in the TDI assay to show a high degree of CB irrespective of the result of the [(35)S]GSH trapping assay. Finally, to combine parameters from the two assays, we introduced intrinsic clearance to describe the formation of RMs (CL(int, RMs)). The Spearman rank correlation coefficient between the extent of CB and CL(int, RMs) was 0.77 (p < 0.0001), which was better than that for the formation rates of [(35)S]GSH adducts. Therefore, we demonstrated that a combination of the [(35)S]GSH trapping and TDI assays is an effective method for detecting compounds potentially capable of generating highly reactive metabolites in the early stages of drug discovery. DOI : 10.1124/dmd.111.039180
OSI-930
Inhibition of human His-tagged CYP3A4 expressed in Escherichia coli TOPP3 using BFC as substrate at 100 uM after 30 mins in presence of NADPH and cytochrome b5
—
Inhibition = 75.0 %
Reference
Title : Inactivation of cytochrome P450 (P450) 3A4 but not P450 3A5 by OSI-930, a thiophene-containing anticancer drug. Authors : Lin HL, Zhang H, Medower C, Hollenberg PF, Johnson WW. Publication Year :
2011 Journal : Drug Metab. Dispos. Abstract : An investigational anticancer agent that contains a thiophene moiety, 3-[(quinolin-4-ylmethyl)-amino]-N-[4-trifluoromethox)phenyl] thiophene-2-carboxamide (OSI-930), was tested to investigate its ability to modulate the activities of several cytochrome P450 enzymes. Results showed that OSI-930 inactivated purified, recombinant cytochrome P450 (P450) 3A4 in the reconstituted system in a mechanism-based manner. The inactivation was dependent on cytochrome b(5) and required NADPH. Catalase did not protect against the inactivation. No inactivation was observed in studies with human 2B6, 2D6, or 3A5 either in the presence or in the absence of b(5). The inactivation of 3A4 by OSI-930 was time- and concentration-dependent. The inactivation of the 7-benzyloxy-4-(trifluoromethyl)coumarin catalytic activity of 3A4 was characterized by a K(I) of 24 μM and a k(inact) of 0.04 min(-1). This K(I) is significantly greater than the clinical OSI-930 C(max) of 1.7 μM at the maximum tolerated dose, indicating that clinical drug interactions of OSI-930 via this pathway are not likely. Spectral analysis of the inactivated protein indicated that the decrease in the reduced CO spectrum at 450 nm was comparable to the amount of inactivation, thereby suggesting that the inactivation was primarily due to modification of the heme. High-pressure liquid chromatography (HPLC) analysis with detection at 400 nm showed a loss of heme comparable to the activity loss, but a modified heme was not detected. This result suggests either that the heme must have been modified enough so as not to be observed in a HPLC chromatograph or, possibly, that it was destroyed. The partition ratio for the inactivation of P450 3A4 was approximately 23, suggesting that this P450 3A4-mediated pathway occurs with approximately 4% frequency during the metabolism of OSI-930. Modeling studies on the binding of OSI-930 to the active site of the P450 3A4 indicated that OSI-930 would be oriented properly in the active site for oxidation of the thiophene sulfur to give the sulfoxide, which has previously been shown to be a significant metabolite of OSI-930. Because OSI-930 is an inactivator of P450 3A4 but does not exhibit any effect on P450 3A5 activity under the same conditions, it may be an appropriate probe for exploring unique aspects of these two very similar P450s. DOI : 10.1124/dmd.110.034074
OSI-930
Inhibition of human His-tagged CYP3A4 expressed in Escherichia coli TOPP3 using BFC as substrate at 100 uM after 30 mins by spectrophotometry in presence of NADPH and cytochrome b5 relative to control
—
Inhibition = 70.0 %
Reference
Title : Inactivation of cytochrome P450 (P450) 3A4 but not P450 3A5 by OSI-930, a thiophene-containing anticancer drug. Authors : Lin HL, Zhang H, Medower C, Hollenberg PF, Johnson WW. Publication Year :
2011 Journal : Drug Metab. Dispos. Abstract : An investigational anticancer agent that contains a thiophene moiety, 3-[(quinolin-4-ylmethyl)-amino]-N-[4-trifluoromethox)phenyl] thiophene-2-carboxamide (OSI-930), was tested to investigate its ability to modulate the activities of several cytochrome P450 enzymes. Results showed that OSI-930 inactivated purified, recombinant cytochrome P450 (P450) 3A4 in the reconstituted system in a mechanism-based manner. The inactivation was dependent on cytochrome b(5) and required NADPH. Catalase did not protect against the inactivation. No inactivation was observed in studies with human 2B6, 2D6, or 3A5 either in the presence or in the absence of b(5). The inactivation of 3A4 by OSI-930 was time- and concentration-dependent. The inactivation of the 7-benzyloxy-4-(trifluoromethyl)coumarin catalytic activity of 3A4 was characterized by a K(I) of 24 μM and a k(inact) of 0.04 min(-1). This K(I) is significantly greater than the clinical OSI-930 C(max) of 1.7 μM at the maximum tolerated dose, indicating that clinical drug interactions of OSI-930 via this pathway are not likely. Spectral analysis of the inactivated protein indicated that the decrease in the reduced CO spectrum at 450 nm was comparable to the amount of inactivation, thereby suggesting that the inactivation was primarily due to modification of the heme. High-pressure liquid chromatography (HPLC) analysis with detection at 400 nm showed a loss of heme comparable to the activity loss, but a modified heme was not detected. This result suggests either that the heme must have been modified enough so as not to be observed in a HPLC chromatograph or, possibly, that it was destroyed. The partition ratio for the inactivation of P450 3A4 was approximately 23, suggesting that this P450 3A4-mediated pathway occurs with approximately 4% frequency during the metabolism of OSI-930. Modeling studies on the binding of OSI-930 to the active site of the P450 3A4 indicated that OSI-930 would be oriented properly in the active site for oxidation of the thiophene sulfur to give the sulfoxide, which has previously been shown to be a significant metabolite of OSI-930. Because OSI-930 is an inactivator of P450 3A4 but does not exhibit any effect on P450 3A5 activity under the same conditions, it may be an appropriate probe for exploring unique aspects of these two very similar P450s. DOI : 10.1124/dmd.110.034074
PACRITINIB
Inhibition of CYP3A4 in human liver microsomes assessed as decrease in formation of 1-hydroxymidazolam from midazolam substrate at 5 uM after 5 mins by LC-MS/MS
Liver
Inhibition = 20.0 %
Reference
Title : Discovery of the macrocycle 11-(2-pyrrolidin-1-yl-ethoxy)-14,19-dioxa-5,7,26-triaza-tetracyclo[19.3.1.1(2,6).1(8,12)]heptacosa-1(25),2(26),3,5,8,10,12(27),16,21,23-decaene (SB1518), a potent Janus kinase 2/fms-like tyrosine kinase-3 (JAK2/FLT3) inhibitor for the treatment of myelofibrosis and lymphoma. Authors : William AD, Lee AC, Blanchard S, Poulsen A, Teo EL, Nagaraj H, Tan E, Chen D, Williams M, Sun ET, Goh KC, Ong WC, Goh SK, Hart S, Jayaraman R, Pasha MK, Ethirajulu K, Wood JM, Dymock BW. Publication Year :
2011 Journal : J. Med. Chem. Abstract : Discovery of the activating mutation V617F in Janus Kinase 2 (JAK2(V617F)), a tyrosine kinase critically involved in receptor signaling, recently ignited interest in JAK2 inhibitor therapy as a treatment for myelofibrosis (MF). Herein, we describe the design and synthesis of a series of small molecule 4-aryl-2-aminopyrimidine macrocycles and their biological evaluation against the JAK family of kinase enzymes and FLT3. The most promising leads were assessed for their in vitro ADME properties culminating in the discovery of 21c, a potent JAK2 (IC(50) = 23 and 19 nM for JAK2(WT) and JAK2(V617F), respectively) and FLT3 (IC(50) = 22 nM) inhibitor with selectivity against JAK1 and JAK3 (IC(50) = 1280 and 520 nM, respectively). Further profiling of 21c in preclinical species and mouse xenograft and allograft models is described. Compound 21c (SB1518) was selected as a development candidate and progressed into clinical trials where it is currently in phase 2 for MF and lymphoma. DOI : 10.1021/jm200326p
PF-05105679
Inhibition of CYP3A4 (unknown origin) at 3 uM
—
Inhibition = 0.0 %
Reference
Title : Discovery of a Selective TRPM8 Antagonist with Clinical Efficacy in Cold-Related Pain. Authors : Andrews MD, Af Forselles K, Beaumont K, Galan SR, Glossop PA, Grenie M, Jessiman A, Kenyon AS, Lunn G, Maw G, Owen RM, Pryde DC, Roberts D, Tran TD. Publication Year :
2015 Journal : ACS Med. Chem. Lett. Abstract : The transient receptor potential (TRP) family of ion channels comprises nonselective cation channels that respond to a wide range of chemical and thermal stimuli. TRPM8, a member of the melastatin subfamily, is activated by cold temperatures (<28 °C), and antagonists of this channel have the potential to treat cold induced allodynia and hyperalgesia. However, TRPM8 has also been implicated in mammalian thermoregulation and antagonists have the potential to induce hypothermia in patients. We report herein the identification and optimization of a series of TRPM8 antagonists that ultimately led to the discovery of PF-05105679. The clinical finding with this compound will be discussed, including both efficacy and its ability to affect thermoregulation processes in humans. DOI : 10.1021/ml500479v
PIPERAQUINE
Inhibition of CYP3A4 (unknown origin)
—
Ki = 90.0 nM
Reference
Title : The Development Process for Discovery and Clinical Advancement of Modern Antimalarials. Authors : Ashton TD, Devine SM, Möhrle JJ, Laleu B, Burrows JN, Charman SA, Creek DJ, Sleebs BE. Publication Year :
2019 Journal : J Med Chem Abstract : Malaria is a devastating disease caused by <i>Plasmodium</i> parasites, resulting in approximately 435000 deaths in 2018. The impact of malaria is compounded by the emergence of widespread resistance to current antimalarial therapies. Recently, a new strategy was initiated to screen small molecule collections against the <i>Plasmodium</i> parasite enabling the identification of new antimalarial chemotypes with novel modes of action. This initiative ushered in the modern era of antimalarial drug development, and as a result, numerous lead candidates are advancing toward or are currently in human clinical trials. In this Perspective, we describe the development pathway of four of the most clinically advanced modern antimalarials, KAE609, KAF156, DSM265, and MMV048. Additionally, the mechanism of action and life-cycle stage specificity of the four antimalarials is discussed in relation to aligning with global strategies to treat and eliminate malaria. This perspective serves as a guide to the expectations of modern antimalarial drug development. DOI : 10.1021/acs.jmedchem.9b00761
PRAVASTATIN
Time dependent inhibition of CYP3A4 (unknown origin) at 100 uM by LC/MS system
—
Inhibition < 10.0 %
Reference
Title : Combination of GSH trapping and time-dependent inhibition assays as a predictive method of drugs generating highly reactive metabolites. Authors : Nakayama S, Takakusa H, Watanabe A, Miyaji Y, Suzuki W, Sugiyama D, Shiosakai K, Honda K, Okudaira N, Izumi T, Okazaki O. Publication Year :
2011 Journal : Drug Metab. Dispos. Abstract : Covalent binding (CB) of reactive metabolites (RMs) is potentially involved in severe adverse drug reactions. Because the CB assay is of low throughput and costly, a qualitative trapping assay using agents such as [(35)S]GSH is often performed in the early stages of drug discovery. However, trapping methods alone cannot replace the CB assay. We hypothesized that the time-dependent inhibition (TDI) assay might be complementary to the [(35)S]GSH trapping assay in detecting RMs. We performed CB assays, [(35)S]GSH trapping assays, and TDI assays for 42 structurally diverse compounds. First, we showed that the [(35)S]GSH trapping assay alone does not correlate with the extent of CB. Four compounds that the [(35)S]GSH trapping assay failed to detect but that showed high extent of CB were inactivators of the enzyme in the TDI assay. There was a tendency for compounds judged as positive in the TDI assay to show a high degree of CB irrespective of the result of the [(35)S]GSH trapping assay. Finally, to combine parameters from the two assays, we introduced intrinsic clearance to describe the formation of RMs (CL(int, RMs)). The Spearman rank correlation coefficient between the extent of CB and CL(int, RMs) was 0.77 (p < 0.0001), which was better than that for the formation rates of [(35)S]GSH adducts. Therefore, we demonstrated that a combination of the [(35)S]GSH trapping and TDI assays is an effective method for detecting compounds potentially capable of generating highly reactive metabolites in the early stages of drug discovery. DOI : 10.1124/dmd.111.039180
PROCAINAMIDE
Time dependent inhibition of CYP3A4 (unknown origin) at 100 uM by LC/MS system
—
Inhibition < 10.0 %
Reference
Title : Combination of GSH trapping and time-dependent inhibition assays as a predictive method of drugs generating highly reactive metabolites. Authors : Nakayama S, Takakusa H, Watanabe A, Miyaji Y, Suzuki W, Sugiyama D, Shiosakai K, Honda K, Okudaira N, Izumi T, Okazaki O. Publication Year :
2011 Journal : Drug Metab. Dispos. Abstract : Covalent binding (CB) of reactive metabolites (RMs) is potentially involved in severe adverse drug reactions. Because the CB assay is of low throughput and costly, a qualitative trapping assay using agents such as [(35)S]GSH is often performed in the early stages of drug discovery. However, trapping methods alone cannot replace the CB assay. We hypothesized that the time-dependent inhibition (TDI) assay might be complementary to the [(35)S]GSH trapping assay in detecting RMs. We performed CB assays, [(35)S]GSH trapping assays, and TDI assays for 42 structurally diverse compounds. First, we showed that the [(35)S]GSH trapping assay alone does not correlate with the extent of CB. Four compounds that the [(35)S]GSH trapping assay failed to detect but that showed high extent of CB were inactivators of the enzyme in the TDI assay. There was a tendency for compounds judged as positive in the TDI assay to show a high degree of CB irrespective of the result of the [(35)S]GSH trapping assay. Finally, to combine parameters from the two assays, we introduced intrinsic clearance to describe the formation of RMs (CL(int, RMs)). The Spearman rank correlation coefficient between the extent of CB and CL(int, RMs) was 0.77 (p < 0.0001), which was better than that for the formation rates of [(35)S]GSH adducts. Therefore, we demonstrated that a combination of the [(35)S]GSH trapping and TDI assays is an effective method for detecting compounds potentially capable of generating highly reactive metabolites in the early stages of drug discovery. DOI : 10.1124/dmd.111.039180
PROPRANOLOL
Time dependent inhibition of CYP3A4 (unknown origin) at 100 uM by LC/MS system
—
Inhibition < 10.0 %
Reference
Title : Combination of GSH trapping and time-dependent inhibition assays as a predictive method of drugs generating highly reactive metabolites. Authors : Nakayama S, Takakusa H, Watanabe A, Miyaji Y, Suzuki W, Sugiyama D, Shiosakai K, Honda K, Okudaira N, Izumi T, Okazaki O. Publication Year :
2011 Journal : Drug Metab. Dispos. Abstract : Covalent binding (CB) of reactive metabolites (RMs) is potentially involved in severe adverse drug reactions. Because the CB assay is of low throughput and costly, a qualitative trapping assay using agents such as [(35)S]GSH is often performed in the early stages of drug discovery. However, trapping methods alone cannot replace the CB assay. We hypothesized that the time-dependent inhibition (TDI) assay might be complementary to the [(35)S]GSH trapping assay in detecting RMs. We performed CB assays, [(35)S]GSH trapping assays, and TDI assays for 42 structurally diverse compounds. First, we showed that the [(35)S]GSH trapping assay alone does not correlate with the extent of CB. Four compounds that the [(35)S]GSH trapping assay failed to detect but that showed high extent of CB were inactivators of the enzyme in the TDI assay. There was a tendency for compounds judged as positive in the TDI assay to show a high degree of CB irrespective of the result of the [(35)S]GSH trapping assay. Finally, to combine parameters from the two assays, we introduced intrinsic clearance to describe the formation of RMs (CL(int, RMs)). The Spearman rank correlation coefficient between the extent of CB and CL(int, RMs) was 0.77 (p < 0.0001), which was better than that for the formation rates of [(35)S]GSH adducts. Therefore, we demonstrated that a combination of the [(35)S]GSH trapping and TDI assays is an effective method for detecting compounds potentially capable of generating highly reactive metabolites in the early stages of drug discovery. DOI : 10.1124/dmd.111.039180
QUINIDINE
Inhibition of CYP3A4 in human liver microsomes assessed as midazolam hydroxylation to 1'-hydroxymidazolam at 10 uM after 10 mins relative to control
Liver
Inhibition = 36.0 %
Reference
Title : Rational development of 4-aminopyridyl-based inhibitors targeting Trypanosoma cruzi CYP51 as anti-chagas agents. Authors : Choi JY, Calvet CM, Gunatilleke SS, Ruiz C, Cameron MD, McKerrow JH, Podust LM, Roush WR. Publication Year :
2013 Journal : J. Med. Chem. Abstract : A new series of 4-aminopyridyl-based lead inhibitors targeting Trypanosoma cruzi CYP51 (TcCYP51) has been developed using structure-based drug design as well as structure-property relationship (SPR) analyses. The screening hit starting point, LP10 (KD ≤ 42 nM; EC50 = 0.65 μM), has been optimized to give the potential leads 14t, 27i, 27q, 27r, and 27t, which have low-nanomolar binding affinity to TcCYP51 and significant activity against T. cruzi amastigotes cultured in human myoblasts (EC50 = 14-18 nM for 27i and 27r). Many of the optimized compounds have improved microsome stability, and most are selective against human CYPs 1A2, 2D6, and 3A4 (<50% inhibition at 1 μM). A rationale for the improvement in microsome stability and selectivity of inhibitors against human metabolic CYP enzymes is presented. In addition, the binding mode of 14t with the Trypanosoma brucei CYP51 (TbCYP51) orthologue has been characterized by X-ray structure analysis. DOI : 10.1021/jm401067s
RELUGOLIX
Inhibition of human CYP3A4 assessed as conversion of testosterone to 6beta-hydroxytestosterone at 10 uM after 30 mins by HPLC analysis
—
Inhibition = 0.0 %
Reference
Title : Discovery of 1-{4-[1-(2,6-difluorobenzyl)-5-[(dimethylamino)methyl]-3-(6-methoxypyridazin-3-yl)-2,4-dioxo-1,2,3,4-tetrahydrothieno[2,3-d]pyrimidin-6-yl]phenyl}-3-methoxyurea (TAK-385) as a potent, orally active, non-peptide antagonist of the human gonadotropin-releasing hormone receptor. Authors : Miwa K, Hitaka T, Imada T, Sasaki S, Yoshimatsu M, Kusaka M, Tanaka A, Nakata D, Furuya S, Endo S, Hamamura K, Kitazaki T. Publication Year :
2011 Journal : J. Med. Chem. Abstract : We previously discovered an orally active human gonadotropin-releasing hormone (GnRH) receptor antagonist, thieno[2,3-d]pyrimidine-2,4-dione derivative 1 (sufugolix). To reduce the cytochrome P450 (CYP) inhibitory activity and improve in vivo GnRH antagonistic activity, further optimization of this scaffold was carried out. We focused our synthetic efforts on chemical modification at the 5 and 3 positions of the thieno[2,3-d]pyrimidine-2,4-dione ring based on computational modeling, which resulted in the discovery of 1-{4-[1-(2,6-difluorobenzyl)-5-[(dimethylamino)methyl]-3-(6-methoxypyridazin-3-yl)-2,4-dioxo-1,2,3,4-tetrahydrothieno[2,3-d]pyrimidin-6-yl]phenyl}-3-methoxyurea (16b) as a highly potent and orally active GnRH antagonist. Compound 16b showed potent in vitro GnRH antagonistic activity in the presence of fetal bovine serum (FBS) without CYP inhibition. Oral administration of 16b maintained the suppressive effect of the plasma luteinizing hormone levels in castrated cynomolgus monkeys at a 3 mg/kg dose for more than 24 h. Compound 16b is currently under clinical development with the code name of TAK-385. DOI : 10.1021/jm200216q
RIMONABANT
Inhibition of CYP3A4 in human liver microsomes assessed as inhibition of midazolam metabolism at 5 uM
Liver
Inhibition < 5.0 %
Reference
Title : Aminopyrazine CB1 receptor inverse agonists. Authors : Wustrow DJ, Maynard GD, Yuan J, Zhao H, Mao J, Guo Q, Kershaw M, Hammer J, Brodbeck RM, Near KE, Zhou D, Beers DS, Chenard BL, Krause JE, Hutchison AJ. Publication Year :
2008 Journal : Bioorg. Med. Chem. Lett. Abstract : A series of 5,6-diaryl-2-amino-pyrazines were prepared and found to have antagonist-like properties at the CB1 receptor. Subsequent SAR studies optimized both receptor potency and drug-like properties including solubility and Cytochrome-P450 inhibition potential. Optimized compounds were demonstrated to be inverse agonists and compared in vivo with rimonabant for their ability to inhibit food intake, to occupy central CB1 receptors and to influence hormonal markers associated with obesity. DOI : 10.1016/j.bmcl.2008.04.022
RIMONABANT
Inhibition of CYP3A4 in human liver microsomes after 30 mins
Liver
IC50 > 20.0 nM
Reference
Title : Discovery of 2-(4-((1H-1,2,4-triazol-1-yl)methyl)-5-(4-bromophenyl)-1-(2-chlorophenyl)-1H-pyrazol-3-yl)-5-tert-butyl-1,3,4-thiadiazole (GCC2680) as a potent, selective and orally efficacious cannabinoid-1 receptor antagonist. Authors : Lee J, Seo HJ, Lee SH, Kim J, Jung ME, Lee SH, Song KS, Lee J, Kang SY, Kim MJ, Kim MS, Son EJ, Lee M, Han HK. Publication Year :
2010 Journal : Bioorg. Med. Chem. Abstract : Structure-activity relationship studies in a series of diarylpyrazolyl thiadiazoles identified cannabinoid-1 receptor antagonists with excellent potency and selectivity. Based on its exceptional in vivo efficacy in animal models and its favorable pharmacokinetic and toxicological profiles, 2-(4-((1H-1,2,4-triazol-1-yl)methyl)-5-(4-bromophenyl)-1-(2-chlorophenyl)-1H-pyrazol-3-yl)-5-tert-butyl-1,3,4-thiadiazole (GCC2680) was selected as a preclinical candidate for the treatment of obesity. DOI : 10.1016/j.bmc.2010.07.013
ROSIGLITAZONE
Time dependent inhibition of CYP3A4 (unknown origin) at 100 uM by LC/MS system
—
Inhibition = 56.0 %
Reference
Title : Combination of GSH trapping and time-dependent inhibition assays as a predictive method of drugs generating highly reactive metabolites. Authors : Nakayama S, Takakusa H, Watanabe A, Miyaji Y, Suzuki W, Sugiyama D, Shiosakai K, Honda K, Okudaira N, Izumi T, Okazaki O. Publication Year :
2011 Journal : Drug Metab. Dispos. Abstract : Covalent binding (CB) of reactive metabolites (RMs) is potentially involved in severe adverse drug reactions. Because the CB assay is of low throughput and costly, a qualitative trapping assay using agents such as [(35)S]GSH is often performed in the early stages of drug discovery. However, trapping methods alone cannot replace the CB assay. We hypothesized that the time-dependent inhibition (TDI) assay might be complementary to the [(35)S]GSH trapping assay in detecting RMs. We performed CB assays, [(35)S]GSH trapping assays, and TDI assays for 42 structurally diverse compounds. First, we showed that the [(35)S]GSH trapping assay alone does not correlate with the extent of CB. Four compounds that the [(35)S]GSH trapping assay failed to detect but that showed high extent of CB were inactivators of the enzyme in the TDI assay. There was a tendency for compounds judged as positive in the TDI assay to show a high degree of CB irrespective of the result of the [(35)S]GSH trapping assay. Finally, to combine parameters from the two assays, we introduced intrinsic clearance to describe the formation of RMs (CL(int, RMs)). The Spearman rank correlation coefficient between the extent of CB and CL(int, RMs) was 0.77 (p < 0.0001), which was better than that for the formation rates of [(35)S]GSH adducts. Therefore, we demonstrated that a combination of the [(35)S]GSH trapping and TDI assays is an effective method for detecting compounds potentially capable of generating highly reactive metabolites in the early stages of drug discovery. DOI : 10.1124/dmd.111.039180
TALAZOPARIB
Inhibition of CYP3A4 (unknown origin) at 10 uM
—
Inhibition < 12.0 %
Reference
Title : Discovery and Characterization of (8S,9R)-5-Fluoro-8-(4-fluorophenyl)-9-(1-methyl-1H-1,2,4-triazol-5-yl)-2,7,8,9-tetrahydro-3H-pyrido[4,3,2-de]phthalazin-3-one (BMN 673, Talazoparib), a Novel, Highly Potent, and Orally Efficacious Poly(ADP-ribose) Polymerase-1/2 Inhibitor, as an Anticancer Agent. Authors : Wang B, Chu D, Feng Y, Shen Y, Aoyagi-Scharber M, Post LE. Publication Year :
2016 Journal : J. Med. Chem. Abstract : We discovered and developed a novel series of tetrahydropyridophthlazinones as poly(ADP-ribose) polymerase (PARP) 1 and 2 inhibitors. Lead optimization led to the identification of (8S,9R)-47 (talazoparib; BMN 673; (8S,9R)-5-fluoro-8-(4-fluorophenyl)-9-(1-methyl-1H-1,2,4-triazol-5-yl)-2,7,8,9-tetrahydro-3H-pyrido[4,3,2-de]phthalazin-3-one). The novel stereospecific dual chiral-center-embedded structure of this compound has enabled extensive and unique binding interactions with PARP1/2 proteins. (8S,9R)-47 demonstrates excellent potency, inhibiting PARP1 and PARP2 enzyme activity with Ki = 1.2 and 0.87 nM, respectively. It inhibits PARP-mediated PARylation in a whole-cell assay with an EC50 of 2.51 nM and prevents proliferation of cancer cells carrying mutant BRCA1/2, with EC50 = 0.3 nM (MX-1) and 5 nM (Capan-1), respectively. (8S,9R)-47 is orally available, displaying favorable pharmacokinetic (PK) properties and remarkable antitumor efficacy in the BRCA1 mutant MX-1 breast cancer xenograft model following oral administration as a single-agent or in combination with chemotherapy agents such as temozolomide and cisplatin. (8S,9R)-47 has completed phase 1 clinical trial and is currently being studied in phase 2 and 3 clinical trials for the treatment of locally advanced and/or metastatic breast cancer with germline BRCA1/2 deleterious mutations. DOI : 10.1021/acs.jmedchem.5b01498
TAMOXIFEN
Time dependent inhibition of CYP3A4 (unknown origin) at 100 uM by LC/MS system
—
Inhibition < 10.0 %
Reference
Title : Combination of GSH trapping and time-dependent inhibition assays as a predictive method of drugs generating highly reactive metabolites. Authors : Nakayama S, Takakusa H, Watanabe A, Miyaji Y, Suzuki W, Sugiyama D, Shiosakai K, Honda K, Okudaira N, Izumi T, Okazaki O. Publication Year :
2011 Journal : Drug Metab. Dispos. Abstract : Covalent binding (CB) of reactive metabolites (RMs) is potentially involved in severe adverse drug reactions. Because the CB assay is of low throughput and costly, a qualitative trapping assay using agents such as [(35)S]GSH is often performed in the early stages of drug discovery. However, trapping methods alone cannot replace the CB assay. We hypothesized that the time-dependent inhibition (TDI) assay might be complementary to the [(35)S]GSH trapping assay in detecting RMs. We performed CB assays, [(35)S]GSH trapping assays, and TDI assays for 42 structurally diverse compounds. First, we showed that the [(35)S]GSH trapping assay alone does not correlate with the extent of CB. Four compounds that the [(35)S]GSH trapping assay failed to detect but that showed high extent of CB were inactivators of the enzyme in the TDI assay. There was a tendency for compounds judged as positive in the TDI assay to show a high degree of CB irrespective of the result of the [(35)S]GSH trapping assay. Finally, to combine parameters from the two assays, we introduced intrinsic clearance to describe the formation of RMs (CL(int, RMs)). The Spearman rank correlation coefficient between the extent of CB and CL(int, RMs) was 0.77 (p < 0.0001), which was better than that for the formation rates of [(35)S]GSH adducts. Therefore, we demonstrated that a combination of the [(35)S]GSH trapping and TDI assays is an effective method for detecting compounds potentially capable of generating highly reactive metabolites in the early stages of drug discovery. DOI : 10.1124/dmd.111.039180
TAMOXIFEN
Mechanism based inhibition of human cytochrome P450 3A4 measured by testosterone 6-beta hydroxylation
—
Ki = 200.0 nM
Reference
Title : Cytochrome p450 enzymes mechanism based inhibitors: common sub-structures and reactivity. Authors : Fontana E, Dansette PM, Poli SM. Publication Year :
2005 Journal : Curr. Drug Metab. Abstract : The inhibition of human cytochrome P450s (CYPs) is one of the most common mechanisms which can lead to drug-drug interactions. The inhibition of CYPs can be reversible (competitive or non-competitive) or irreversible. Irreversible inhibition usually derives from activation of a drug by CYPs into a reactive metabolite, which tightly binds to the enzyme active site, leading to a long lasting inactivation. This process is called "mechanism based inhibition" or "suicide inhibition". The irreversible inactivation usually implies the formation of a covalent bond between the metabolite and the enzyme, which can lead to hapten formation and can in some cases trigger an autoimmune-response. For these reasons it is of utmost importance to study the mechanism of the CYP inhibition of new potential drugs as early as possible during the drug discovery process. The literature on CYPs is vast and covers numerous aspects of their biology and biochemistry, however to our knowledge there is no general and systematic review focusing on mechanism-based inhibitors; we have reviewed the literature and compiled all the available data on chemical entities, which are known to be CYP suicide inhibitors. Each compound is reported together with its chemical structure, the CYP isoform and the parameters describing the inactivation. Literature references are reported together with their PMID (PubMed ID number) to allow a fast retrieval of the papers. This review offers a quick reference to help predict liabilities of new chemical entities without carrying out extensive in vitro work, and will hopefully help in designing safer drugs. DOI : 10.2174/138920005774330639
TARANABANT
Inhibition of CYP3A4 in human liver microsomes after 30 mins
Liver
IC50 > 20.0 nM
Reference
Title : Discovery of 2-(4-((1H-1,2,4-triazol-1-yl)methyl)-5-(4-bromophenyl)-1-(2-chlorophenyl)-1H-pyrazol-3-yl)-5-tert-butyl-1,3,4-thiadiazole (GCC2680) as a potent, selective and orally efficacious cannabinoid-1 receptor antagonist. Authors : Lee J, Seo HJ, Lee SH, Kim J, Jung ME, Lee SH, Song KS, Lee J, Kang SY, Kim MJ, Kim MS, Son EJ, Lee M, Han HK. Publication Year :
2010 Journal : Bioorg. Med. Chem. Abstract : Structure-activity relationship studies in a series of diarylpyrazolyl thiadiazoles identified cannabinoid-1 receptor antagonists with excellent potency and selectivity. Based on its exceptional in vivo efficacy in animal models and its favorable pharmacokinetic and toxicological profiles, 2-(4-((1H-1,2,4-triazol-1-yl)methyl)-5-(4-bromophenyl)-1-(2-chlorophenyl)-1H-pyrazol-3-yl)-5-tert-butyl-1,3,4-thiadiazole (GCC2680) was selected as a preclinical candidate for the treatment of obesity. DOI : 10.1016/j.bmc.2010.07.013
TICLOPIDINE
Time dependent inhibition of CYP3A4 (unknown origin) at 100 uM by LC/MS system
—
Inhibition = 26.0 %
Reference
Title : Combination of GSH trapping and time-dependent inhibition assays as a predictive method of drugs generating highly reactive metabolites. Authors : Nakayama S, Takakusa H, Watanabe A, Miyaji Y, Suzuki W, Sugiyama D, Shiosakai K, Honda K, Okudaira N, Izumi T, Okazaki O. Publication Year :
2011 Journal : Drug Metab. Dispos. Abstract : Covalent binding (CB) of reactive metabolites (RMs) is potentially involved in severe adverse drug reactions. Because the CB assay is of low throughput and costly, a qualitative trapping assay using agents such as [(35)S]GSH is often performed in the early stages of drug discovery. However, trapping methods alone cannot replace the CB assay. We hypothesized that the time-dependent inhibition (TDI) assay might be complementary to the [(35)S]GSH trapping assay in detecting RMs. We performed CB assays, [(35)S]GSH trapping assays, and TDI assays for 42 structurally diverse compounds. First, we showed that the [(35)S]GSH trapping assay alone does not correlate with the extent of CB. Four compounds that the [(35)S]GSH trapping assay failed to detect but that showed high extent of CB were inactivators of the enzyme in the TDI assay. There was a tendency for compounds judged as positive in the TDI assay to show a high degree of CB irrespective of the result of the [(35)S]GSH trapping assay. Finally, to combine parameters from the two assays, we introduced intrinsic clearance to describe the formation of RMs (CL(int, RMs)). The Spearman rank correlation coefficient between the extent of CB and CL(int, RMs) was 0.77 (p < 0.0001), which was better than that for the formation rates of [(35)S]GSH adducts. Therefore, we demonstrated that a combination of the [(35)S]GSH trapping and TDI assays is an effective method for detecting compounds potentially capable of generating highly reactive metabolites in the early stages of drug discovery. DOI : 10.1124/dmd.111.039180
TOPIROXOSTAT
Inhibition of CYP3A4 in human liver microsomes assessed as inhibition of testosterone hydroxylation at 10 uM by HPLC analysis
Liver
Inhibition = 18.6 %
Reference
Title : Discovery of 3-(2-cyano-4-pyridyl)-5-(4-pyridyl)-1,2,4-triazole, FYX-051 - a xanthine oxidoreductase inhibitor for the treatment of hyperuricemia [corrected]. Authors : Sato T, Ashizawa N, Matsumoto K, Iwanaga T, Nakamura H, Inoue T, Nagata O. Publication Year :
2009 Journal : Bioorg. Med. Chem. Lett. Abstract : Our previous study identified 2-[2-(2-methoxyethoxy)ethoxy]-5-[5-(2-methyl-4-pyridyl)-1H-[1,2,4] triazol-3-yl]benzonitrile (2)[corrected]as a safe and potent xanthine oxidoreductase (XOR) inhibitor for the treatment of hyperuricemia. Here, we synthesized a series of 3,5-dipyridyl-1,2,4-triazole derivatives and, in particular, examined their in vivo activity in lowering the serum uric acid levels in rats. As a result, we identified 3-(2-cyano-4-pyridyl)-5-(4-pyridyl)-1,2,4-triazole (FYX-051, compound 39) [corrected] to be one of the most potent XOR inhibitors; it exhibited an extremely potent in vivo activity, weak CYP3A4-inhibitory activity and a better pharmacokinetic profile than compound 2. Compound 39 is currently being evaluated in a phase 2 clinical trial. DOI : 10.1016/j.bmcl.2009.08.091
VERAPAMIL
Time dependent inhibition of CYP3A4 (unknown origin) at 100 uM by LC/MS system
—
Inhibition = 51.0 %
Reference
Title : Combination of GSH trapping and time-dependent inhibition assays as a predictive method of drugs generating highly reactive metabolites. Authors : Nakayama S, Takakusa H, Watanabe A, Miyaji Y, Suzuki W, Sugiyama D, Shiosakai K, Honda K, Okudaira N, Izumi T, Okazaki O. Publication Year :
2011 Journal : Drug Metab. Dispos. Abstract : Covalent binding (CB) of reactive metabolites (RMs) is potentially involved in severe adverse drug reactions. Because the CB assay is of low throughput and costly, a qualitative trapping assay using agents such as [(35)S]GSH is often performed in the early stages of drug discovery. However, trapping methods alone cannot replace the CB assay. We hypothesized that the time-dependent inhibition (TDI) assay might be complementary to the [(35)S]GSH trapping assay in detecting RMs. We performed CB assays, [(35)S]GSH trapping assays, and TDI assays for 42 structurally diverse compounds. First, we showed that the [(35)S]GSH trapping assay alone does not correlate with the extent of CB. Four compounds that the [(35)S]GSH trapping assay failed to detect but that showed high extent of CB were inactivators of the enzyme in the TDI assay. There was a tendency for compounds judged as positive in the TDI assay to show a high degree of CB irrespective of the result of the [(35)S]GSH trapping assay. Finally, to combine parameters from the two assays, we introduced intrinsic clearance to describe the formation of RMs (CL(int, RMs)). The Spearman rank correlation coefficient between the extent of CB and CL(int, RMs) was 0.77 (p < 0.0001), which was better than that for the formation rates of [(35)S]GSH adducts. Therefore, we demonstrated that a combination of the [(35)S]GSH trapping and TDI assays is an effective method for detecting compounds potentially capable of generating highly reactive metabolites in the early stages of drug discovery. DOI : 10.1124/dmd.111.039180
VERAPAMIL
Inhibition of human CYP3A4 preincubated for 10 mins followed by substrate and NADP addition measured after 30 mins by fluorescence assay
—
IC50 = 11.0 nM
Reference
Title : Authors : Publication Year : Journal :
VORAPAXAR
Activity of human recombinant CYP3A4 assessed as enzyme-mediated ((4aR,6R)-9-{2-[(1R,4S)-5-((S)-3-Fluoro-phenyl)-pyridin-2-yl]-vinyl}-1-methyl-3-oxo-dodecahydro-naphtho[2,3-c]furan-6-yl)-carbamic acid(R)-2-hydroxy-ethyl ester formation at 10 uM after 120 mins by LC-MS/MS analysis in presence of astemizole
—
Inhibition = 76.0 %
Reference
Title : Identification of human liver cytochrome P450 enzymes involved in the metabolism of SCH 530348 (Vorapaxar), a potent oral thrombin protease-activated receptor 1 antagonist. Authors : Ghosal A, Lu X, Penner N, Gao L, Ramanathan R, Chowdhury SK, Kishnani NS, Alton KB. Publication Year :
2011 Journal : Drug Metab. Dispos. Abstract : Vorapaxar (SCH 530348), a potent oral thrombin protease-activated receptor 1 antagonist, is being developed as an antiplatelet agent for patients with established vascular disease. The objective of this study was to identify the human liver cytochrome P450 (P450) enzyme(s) responsible for the metabolism of SCH 530348. Human liver microsomes metabolized SCH 530348 to M19, an amine metabolite formed via carbamate cleavage, and M20 (monohydroxy-SCH 530348). Recombinant human CYP3A4 exhibited the most activity (11.5% profiled radioactivity) for the formation of M19, followed by markedly less substrate conversion with CYP1A1 and CYP2C19. Trace levels of M19, a major excreted human metabolite, were detected with CYP1A2, CYP3A5, and CYP4F3A. Formation of M19 by human liver microsomes was inhibited 89% by ketoconazole (IC(50), 0.73 μM), 34% by tranylcypromine, and 89% by anti-CYP3A4 monoclonal antibody. There was a significant correlation between the rate of M19 formation and midazolam 1'-hydroxylation (r = 0.75) or M19 formation and testosterone 6β-hydroxylation (r = 0.92). The results of screening, inhibition, and correlation studies confirmed that CYP3A4 is the major P450 enzyme responsible for M19 formation from SCH 530348. In contrast, formation of M20, a major circulating human metabolite at steady state, was primarily catalyzed by CYP3A4 and CYP2J2. M20 is pharmacologically equipotent to SCH 530348, whereas M19 is an inactive metabolite. Formation of M20 by human liver microsomes was inhibited 89% by ketoconazole, 75% by astemizole (a CYP2J2 inhibitor), and 43% by CYP3A4 monoclonal antibody. These results suggest that CYP3A4 and CYP2J2 are both involved in the formation of M20 metabolite. DOI : 10.1124/dmd.110.035493
WARFARIN
Time dependent inhibition of CYP3A4 (unknown origin) at 100 uM by LC/MS system
—
Inhibition < 10.0 %
Reference
Title : Combination of GSH trapping and time-dependent inhibition assays as a predictive method of drugs generating highly reactive metabolites. Authors : Nakayama S, Takakusa H, Watanabe A, Miyaji Y, Suzuki W, Sugiyama D, Shiosakai K, Honda K, Okudaira N, Izumi T, Okazaki O. Publication Year :
2011 Journal : Drug Metab. Dispos. Abstract : Covalent binding (CB) of reactive metabolites (RMs) is potentially involved in severe adverse drug reactions. Because the CB assay is of low throughput and costly, a qualitative trapping assay using agents such as [(35)S]GSH is often performed in the early stages of drug discovery. However, trapping methods alone cannot replace the CB assay. We hypothesized that the time-dependent inhibition (TDI) assay might be complementary to the [(35)S]GSH trapping assay in detecting RMs. We performed CB assays, [(35)S]GSH trapping assays, and TDI assays for 42 structurally diverse compounds. First, we showed that the [(35)S]GSH trapping assay alone does not correlate with the extent of CB. Four compounds that the [(35)S]GSH trapping assay failed to detect but that showed high extent of CB were inactivators of the enzyme in the TDI assay. There was a tendency for compounds judged as positive in the TDI assay to show a high degree of CB irrespective of the result of the [(35)S]GSH trapping assay. Finally, to combine parameters from the two assays, we introduced intrinsic clearance to describe the formation of RMs (CL(int, RMs)). The Spearman rank correlation coefficient between the extent of CB and CL(int, RMs) was 0.77 (p < 0.0001), which was better than that for the formation rates of [(35)S]GSH adducts. Therefore, we demonstrated that a combination of the [(35)S]GSH trapping and TDI assays is an effective method for detecting compounds potentially capable of generating highly reactive metabolites in the early stages of drug discovery. DOI : 10.1124/dmd.111.039180
ARTEMISININ
Time-dependent inhibition of CYP3A4 in human liver microsomes
Liver
Inhibition < 10.0 %
Reference
Title : Evaluation of P450 inhibition and induction by artemisinin antimalarials in human liver microsomes and primary human hepatocytes. Authors : Xing J, Kirby BJ, Whittington D, Wan Y, Goodlett DR. Publication Year :
2012 Journal : Drug Metab. Dispos. Abstract : Artemisinin drugs have become the first-line antimalarials in areas of multidrug resistance. However, monotherapy with artemisinin drugs results in comparatively high recrudescence rates. Autoinduction of cytochrome P450 (P450)-mediated metabolism, resulting in reduced exposure, has been supposed to be the underlying mechanism. To better understand the autoinduction and metabolic drug-drug interactions (DDIs), we evaluated the P450s (particularly CYP2B6 and CYP3A4) inhibited or induced by two artemisinin drugs, Qing-hao-su (QHS) and dihydroartemisinin (DHA) using human liver microsome, recombinant P450 enzymes, and primary human hepatocytes. The results suggested that QHS was a weak reversible inhibitor of CYP2B6 (K(i) 4.6 μM), but not CYP3A4 (IC₅₀ ∼ 50 μM) and did not show measurable time-dependent inhibition of either CYP2B6 or CYP3A4. DHA inhibited neither CYP2B6 nor CYP3A4 (IC₅₀ > 125 μM). In addition, it was found that QHS induced the activity of CYP3A4 (E(max) 3.5-fold and EC₅₀ 5.9 μM) and CYP2B6 (E(max) 1.9-fold and EC₅₀ 0.6 μM). Of the other P450s, UDP glucuronosyltransferases, and transporters studied, QHS and DHA had no significant effect except for minor induction of mRNA expression of CYP1A2 (E(max) 7.9-fold and EC₅₀ 5.2 μM) and CYP2A6 (E(max) 11.7-fold and EC₅₀ 4.0 μM) by QHS. Quantitative prediction of P450-mediated DDIs indicate autoinduction of QHS clearance with the AUC(i)/AUC ratio decreasing to 59%, as a result of a 1.9-fold increase in CYP3A4 and a 1.6-fold increase in CYP2B6 activity. These data suggest that QHS drugs are potential inducers of P450 enzymes, and the possible drug interactions (or lack thereof) with artemisinin drugs may be clinically relevant. DOI : 10.1124/dmd.112.045765
IMIPRAMINE
Time dependent inhibition of CYP3A4 (unknown origin) at 100 uM by LC/MS system
—
Inhibition < 10.0 %
Reference
Title : Combination of GSH trapping and time-dependent inhibition assays as a predictive method of drugs generating highly reactive metabolites. Authors : Nakayama S, Takakusa H, Watanabe A, Miyaji Y, Suzuki W, Sugiyama D, Shiosakai K, Honda K, Okudaira N, Izumi T, Okazaki O. Publication Year :
2011 Journal : Drug Metab. Dispos. Abstract : Covalent binding (CB) of reactive metabolites (RMs) is potentially involved in severe adverse drug reactions. Because the CB assay is of low throughput and costly, a qualitative trapping assay using agents such as [(35)S]GSH is often performed in the early stages of drug discovery. However, trapping methods alone cannot replace the CB assay. We hypothesized that the time-dependent inhibition (TDI) assay might be complementary to the [(35)S]GSH trapping assay in detecting RMs. We performed CB assays, [(35)S]GSH trapping assays, and TDI assays for 42 structurally diverse compounds. First, we showed that the [(35)S]GSH trapping assay alone does not correlate with the extent of CB. Four compounds that the [(35)S]GSH trapping assay failed to detect but that showed high extent of CB were inactivators of the enzyme in the TDI assay. There was a tendency for compounds judged as positive in the TDI assay to show a high degree of CB irrespective of the result of the [(35)S]GSH trapping assay. Finally, to combine parameters from the two assays, we introduced intrinsic clearance to describe the formation of RMs (CL(int, RMs)). The Spearman rank correlation coefficient between the extent of CB and CL(int, RMs) was 0.77 (p < 0.0001), which was better than that for the formation rates of [(35)S]GSH adducts. Therefore, we demonstrated that a combination of the [(35)S]GSH trapping and TDI assays is an effective method for detecting compounds potentially capable of generating highly reactive metabolites in the early stages of drug discovery. DOI : 10.1124/dmd.111.039180
DILTIAZEM
Mechanism based inhibition of human cytochrome P450 3A4 measured by testosterone 6-beta hydroxylation
—
Ki = 500.0 nM
Reference
Title : Cytochrome p450 enzymes mechanism based inhibitors: common sub-structures and reactivity. Authors : Fontana E, Dansette PM, Poli SM. Publication Year :
2005 Journal : Curr. Drug Metab. Abstract : The inhibition of human cytochrome P450s (CYPs) is one of the most common mechanisms which can lead to drug-drug interactions. The inhibition of CYPs can be reversible (competitive or non-competitive) or irreversible. Irreversible inhibition usually derives from activation of a drug by CYPs into a reactive metabolite, which tightly binds to the enzyme active site, leading to a long lasting inactivation. This process is called "mechanism based inhibition" or "suicide inhibition". The irreversible inactivation usually implies the formation of a covalent bond between the metabolite and the enzyme, which can lead to hapten formation and can in some cases trigger an autoimmune-response. For these reasons it is of utmost importance to study the mechanism of the CYP inhibition of new potential drugs as early as possible during the drug discovery process. The literature on CYPs is vast and covers numerous aspects of their biology and biochemistry, however to our knowledge there is no general and systematic review focusing on mechanism-based inhibitors; we have reviewed the literature and compiled all the available data on chemical entities, which are known to be CYP suicide inhibitors. Each compound is reported together with its chemical structure, the CYP isoform and the parameters describing the inactivation. Literature references are reported together with their PMID (PubMed ID number) to allow a fast retrieval of the papers. This review offers a quick reference to help predict liabilities of new chemical entities without carrying out extensive in vitro work, and will hopefully help in designing safer drugs. DOI : 10.2174/138920005774330639
DILTIAZEM
Inhibition of CYP3A4-mediated N-demethylation in human liver microsomes
Liver
Ki = 100.0 nM
Reference
Title : Mechanism-based inactivation (MBI) of cytochrome P450 enzymes: structure-activity relationships and discovery strategies to mitigate drug-drug interaction risks. Authors : Orr ST, Ripp SL, Ballard TE, Henderson JL, Scott DO, Obach RS, Sun H, Kalgutkar AS. Publication Year :
2012 Journal : J. Med. Chem. DOI : 10.1021/jm300065h
DILTIAZEM
Time dependent inhibition of CYP3A4 in human liver microsomes using midazolam as substrate preincubated for 90 mins followed by 10-fold dilution and substrate addition by LC-MS/MS analysis in presence of NADPH
Liver
IC50 = 100.0 nM
Reference
Title : Sequential metabolism of AMG 487, a novel CXCR3 antagonist, results in formation of quinone reactive metabolites that covalently modify CYP3A4 Cys239 and cause time-dependent inhibition of the enzyme. Authors : Henne KR, Tran TB, VandenBrink BM, Rock DA, Aidasani DK, Subramanian R, Mason AK, Stresser DM, Teffera Y, Wong SG, Johnson MG, Chen X, Tonn GR, Wong BK. Publication Year :
2012 Journal : Drug Metab. Dispos. Abstract : CYP3A4-mediated biotransformation of (R)-N-(1-(3-(4-ethoxyphenyl)-4-oxo-3,4-dihydropyrido[2,3-d]pyrimidin-2-yl)ethyl)-N-(pyridin-3-ylmethyl)-2-(4-(trifluoromethoxy)phenyl)acetamide (AMG 487) was previously shown to generate an inhibitory metabolite linked to dose- and time-dependent pharmacokinetics in humans. Although in vitro activity loss assays failed to demonstrate CYP3A4 time-dependent inhibition (TDI) with AMG 487, its M2 phenol metabolite readily produced TDI when remaining activity was assessed using either midazolam or testosterone (K(I) = 0.73-0.74 μM, k(inact) = 0.088-0.099 min(-1)). TDI investigations using an IC(50) shift method successfully produced inhibition attributable to AMG 487, but only when preincubations were extended from 30 to 90 min. The shift magnitude was ∼3× for midazolam activity, but no shift was observed for testosterone activity. Subsequent partition ratio determinations conducted for M2 using recombinant CYP3A4 showed that inactivation was a relatively inefficient process (r = 36). CYP3A4-mediated biotransformation of [(3)H]M2 in the presence of GSH led to identification of two new metabolites, M4 and M5, which shifted focus away from M2 being directly responsible for TDI. M4 (hydroxylated M2) was further metabolized to form reactive intermediates that, upon reaction with GSH, produced isomeric adducts, collectively designated M5. Incubations conducted in the presence of [(18)O]H(2)O confirmed incorporation of oxygen from O(2) for the majority of M4 and M5 formed (>75%). Further evidence of a primary role for M4 in CYP3A4 TDI was generated by protein labeling and proteolysis experiments, in which M4 was found to be covalently bound to Cys239 of CYP3A4. These investigations confirmed a primarily role for M4 in CYP3A4 inactivation, suggesting that a more complex metabolic pathway was responsible for generation of inhibitory metabolites affecting AMG 487 human pharmacokinetics. DOI : 10.1124/dmd.112.045708
DILTIAZEM
Time dependent inhibition of CYP3A4 in human liver microsomes using testosterone as substrate preincubated for 90 mins followed by 10-fold dilution and substrate addition by LC-MS/MS analysis in presence of NADPH
Liver
IC50 = 100.0 nM
Reference
Title : Sequential metabolism of AMG 487, a novel CXCR3 antagonist, results in formation of quinone reactive metabolites that covalently modify CYP3A4 Cys239 and cause time-dependent inhibition of the enzyme. Authors : Henne KR, Tran TB, VandenBrink BM, Rock DA, Aidasani DK, Subramanian R, Mason AK, Stresser DM, Teffera Y, Wong SG, Johnson MG, Chen X, Tonn GR, Wong BK. Publication Year :
2012 Journal : Drug Metab. Dispos. Abstract : CYP3A4-mediated biotransformation of (R)-N-(1-(3-(4-ethoxyphenyl)-4-oxo-3,4-dihydropyrido[2,3-d]pyrimidin-2-yl)ethyl)-N-(pyridin-3-ylmethyl)-2-(4-(trifluoromethoxy)phenyl)acetamide (AMG 487) was previously shown to generate an inhibitory metabolite linked to dose- and time-dependent pharmacokinetics in humans. Although in vitro activity loss assays failed to demonstrate CYP3A4 time-dependent inhibition (TDI) with AMG 487, its M2 phenol metabolite readily produced TDI when remaining activity was assessed using either midazolam or testosterone (K(I) = 0.73-0.74 μM, k(inact) = 0.088-0.099 min(-1)). TDI investigations using an IC(50) shift method successfully produced inhibition attributable to AMG 487, but only when preincubations were extended from 30 to 90 min. The shift magnitude was ∼3× for midazolam activity, but no shift was observed for testosterone activity. Subsequent partition ratio determinations conducted for M2 using recombinant CYP3A4 showed that inactivation was a relatively inefficient process (r = 36). CYP3A4-mediated biotransformation of [(3)H]M2 in the presence of GSH led to identification of two new metabolites, M4 and M5, which shifted focus away from M2 being directly responsible for TDI. M4 (hydroxylated M2) was further metabolized to form reactive intermediates that, upon reaction with GSH, produced isomeric adducts, collectively designated M5. Incubations conducted in the presence of [(18)O]H(2)O confirmed incorporation of oxygen from O(2) for the majority of M4 and M5 formed (>75%). Further evidence of a primary role for M4 in CYP3A4 TDI was generated by protein labeling and proteolysis experiments, in which M4 was found to be covalently bound to Cys239 of CYP3A4. These investigations confirmed a primarily role for M4 in CYP3A4 inactivation, suggesting that a more complex metabolic pathway was responsible for generation of inhibitory metabolites affecting AMG 487 human pharmacokinetics. DOI : 10.1124/dmd.112.045708
ABIRATERONE
Inhibition of C-terminal His-tagged recombinant human CYP3A4 assessed as decrease in metabolism of luciferin isopropyl alcohol preincubated for 3 mins followed by NADPH addition measured after 30 mins
—
IC50 = 230.0 nM
Reference
Title : Structure-Based Design of Inhibitors with Improved Selectivity for Steroidogenic Cytochrome P450 17A1 over Cytochrome P450 21A2. Authors : Fehl C, Vogt CD, Yadav R, Li K, Scott EE, Aubé J. Publication Year :
2018 Journal : J Med Chem Abstract : Inhibition of androgen biosynthesis is clinically effective for treating androgen-responsive prostate cancer. Abiraterone is a clinical first-in-class inhibitor of cytochrome P450 17A1 (CYP17A1) required for androgen biosynthesis. However, abiraterone also causes hypertension, hypokalemia, and edema, likely due in part to off-target inhibition of another steroidogenic cytochrome P450, CYP21A2. Abiraterone analogs were designed based on structural evidence that B-ring substituents may favorably interact with polar residues in binding CYP17A1 and sterically clash with residues in the CYP21A2 active site. The best analogs increased selectivity of CYP17A1 inhibition up to 84-fold compared with 6.6-fold for abiraterone. Cocrystallization with CYP17A1 validated the intended new contacts with CYP17A1 active site residues. Docking these analogs into CYP21A2 identified steric clashes that likely underlie decreased binding and CYP21A2 inhibition. Overall, these analogs may offer a clinical advantage in the form of reduced side effects. DOI : 10.1021/acs.jmedchem.8b00419
CHLOROQUINE
Inhibition of CYP3A4 in human liver microsomes assessed as metabolite produced at 1 uM using midazolam as substrate by LC-MS/MS analysis relative to control
Liver
Inhibition = 105.0 %
Reference
Title : Development of a new generation of 4-aminoquinoline antimalarial compounds using predictive pharmacokinetic and toxicology models. Authors : Ray S, Madrid PB, Catz P, LeValley SE, Furniss MJ, Rausch LL, Guy RK, DeRisi JL, Iyer LV, Green CE, Mirsalis JC. Publication Year :
2010 Journal : J. Med. Chem. Abstract : Among the known antimalarial drugs, chloroquine (CQ) and other 4-aminoquinolines have shown high potency and good bioavailability. Yet complications associated with drug resistance necessitate the discovery of effective new antimalarial agents. ADMET prediction studies were employed to evaluate a library of new molecules based on the 4-aminoquinolone-related structure of CQ. Extensive in vitro screening and in vivo pharmacokinetic studies in mice helped to identify two lead molecules, 18 and 4, with promising in vitro therapeutic efficacy, improved ADMET properties, low risk for drug-drug interactions, and desirable pharmacokinetic profiles. Both 18 and 4 are highly potent antimalarial compounds, with IC(50) values of 5.6 and 17.3 nM, respectively, against the W2 (CQ-resistant) strain of Plasmodium falciparum (for CQ, IC(50) = 382 nM). When tested in mice, these compounds were found to have biological half-lives and plasma exposure values similar to or higher than those of CQ; they are therefore desirable candidates to pursue in future clinical trials. DOI : 10.1021/jm100057h
CHLOROQUINE
Inhibition of CYP3A4 in human liver microsomes assessed as metabolite produced at 1 uM using testosterone as substrate by LC-MS/MS analysis relative to control
Liver
Inhibition = 102.0 %
Reference
Title : Development of a new generation of 4-aminoquinoline antimalarial compounds using predictive pharmacokinetic and toxicology models. Authors : Ray S, Madrid PB, Catz P, LeValley SE, Furniss MJ, Rausch LL, Guy RK, DeRisi JL, Iyer LV, Green CE, Mirsalis JC. Publication Year :
2010 Journal : J. Med. Chem. Abstract : Among the known antimalarial drugs, chloroquine (CQ) and other 4-aminoquinolines have shown high potency and good bioavailability. Yet complications associated with drug resistance necessitate the discovery of effective new antimalarial agents. ADMET prediction studies were employed to evaluate a library of new molecules based on the 4-aminoquinolone-related structure of CQ. Extensive in vitro screening and in vivo pharmacokinetic studies in mice helped to identify two lead molecules, 18 and 4, with promising in vitro therapeutic efficacy, improved ADMET properties, low risk for drug-drug interactions, and desirable pharmacokinetic profiles. Both 18 and 4 are highly potent antimalarial compounds, with IC(50) values of 5.6 and 17.3 nM, respectively, against the W2 (CQ-resistant) strain of Plasmodium falciparum (for CQ, IC(50) = 382 nM). When tested in mice, these compounds were found to have biological half-lives and plasma exposure values similar to or higher than those of CQ; they are therefore desirable candidates to pursue in future clinical trials. DOI : 10.1021/jm100057h
CHLOROQUINE
Inhibition of CYP3A4 in human liver microsomes assessed as metabolite produced at 10 uM using midazolam as substrate by LC-MS/MS analysis relative to control
Liver
Inhibition = 110.0 %
Reference
Title : Development of a new generation of 4-aminoquinoline antimalarial compounds using predictive pharmacokinetic and toxicology models. Authors : Ray S, Madrid PB, Catz P, LeValley SE, Furniss MJ, Rausch LL, Guy RK, DeRisi JL, Iyer LV, Green CE, Mirsalis JC. Publication Year :
2010 Journal : J. Med. Chem. Abstract : Among the known antimalarial drugs, chloroquine (CQ) and other 4-aminoquinolines have shown high potency and good bioavailability. Yet complications associated with drug resistance necessitate the discovery of effective new antimalarial agents. ADMET prediction studies were employed to evaluate a library of new molecules based on the 4-aminoquinolone-related structure of CQ. Extensive in vitro screening and in vivo pharmacokinetic studies in mice helped to identify two lead molecules, 18 and 4, with promising in vitro therapeutic efficacy, improved ADMET properties, low risk for drug-drug interactions, and desirable pharmacokinetic profiles. Both 18 and 4 are highly potent antimalarial compounds, with IC(50) values of 5.6 and 17.3 nM, respectively, against the W2 (CQ-resistant) strain of Plasmodium falciparum (for CQ, IC(50) = 382 nM). When tested in mice, these compounds were found to have biological half-lives and plasma exposure values similar to or higher than those of CQ; they are therefore desirable candidates to pursue in future clinical trials. DOI : 10.1021/jm100057h
CHLOROQUINE
Inhibition of CYP3A4 in human liver microsomes assessed as metabolite produced at 10 uM using testosterone as substrate by LC-MS/MS analysis relative to control
Liver
Inhibition = 106.0 %
Reference
Title : Development of a new generation of 4-aminoquinoline antimalarial compounds using predictive pharmacokinetic and toxicology models. Authors : Ray S, Madrid PB, Catz P, LeValley SE, Furniss MJ, Rausch LL, Guy RK, DeRisi JL, Iyer LV, Green CE, Mirsalis JC. Publication Year :
2010 Journal : J. Med. Chem. Abstract : Among the known antimalarial drugs, chloroquine (CQ) and other 4-aminoquinolines have shown high potency and good bioavailability. Yet complications associated with drug resistance necessitate the discovery of effective new antimalarial agents. ADMET prediction studies were employed to evaluate a library of new molecules based on the 4-aminoquinolone-related structure of CQ. Extensive in vitro screening and in vivo pharmacokinetic studies in mice helped to identify two lead molecules, 18 and 4, with promising in vitro therapeutic efficacy, improved ADMET properties, low risk for drug-drug interactions, and desirable pharmacokinetic profiles. Both 18 and 4 are highly potent antimalarial compounds, with IC(50) values of 5.6 and 17.3 nM, respectively, against the W2 (CQ-resistant) strain of Plasmodium falciparum (for CQ, IC(50) = 382 nM). When tested in mice, these compounds were found to have biological half-lives and plasma exposure values similar to or higher than those of CQ; they are therefore desirable candidates to pursue in future clinical trials. DOI : 10.1021/jm100057h
CLOPIDOGREL
Time dependent inhibition of CYP3A4 (unknown origin) at 100 uM by LC/MS system
—
Inhibition = 35.0 %
Reference
Title : Combination of GSH trapping and time-dependent inhibition assays as a predictive method of drugs generating highly reactive metabolites. Authors : Nakayama S, Takakusa H, Watanabe A, Miyaji Y, Suzuki W, Sugiyama D, Shiosakai K, Honda K, Okudaira N, Izumi T, Okazaki O. Publication Year :
2011 Journal : Drug Metab. Dispos. Abstract : Covalent binding (CB) of reactive metabolites (RMs) is potentially involved in severe adverse drug reactions. Because the CB assay is of low throughput and costly, a qualitative trapping assay using agents such as [(35)S]GSH is often performed in the early stages of drug discovery. However, trapping methods alone cannot replace the CB assay. We hypothesized that the time-dependent inhibition (TDI) assay might be complementary to the [(35)S]GSH trapping assay in detecting RMs. We performed CB assays, [(35)S]GSH trapping assays, and TDI assays for 42 structurally diverse compounds. First, we showed that the [(35)S]GSH trapping assay alone does not correlate with the extent of CB. Four compounds that the [(35)S]GSH trapping assay failed to detect but that showed high extent of CB were inactivators of the enzyme in the TDI assay. There was a tendency for compounds judged as positive in the TDI assay to show a high degree of CB irrespective of the result of the [(35)S]GSH trapping assay. Finally, to combine parameters from the two assays, we introduced intrinsic clearance to describe the formation of RMs (CL(int, RMs)). The Spearman rank correlation coefficient between the extent of CB and CL(int, RMs) was 0.77 (p < 0.0001), which was better than that for the formation rates of [(35)S]GSH adducts. Therefore, we demonstrated that a combination of the [(35)S]GSH trapping and TDI assays is an effective method for detecting compounds potentially capable of generating highly reactive metabolites in the early stages of drug discovery. DOI : 10.1124/dmd.111.039180
PIOGLITAZONE
Time dependent inhibition of CYP3A4 (unknown origin) at 100 uM by LC/MS system
—
Inhibition = 61.0 %
Reference
Title : Combination of GSH trapping and time-dependent inhibition assays as a predictive method of drugs generating highly reactive metabolites. Authors : Nakayama S, Takakusa H, Watanabe A, Miyaji Y, Suzuki W, Sugiyama D, Shiosakai K, Honda K, Okudaira N, Izumi T, Okazaki O. Publication Year :
2011 Journal : Drug Metab. Dispos. Abstract : Covalent binding (CB) of reactive metabolites (RMs) is potentially involved in severe adverse drug reactions. Because the CB assay is of low throughput and costly, a qualitative trapping assay using agents such as [(35)S]GSH is often performed in the early stages of drug discovery. However, trapping methods alone cannot replace the CB assay. We hypothesized that the time-dependent inhibition (TDI) assay might be complementary to the [(35)S]GSH trapping assay in detecting RMs. We performed CB assays, [(35)S]GSH trapping assays, and TDI assays for 42 structurally diverse compounds. First, we showed that the [(35)S]GSH trapping assay alone does not correlate with the extent of CB. Four compounds that the [(35)S]GSH trapping assay failed to detect but that showed high extent of CB were inactivators of the enzyme in the TDI assay. There was a tendency for compounds judged as positive in the TDI assay to show a high degree of CB irrespective of the result of the [(35)S]GSH trapping assay. Finally, to combine parameters from the two assays, we introduced intrinsic clearance to describe the formation of RMs (CL(int, RMs)). The Spearman rank correlation coefficient between the extent of CB and CL(int, RMs) was 0.77 (p < 0.0001), which was better than that for the formation rates of [(35)S]GSH adducts. Therefore, we demonstrated that a combination of the [(35)S]GSH trapping and TDI assays is an effective method for detecting compounds potentially capable of generating highly reactive metabolites in the early stages of drug discovery. DOI : 10.1124/dmd.111.039180
ZOMEPIRAC
Time dependent inhibition of CYP3A4 (unknown origin) at 100 uM by LC/MS system
—
Inhibition < 10.0 %
Reference
Title : Combination of GSH trapping and time-dependent inhibition assays as a predictive method of drugs generating highly reactive metabolites. Authors : Nakayama S, Takakusa H, Watanabe A, Miyaji Y, Suzuki W, Sugiyama D, Shiosakai K, Honda K, Okudaira N, Izumi T, Okazaki O. Publication Year :
2011 Journal : Drug Metab. Dispos. Abstract : Covalent binding (CB) of reactive metabolites (RMs) is potentially involved in severe adverse drug reactions. Because the CB assay is of low throughput and costly, a qualitative trapping assay using agents such as [(35)S]GSH is often performed in the early stages of drug discovery. However, trapping methods alone cannot replace the CB assay. We hypothesized that the time-dependent inhibition (TDI) assay might be complementary to the [(35)S]GSH trapping assay in detecting RMs. We performed CB assays, [(35)S]GSH trapping assays, and TDI assays for 42 structurally diverse compounds. First, we showed that the [(35)S]GSH trapping assay alone does not correlate with the extent of CB. Four compounds that the [(35)S]GSH trapping assay failed to detect but that showed high extent of CB were inactivators of the enzyme in the TDI assay. There was a tendency for compounds judged as positive in the TDI assay to show a high degree of CB irrespective of the result of the [(35)S]GSH trapping assay. Finally, to combine parameters from the two assays, we introduced intrinsic clearance to describe the formation of RMs (CL(int, RMs)). The Spearman rank correlation coefficient between the extent of CB and CL(int, RMs) was 0.77 (p < 0.0001), which was better than that for the formation rates of [(35)S]GSH adducts. Therefore, we demonstrated that a combination of the [(35)S]GSH trapping and TDI assays is an effective method for detecting compounds potentially capable of generating highly reactive metabolites in the early stages of drug discovery. DOI : 10.1124/dmd.111.039180
DORAMAPIMOD
Inhibition of CYP3A4 (unknown origin) at 30 uM
—
Inhibition = -23.0 %
Reference
Title : Discovery of Narrow Spectrum Kinase Inhibitors: New Therapeutic Agents for the Treatment of COPD and Steroid-Resistant Asthma. Authors : Onions ST, Ito K, Charron CE, Brown RJ, Colucci M, Frickel F, Hardy G, Joly K, King-Underwood J, Kizawa Y, Knowles I, Murray PJ, Novak A, Rani A, Rapeport G, Smith A, Strong P, Taddei DM, Williams JG. Publication Year :
2016 Journal : J. Med. Chem. Abstract : The discovery of a novel series of therapeutic agents that has been designed and optimized for treating chronic obstructive pulmonary disease is reported. The pharmacological strategy was based on the identification of compounds that inhibit a defined subset of kinase enzymes modulating inflammatory processes that would be effective against steroid refractory disease and exhibit a sustained duration of action after inhaled delivery. DOI : 10.1021/acs.jmedchem.5b01029
AMINOPYRINE
Time dependent inhibition of CYP3A4 (unknown origin) at 100 uM by LC/MS system
—
Inhibition < 10.0 %
Reference
Title : Combination of GSH trapping and time-dependent inhibition assays as a predictive method of drugs generating highly reactive metabolites. Authors : Nakayama S, Takakusa H, Watanabe A, Miyaji Y, Suzuki W, Sugiyama D, Shiosakai K, Honda K, Okudaira N, Izumi T, Okazaki O. Publication Year :
2011 Journal : Drug Metab. Dispos. Abstract : Covalent binding (CB) of reactive metabolites (RMs) is potentially involved in severe adverse drug reactions. Because the CB assay is of low throughput and costly, a qualitative trapping assay using agents such as [(35)S]GSH is often performed in the early stages of drug discovery. However, trapping methods alone cannot replace the CB assay. We hypothesized that the time-dependent inhibition (TDI) assay might be complementary to the [(35)S]GSH trapping assay in detecting RMs. We performed CB assays, [(35)S]GSH trapping assays, and TDI assays for 42 structurally diverse compounds. First, we showed that the [(35)S]GSH trapping assay alone does not correlate with the extent of CB. Four compounds that the [(35)S]GSH trapping assay failed to detect but that showed high extent of CB were inactivators of the enzyme in the TDI assay. There was a tendency for compounds judged as positive in the TDI assay to show a high degree of CB irrespective of the result of the [(35)S]GSH trapping assay. Finally, to combine parameters from the two assays, we introduced intrinsic clearance to describe the formation of RMs (CL(int, RMs)). The Spearman rank correlation coefficient between the extent of CB and CL(int, RMs) was 0.77 (p < 0.0001), which was better than that for the formation rates of [(35)S]GSH adducts. Therefore, we demonstrated that a combination of the [(35)S]GSH trapping and TDI assays is an effective method for detecting compounds potentially capable of generating highly reactive metabolites in the early stages of drug discovery. DOI : 10.1124/dmd.111.039180
ECONAZOLE
Inhibition of CYP3A4 (unknown origin)
—
IC50 = 430.0 nM
Reference
Title : Strategies for the development of highly selective cytochrome P450 inhibitors: Several CYP targets in current research. Authors : Zhao L, Sun N, Tian L, Zhao S, Sun B, Sun Y, Zhao D. Publication Year :
2019 Journal : Bioorg Med Chem Lett Abstract : Cytochromes P450 (CYPs) play an important role in the metabolism of endogenic and xenobiotic substances, especially drugs. In addition, many CYPs may serve as targets for disease treatment. However, due to the presence of a common heme, the hydrophobicity of the CYP binding cavity, and the high homology within the binding pocket, most CYP inhibitors lack selectivity, which often leads to drug-drug interactions. Therefore, it is meaningful to develop highly selective CYP inhibitors. In this review, we summarize some of the strategies that have been used to develop highly selective CYP inhibitors, such as the weakening of the heme-binding group interaction, reduction of molecular lipophilicity and introduction of small structural changes within compounds. DOI : 10.1016/j.bmcl.2019.06.040
PF-00562271
Inhibition of CYP3A4 (unknown origin) at 10 uM relative to control
—
Inhibition = 47.2 %
Reference
Title : Authors : Publication Year : Journal :
LUMINESPIB
Inhibition of CYP3A4 in human liver microsomes at 10 uM after 20 mins
Liver
Inhibition = 12.0 %
Reference
Title : Discovery of potent N-(isoxazol-5-yl)amides as HSP90 inhibitors. Authors : Chen D, Shen A, Li J, Shi F, Chen W, Ren J, Liu H, Xu Y, Wang X, Yang X, Sun Y, Yang M, He J, Wang Y, Zhang L, Huang M, Geng M, Xiong B, Shen J. Publication Year :
2014 Journal : Eur. J. Med. Chem. Abstract : HSP90 is ubiquitously overexpressed in a broad spectrum of human cancers and has been recognized as an attractive target for cancer treatment. Here, we described the fragment screening, synthesis and structure-activity relationship studies of small molecule inhibitors with 4,5-diarylisoxazole scaffold targeting HSP90. Among them, the compound N-(3-(2,4-dihydroxy-5-isopropylphenyl)-4-(4-((4-morpholinopiperidin-1-yl)methyl)phenyl)isoxazol-5-yl)cyclopropanecarboxamide (108) showed high affinity for binding to HSP90 (FP binding assay, IC50 = 0.030 μM) and inhibited the proliferation of various human cancer cell lines with averaging GI50 about 88 nM. Compound 108 exhibited its functional inhibition of HSP90 by depleting key signaling pathways and concomitantly elevating of HSP70 and HSP27 in U-87MG cells. Further in vivo studies showed that compound 108 strongly suppressed the tumor growth of human glioblastoma xenograft model U-87MG with T/C = 18.35% at 50 mg/kg q3w/2.5w. Moreover, compound 108 also exhibited good pharmacokinetic properties. Together, our study implicates that compound 108 is a promising candidate of HSP90 inhibitor and is currently advanced to preclinical study. DOI : 10.1016/j.ejmech.2014.09.065
CHS-828
Inhibition of CYP3A4 in human liver microsomes using testosterone as substrate in presence of NADPH by LC-MS/MS analysis
Liver
IC50 = 220.0 nM
Reference
Title : Minimizing CYP2C9 Inhibition of Exposed-Pyridine NAMPT (Nicotinamide Phosphoribosyltransferase) Inhibitors. Authors : Zak M, Yuen PW, Liu X, Patel S, Sampath D, Oeh J, Liederer BM, Wang W, O'Brien T, Xiao Y, Skelton N, Hua R, Sodhi J, Wang Y, Zhang L, Zhao G, Zheng X, Ho YC, Bair KW, Dragovich PS. Publication Year :
2016 Journal : J Med Chem Abstract : NAMPT inhibitors may show potential as therapeutics for oncology. Throughout our NAMPT inhibitor program, we found that exposed pyridines or related heterocyclic systems in the left-hand portion of the inhibitors are necessary pharmacophores for potent cellular NAMPT inhibition. However, when combined with a benzyl group in the center of the inhibitors, such pyridine-like moieties also led to consistent and potent inhibition of CYP2C9. In an attempt to reduce CYP2C9 inhibition, a parallel synthesis approach was used to identify central benzyl group replacements with increased Fsp3. A spirocyclic central motif was thus discovered that was combined with left-hand pyridines (or pyridine-like systems) to provide cellularly potent NAMPT inhibitors with minimal CYP2C9 inhibition. Further optimization of potency and ADME properties led to the discovery of compound 68, a highly potent NAMPT inhibitor with outstanding efficacy in a mouse tumor xenograft model and lacking measurable CYP2C9 inhibition at the concentrations tested. DOI : 10.1021/acs.jmedchem.6b00697
CHS-828
Inhibition of CYP3A4 in human liver microsomes using midazolam as substrate in presence of NADPH by LC-MS/MS analysis
Liver
IC50 = 320.0 nM
Reference
Title : Minimizing CYP2C9 Inhibition of Exposed-Pyridine NAMPT (Nicotinamide Phosphoribosyltransferase) Inhibitors. Authors : Zak M, Yuen PW, Liu X, Patel S, Sampath D, Oeh J, Liederer BM, Wang W, O'Brien T, Xiao Y, Skelton N, Hua R, Sodhi J, Wang Y, Zhang L, Zhao G, Zheng X, Ho YC, Bair KW, Dragovich PS. Publication Year :
2016 Journal : J Med Chem Abstract : NAMPT inhibitors may show potential as therapeutics for oncology. Throughout our NAMPT inhibitor program, we found that exposed pyridines or related heterocyclic systems in the left-hand portion of the inhibitors are necessary pharmacophores for potent cellular NAMPT inhibition. However, when combined with a benzyl group in the center of the inhibitors, such pyridine-like moieties also led to consistent and potent inhibition of CYP2C9. In an attempt to reduce CYP2C9 inhibition, a parallel synthesis approach was used to identify central benzyl group replacements with increased Fsp3. A spirocyclic central motif was thus discovered that was combined with left-hand pyridines (or pyridine-like systems) to provide cellularly potent NAMPT inhibitors with minimal CYP2C9 inhibition. Further optimization of potency and ADME properties led to the discovery of compound 68, a highly potent NAMPT inhibitor with outstanding efficacy in a mouse tumor xenograft model and lacking measurable CYP2C9 inhibition at the concentrations tested. DOI : 10.1021/acs.jmedchem.6b00697
ADL-5747
Inhibition of human CYP3A4 at 10 uM
—
Inhibition = 18.0 %
Reference
Title : Spirocyclic delta opioid receptor agonists for the treatment of pain: discovery of N,N-diethyl-3-hydroxy-4-(spiro[chromene-2,4'-piperidine]-4-yl) benzamide (ADL5747). Authors : Le Bourdonnec B, Windh RT, Leister LK, Zhou QJ, Ajello CW, Gu M, Chu GH, Tuthill PA, Barker WM, Koblish M, Wiant DD, Graczyk TM, Belanger S, Cassel JA, Feschenko MS, Brogdon BL, Smith SA, Derelanko MJ, Kutz S, Little PJ, DeHaven RN, DeHaven-Hudkins DL, Dolle RE. Publication Year :
2009 Journal : J. Med. Chem. Abstract : Selective, nonpeptidic delta opioid receptor agonists have been the subject of great interest as potential novel analgesic agents. The discoveries of BW373U86 (1) and SNC80 (2) contributed to the rapid expansion of research in this field. However, poor drug-like properties and low therapeutic indices have prevented clinical evaluation of these agents. Doses of 1 and 2 similar to those required for analgesic activity produce convulsions in rodents and nonhuman primates. Recently, we described a novel series of potent, selective, and orally bioavailable delta opioid receptor agonists. The lead derivative, ADL5859 (4), is currently in phase II proof-of-concept studies for the management of pain. Further structure activity relationship exploration has led to the discovery of ADL5747 (36), which is approximately 50-fold more potent than 4 in an animal model of inflammatory pain. On the basis of its favorable efficacy, safety, and pharmacokinetic profile, 36 was selected as a clinical candidate for the treatment of pain. DOI : 10.1021/jm900773n
FIBOFLAPON
Inhibition of human CYP3A4
—
IC50 > 50.0 nM
Reference
Title : 5-Lipoxygenase-activating protein (FLAP) inhibitors. Part 4: development of 3-[3-tert-butylsulfanyl-1-[4-(6-ethoxypyridin-3-yl)benzyl]-5-(5-methylpyridin-2-ylmethoxy)-1H-indol-2-yl]-2,2-dimethylpropionic acid (AM803), a potent, oral, once daily FLAP inhibitor. Authors : Stock NS, Bain G, Zunic J, Li Y, Ziff J, Roppe J, Santini A, Darlington J, Prodanovich P, King CD, Baccei C, Lee C, Rong H, Chapman C, Broadhead A, Lorrain D, Correa L, Hutchinson JH, Evans JF, Prasit P. Publication Year :
2011 Journal : J. Med. Chem. Abstract : The potent 5-lipoxygenase-activating protein (FLAP) inhibitor 3-[3-tert-butylsulfanyl-1-[4-(6-ethoxypyridin-3-yl)benzyl]-5-(5-methylpyridin-2-ylmethoxy)-1H-indol-2-yl]-2,2-dimethylpropionic acid 11cc is described (AM803, now GSK2190915). Building upon AM103 (1) (Hutchinson et al. J. Med Chem.2009, 52, 5803-5815; Stock et al. Bioorg. Med. Chem. Lett. 2010, 20, 213-217; Stock et al. Bioorg. Med. Chem. Lett.2010, 20, 4598-4601), SAR studies centering around the pyridine moiety led to the discovery of compounds that exhibit significantly increased potency in a human whole blood assay measuring LTB(4) inhibition with longer drug preincubation times (15 min vs 5 h). Further studies identified 11cc with a potency of 2.9 nM in FLAP binding, an IC(50) of 76 nM for inhibition of LTB(4) in human blood (5 h incubation) and excellent preclinical toxicology and pharmacokinetics in rat and dog. 11cc also demonstrated an extended pharmacodynamic effect in a rodent bronchoalveolar lavage (BAL) model. This compound has successfully completed phase 1 clinical studies in healthy volunteers and is currently undergoing phase 2 trials in asthmatic patients. DOI : 10.1021/jm2008369
DAPORINAD
Inhibition of CYP3A4 in human liver microsomes using midazolam as substrate in presence of NADPH by LC-MS/MS analysis
Liver
IC50 = 540.0 nM
Reference
Title : Minimizing CYP2C9 Inhibition of Exposed-Pyridine NAMPT (Nicotinamide Phosphoribosyltransferase) Inhibitors. Authors : Zak M, Yuen PW, Liu X, Patel S, Sampath D, Oeh J, Liederer BM, Wang W, O'Brien T, Xiao Y, Skelton N, Hua R, Sodhi J, Wang Y, Zhang L, Zhao G, Zheng X, Ho YC, Bair KW, Dragovich PS. Publication Year :
2016 Journal : J Med Chem Abstract : NAMPT inhibitors may show potential as therapeutics for oncology. Throughout our NAMPT inhibitor program, we found that exposed pyridines or related heterocyclic systems in the left-hand portion of the inhibitors are necessary pharmacophores for potent cellular NAMPT inhibition. However, when combined with a benzyl group in the center of the inhibitors, such pyridine-like moieties also led to consistent and potent inhibition of CYP2C9. In an attempt to reduce CYP2C9 inhibition, a parallel synthesis approach was used to identify central benzyl group replacements with increased Fsp3. A spirocyclic central motif was thus discovered that was combined with left-hand pyridines (or pyridine-like systems) to provide cellularly potent NAMPT inhibitors with minimal CYP2C9 inhibition. Further optimization of potency and ADME properties led to the discovery of compound 68, a highly potent NAMPT inhibitor with outstanding efficacy in a mouse tumor xenograft model and lacking measurable CYP2C9 inhibition at the concentrations tested. DOI : 10.1021/acs.jmedchem.6b00697
ASTEMIZOLE
Inhibition of CYP3A4-mediated midazolam metabolite formation in human liver microsomes at 0.3 uM after 8 mins by LC-MS/MS analysis relative to control
Liver
Inhibition = 20.3 %
Reference
Title : Identifying a selective substrate and inhibitor pair for the evaluation of CYP2J2 activity. Authors : Lee CA, Jones JP, Katayama J, Kaspera R, Jiang Y, Freiwald S, Smith E, Walker GS, Totah RA. Publication Year :
2012 Journal : Drug Metab. Dispos. Abstract : CYP2J2, an arachidonic acid epoxygenase, is recognized for its role in the first-pass metabolism of astemizole and ebastine. To fully assess the role of CYP2J2 in drug metabolism, a selective substrate and potent specific chemical inhibitor are essential. In this study, we report amiodarone 4-hydoxylation as a specific CYP2J2-catalyzed reaction with no CYP3A4, or other drug-metabolizing enzyme, involvement. Amiodarone 4-hydroxylation enabled the determination of liver relative activity factor and intersystem extrapolation factor for CYP2J2. Amiodarone 4-hydroxylation correlated with astemizole O-demethylation but not with CYP2J2 protein content in a sample of human liver microsomes. To identify a specific CYP2J2 inhibitor, 138 drugs were screened using terfenadine and astemizole as probe substrates with recombinant CYP2J2. Forty-two drugs inhibited CYP2J2 activity by ≥50% at 30 μM, but inhibition was substrate-dependent. Of these, danazol was a potent inhibitor of both hydroxylation of terfenadine (IC(50) = 77 nM) and O-demethylation of astemizole (K(i) = 20 nM), and inhibition was mostly competitive. Danazol inhibited CYP2C9, CYP2C8, and CYP2D6 with IC(50) values of 1.44, 1.95, and 2.74 μM, respectively. Amiodarone or astemizole were included in a seven-probe cocktail for cytochrome P450 (P450) drug-interaction screening potential, and astemizole demonstrated a better profile because it did not appreciably interact with other P450 probes. Thus, danazol, amiodarone, and astemizole will facilitate the ability to determine the metabolic role of CYP2J2 in hepatic and extrahepatic tissues. DOI : 10.1124/dmd.111.043505
TACRINE
Time dependent inhibition of CYP3A4 (unknown origin) at 100 uM by LC/MS system
—
Inhibition < 10.0 %
Reference
Title : Combination of GSH trapping and time-dependent inhibition assays as a predictive method of drugs generating highly reactive metabolites. Authors : Nakayama S, Takakusa H, Watanabe A, Miyaji Y, Suzuki W, Sugiyama D, Shiosakai K, Honda K, Okudaira N, Izumi T, Okazaki O. Publication Year :
2011 Journal : Drug Metab. Dispos. Abstract : Covalent binding (CB) of reactive metabolites (RMs) is potentially involved in severe adverse drug reactions. Because the CB assay is of low throughput and costly, a qualitative trapping assay using agents such as [(35)S]GSH is often performed in the early stages of drug discovery. However, trapping methods alone cannot replace the CB assay. We hypothesized that the time-dependent inhibition (TDI) assay might be complementary to the [(35)S]GSH trapping assay in detecting RMs. We performed CB assays, [(35)S]GSH trapping assays, and TDI assays for 42 structurally diverse compounds. First, we showed that the [(35)S]GSH trapping assay alone does not correlate with the extent of CB. Four compounds that the [(35)S]GSH trapping assay failed to detect but that showed high extent of CB were inactivators of the enzyme in the TDI assay. There was a tendency for compounds judged as positive in the TDI assay to show a high degree of CB irrespective of the result of the [(35)S]GSH trapping assay. Finally, to combine parameters from the two assays, we introduced intrinsic clearance to describe the formation of RMs (CL(int, RMs)). The Spearman rank correlation coefficient between the extent of CB and CL(int, RMs) was 0.77 (p < 0.0001), which was better than that for the formation rates of [(35)S]GSH adducts. Therefore, we demonstrated that a combination of the [(35)S]GSH trapping and TDI assays is an effective method for detecting compounds potentially capable of generating highly reactive metabolites in the early stages of drug discovery. DOI : 10.1124/dmd.111.039180
TACRINE
Inhibition of recombinant human CYP3A4 expressed in insect cells at 100 uM using DBOMF as substrate pretreated for 10 mins followed by substrate addition measured every minute for 15 mins in presence of NADP+ by fluorescence assay relative to control
—
Inhibition = 55.0 %
Reference
Title : Cytochrome P450 binding studies of novel tacrine derivatives: Predicting the risk of hepatotoxicity. Authors : McEneny-King A, Osman W, Edginton AN, Rao PPN. Publication Year :
2017 Journal : Bioorg Med Chem Lett Abstract : The 1,2,3,4-tetrahydroacridine derivative tacrine was the first drug approved to treat Alzheimer's disease (AD). It is known to act as a potent cholinesterase inhibitor. However, tacrine was removed from the market due to its hepatotoxicity concerns as it undergoes metabolism to toxic quinonemethide species through the cytochrome P450 enzyme CYP1A2. Despite these challenges, tacrine serves as a useful template in the development of novel multi-targeting anti-AD agents. In this regard, we sought to evaluate the risk of hepatotoxicity in a series of C9 substituted tacrine derivatives that exhibit cholinesterase inhibition properties. The hepatotoxic potential of tacrine derivatives was evaluated using recombinant cytochrome (CYP) P450 CYP1A2 and CYP3A4 enzymes. Molecular docking studies were conducted to predict their binding modes and potential risk of forming hepatotoxic metabolites. Tacrine derivatives compound 1 (N-(3,4-dimethoxybenzyl)-1,2,3,4-tetrahydroacridin-9-amine) and 2 (6-chloro-N-(3,4-dimethoxybenzyl)-1,2,3,4-tetrahydroacridin-9-amine) which possess a C9 3,4-dimethoxybenzylamino substituent exhibited weak binding to CYP1A2 enzyme (1, IC50=33.0µM; 2, IC50=8.5µM) compared to tacrine (CYP1A2 IC50=1.5µM). Modeling studies show that the presence of a bulky 3,4-dimethoxybenzylamino C9 substituent prevents the orientation of the 1,2,3,4-tetrahydroacridine ring close to the heme-iron center of CYP1A2 thereby reducing the risk of forming hepatotoxic species. DOI : 10.1016/j.bmcl.2017.04.006
TAK-715
Inhibitory activity against cytochrome P450 3A4
—
Inhibition = 4.3 %
Reference
Title : Novel inhibitor of p38 MAP kinase as an anti-TNF-alpha drug: discovery of N-[4-[2-ethyl-4-(3-methylphenyl)-1,3-thiazol-5-yl]-2-pyridyl]benzamide (TAK-715) as a potent and orally active anti-rheumatoid arthritis agent. Authors : Miwatashi S, Arikawa Y, Kotani E, Miyamoto M, Naruo K, Kimura H, Tanaka T, Asahi S, Ohkawa S. Publication Year :
2005 Journal : J. Med. Chem. Abstract : The p38 mitogen-activated protein (MAP) kinase has been implicated in the proinflammatory cytokine signal pathway, and its inhibitors are potentially useful for the treatment of chronic inflammatory diseases such as rheumatoid arthritis (RA) and inflammatory bowel disease. To develop a new drug for RA, we synthesized a novel series of 4-phenyl-5-pyridyl-1,3-thiazoles and evaluated their inhibition of p38 MAP kinase, lipopolysaccharide (LPS)-stimulated release of tumor necrosis factor-alpha (TNF-alpha) from human monocytic THP-1 cells in vitro, and LPS-induced TNF-alpha production in vivo in mice. During the course of the study, we found that these compounds risk the inhibition of cytochrome P450 (CYP) isoforms by coordination of the 4-pyridyl nitrogen with heme iron. We therefore investigated the effects of substitution at the 2-position of the pyridyl ring on the inhibitory activity of p38 MAP kinase and CYPs in more detail. As a result, N-[4-[2-ethyl-4-(3-methylphenyl)-1,3-thiazol-5-yl]-2-pyridyl]benzamide (8h, TAK-715) exhibited potent inhibitory activity in these assays (inhibition of p38alpha, IC50 = 7.1 nM; LPS-stimulated release of TNF-alpha from THP-1, IC50 = 48 nM; LPS-induced TNF-alpha production in mice, 87.6% inhibition at 10 mg/kg, po) and no inhibitory activity for major CYPs, including CYP3A4. This compound also showed good bioavailability in mice and rats and significant efficacy in a rat adjuvant-induced arthritis model. Compound 8h was selected as a clinical candidate and is now under clinical investigation for the treatment of RA. DOI : 10.1021/jm050165o
SUFUGOLIX
Inhibition of human CYP3A4 assessed as conversion of testosterone to 6beta-hydroxytestosterone at 10 uM after 30 mins by HPLC analysis
—
Inhibition = 36.0 %
Reference
Title : Discovery of 1-{4-[1-(2,6-difluorobenzyl)-5-[(dimethylamino)methyl]-3-(6-methoxypyridazin-3-yl)-2,4-dioxo-1,2,3,4-tetrahydrothieno[2,3-d]pyrimidin-6-yl]phenyl}-3-methoxyurea (TAK-385) as a potent, orally active, non-peptide antagonist of the human gonadotropin-releasing hormone receptor. Authors : Miwa K, Hitaka T, Imada T, Sasaki S, Yoshimatsu M, Kusaka M, Tanaka A, Nakata D, Furuya S, Endo S, Hamamura K, Kitazaki T. Publication Year :
2011 Journal : J. Med. Chem. Abstract : We previously discovered an orally active human gonadotropin-releasing hormone (GnRH) receptor antagonist, thieno[2,3-d]pyrimidine-2,4-dione derivative 1 (sufugolix). To reduce the cytochrome P450 (CYP) inhibitory activity and improve in vivo GnRH antagonistic activity, further optimization of this scaffold was carried out. We focused our synthetic efforts on chemical modification at the 5 and 3 positions of the thieno[2,3-d]pyrimidine-2,4-dione ring based on computational modeling, which resulted in the discovery of 1-{4-[1-(2,6-difluorobenzyl)-5-[(dimethylamino)methyl]-3-(6-methoxypyridazin-3-yl)-2,4-dioxo-1,2,3,4-tetrahydrothieno[2,3-d]pyrimidin-6-yl]phenyl}-3-methoxyurea (16b) as a highly potent and orally active GnRH antagonist. Compound 16b showed potent in vitro GnRH antagonistic activity in the presence of fetal bovine serum (FBS) without CYP inhibition. Oral administration of 16b maintained the suppressive effect of the plasma luteinizing hormone levels in castrated cynomolgus monkeys at a 3 mg/kg dose for more than 24 h. Compound 16b is currently under clinical development with the code name of TAK-385. DOI : 10.1021/jm200216q
BERGAPTEN
Inhibition of recombinant human CYP3A4 expressed in baker's yeast-derived microsomes (Sacchrosomes) at 10 uM using dibenzylfluorescein substrate by EROD assay relative to control
—
Inhibition = 3.0 %
Reference
Title : Identification of karanjin isolated from the Indian beech tree as a potent CYP1 enzyme inhibitor with cellular efficacy <i>via</i> screening of a natural product repository. Authors : Joshi P, Sonawane VR, Williams IS, McCann GJP, Gatchie L, Sharma R, Satti N, Chaudhuri B, Bharate SB. Publication Year :
2018 Journal : MedChemComm Abstract : CYP1A1 is thought to mediate carcinogenesis in oral, lung and epithelial cancers. In order to identify a CYP1A1 inhibitor from an edible plant, 394 natural products in the IIIM's natural product repository were screened, at 10 μM concentration, using CYP1A1-Sacchrosomes™ (<i>i.e.</i> microsomal enzyme isolated from recombinant baker's yeast). Twenty-seven natural products were identified that inhibited 40-97% of CYP1A1's 7-ethoxyresorufin-<i>O</i>-deethylase activity. The IC<sub>50</sub> values of the 'hits', belonging to different chemical scaffolds, were determined. Their selectivity was studied against a panel of 8 CYP-Sacchrosomes™. In order to assess cellular efficacy, the 'hits' were screened for their capability to inhibit CYP enzymes expressed within live recombinant human embryonic kidney (HEK293) cells from plasmids encoding specific CYP genes (1A2, 1B1, 2C9, 2C19, 2D6, 3A4). Isopimpinellin (IN-475; IC<sub>50</sub>, 20 nM) and karanjin (IN-195; IC<sub>50</sub>, 30 nM) showed the most potent inhibition of CYP1A1 in human cells. Isopimpinellin is found in celery, parsnip, fruits and in the rind and pulp of limes whereas different parts of the Indian beech tree, which contain karanjin, have been used in traditional medicine. Both isopimpinellin and karanjin negate the cellular toxicity of CYP1A1-mediated benzo[<i>a</i>]pyrene. Molecular docking and molecular dynamic simulations with CYP isoforms rationalize the observed trends in the potency and selectivity of isopimpinellin and karanjin. DOI : 10.1039/C7MD00388A
TICRYNAFEN
Time dependent inhibition of CYP3A4 (unknown origin) at 100 uM by LC/MS system
—
Inhibition < 10.0 %
Reference
Title : Combination of GSH trapping and time-dependent inhibition assays as a predictive method of drugs generating highly reactive metabolites. Authors : Nakayama S, Takakusa H, Watanabe A, Miyaji Y, Suzuki W, Sugiyama D, Shiosakai K, Honda K, Okudaira N, Izumi T, Okazaki O. Publication Year :
2011 Journal : Drug Metab. Dispos. Abstract : Covalent binding (CB) of reactive metabolites (RMs) is potentially involved in severe adverse drug reactions. Because the CB assay is of low throughput and costly, a qualitative trapping assay using agents such as [(35)S]GSH is often performed in the early stages of drug discovery. However, trapping methods alone cannot replace the CB assay. We hypothesized that the time-dependent inhibition (TDI) assay might be complementary to the [(35)S]GSH trapping assay in detecting RMs. We performed CB assays, [(35)S]GSH trapping assays, and TDI assays for 42 structurally diverse compounds. First, we showed that the [(35)S]GSH trapping assay alone does not correlate with the extent of CB. Four compounds that the [(35)S]GSH trapping assay failed to detect but that showed high extent of CB were inactivators of the enzyme in the TDI assay. There was a tendency for compounds judged as positive in the TDI assay to show a high degree of CB irrespective of the result of the [(35)S]GSH trapping assay. Finally, to combine parameters from the two assays, we introduced intrinsic clearance to describe the formation of RMs (CL(int, RMs)). The Spearman rank correlation coefficient between the extent of CB and CL(int, RMs) was 0.77 (p < 0.0001), which was better than that for the formation rates of [(35)S]GSH adducts. Therefore, we demonstrated that a combination of the [(35)S]GSH trapping and TDI assays is an effective method for detecting compounds potentially capable of generating highly reactive metabolites in the early stages of drug discovery. DOI : 10.1124/dmd.111.039180
MAVELERTINIB
Inhibition of human CYP3A4 at 10 uM
—
Inhibition < 30.0 %
Reference
Title : Discovery of N-((3R,4R)-4-Fluoro-1-(6-((3-methoxy-1-methyl-1H-pyrazol-4-yl)amino)-9-methyl-9H-purin-2-yl)pyrrolidine-3-yl)acrylamide (PF-06747775) through Structure-Based Drug Design: A High Affinity Irreversible Inhibitor Targeting Oncogenic EGFR Mutants with Selectivity over Wild-Type EGFR. Authors : Planken S, Behenna DC, Nair SK, Johnson TO, Nagata A, Almaden C, Bailey S, Ballard TE, Bernier L, Cheng H, Cho-Schultz S, Dalvie D, Deal JG, Dinh DM, Edwards MP, Ferre RA, Gajiwala KS, Hemkens M, Kania RS, Kath JC, Matthews J, Murray BW, Niessen S, Orr ST, Pairish M, Sach NW, Shen H, Shi M, Solowiej J, Tran K, Tseng E, Vicini P, Wang Y, Weinrich SL, Zhou R, Zientek M, Liu L, Luo Y, Xin S, Zhang C, Lafontaine J. Publication Year :
2017 Journal : J Med Chem Abstract : Mutant epidermal growth factor receptor (EGFR) is a major driver of non-small-cell lung cancer (NSCLC). Marketed first generation inhibitors, such as erlotinib, effect a transient beneficial response in EGFR mutant NSCLC patients before resistance mechanisms render these inhibitors ineffective. Secondary oncogenic EGFR mutations account for approximately 50% of relapses, the most common being the gatekeeper T790M substitution that renders existing therapies ineffective. The discovery of PF-06459988 (1), an irreversible pyrrolopyrimidine inhibitor of EGFR T790M mutants, was recently disclosed.1 Herein, we describe our continued efforts to achieve potency across EGFR oncogenic mutations and improved kinome selectivity, resulting in the discovery of clinical candidate PF-06747775 (21), which provides potent EGFR activity against the four common mutants (exon 19 deletion (Del), L858R, and double mutants T790M/L858R and T790M/Del), selectivity over wild-type EGFR, and desirable ADME properties. Compound 21 is currently being evaluated in phase-I clinical trials of mutant EGFR driven NSCLC. DOI : 10.1021/acs.jmedchem.6b01894
LY-2886721
Inhibition of CYP3A4 in human liver microsomes at 10 uM using midazolam as substrate preincubated for 10 mins followed by substrate and NADPH addition measured after 3 mins by LC-MS/MS analysis
Liver
Inhibition = 0.0 %
Reference
Title : Authors : Publication Year : Journal :
ETACONAZOLE
Inhibition of human recombinant CYP3A4 expressed in Saccharomyces cerevisiae by fluorescence assay
—
IC50 = 210.0 nM
Reference
Title : Characterization of type II ligands in CYP2C9 and CYP3A4. Authors : Ahlström MM, Zamora I. Publication Year :
2008 Journal : J. Med. Chem. Abstract : Type II cytochrome P450 (CYP) ligands cause inhibition by direct coordination to the heme iron atom. This interaction usually leads to high inhibitory potential, which can cause drug-drug interaction. The approach to design compounds with diminished CYP inhibition is different depending on whether the compound binds (type II ligand) or not (type I ligand) to the iron atom of the heme group. In this study, the structural characteristics of nitrogen-containing compounds, which bind to the iron atom in two CYP isoforms (CYP2C9 and CYP3A4), were investigated. The in vitro assays applied were fluorescence inhibition assay, difference spectra measurements, and K s determination. Computational modeling as an alternative method to difference spectra measurements to distinguish between type I and type II ligands was also explored. Since two CYP isoforms were used, information about the isoform specificity of type II ligands was also analyzed. The in silico method developed in this study applied together with the information gained from the experimental measurements may result in better decisions during the drug discovery process. DOI : 10.1021/jm701121y
ETACONAZOLE
Binding affinity to purified human recombinant CYP3A4 expressed in Escherichia coli assessed as spectral dissociation constant
—
Kd = 210.0 nM
Reference
Title : Characterization of type II ligands in CYP2C9 and CYP3A4. Authors : Ahlström MM, Zamora I. Publication Year :
2008 Journal : J. Med. Chem. Abstract : Type II cytochrome P450 (CYP) ligands cause inhibition by direct coordination to the heme iron atom. This interaction usually leads to high inhibitory potential, which can cause drug-drug interaction. The approach to design compounds with diminished CYP inhibition is different depending on whether the compound binds (type II ligand) or not (type I ligand) to the iron atom of the heme group. In this study, the structural characteristics of nitrogen-containing compounds, which bind to the iron atom in two CYP isoforms (CYP2C9 and CYP3A4), were investigated. The in vitro assays applied were fluorescence inhibition assay, difference spectra measurements, and K s determination. Computational modeling as an alternative method to difference spectra measurements to distinguish between type I and type II ligands was also explored. Since two CYP isoforms were used, information about the isoform specificity of type II ligands was also analyzed. The in silico method developed in this study applied together with the information gained from the experimental measurements may result in better decisions during the drug discovery process. DOI : 10.1021/jm701121y
AZD-3759
Inhibition of CYP3A4 (unknown origin)
—
IC50 > 50.0 nM
Reference
Title : Discovery and Evaluation of Clinical Candidate AZD3759, a Potent, Oral Active, Central Nervous System-Penetrant, Epidermal Growth Factor Receptor Tyrosine Kinase Inhibitor. Authors : Zeng Q, Wang J, Cheng Z, Chen K, Johnström P, Varnäs K, Li DY, Yang ZF, Zhang X. Publication Year :
2015 Journal : J. Med. Chem. Abstract : Recent reports suggest that an increasing number of patients with lung cancer, especially those with activating mutations of the epidermal growth factor receptor (EGFR), also present with brain metastases and leptomeningeal metastases. These patients have poor prognosis as there are no approved drugs for these indications. Available agents have poor efficacy for these patients even at well above their standard dose. Herein, we report the discovery of (4-[(3-chloro-2-fluorophenyl)amino]-7-methoxyquinazolin-6-yl (2R)-2,4-dimethylpiperazine-1-carboxylate 1m (AZD3759), an investigational drug currently in Phase 1 clinical trial, which has excellent central nervous system penetration and which induces profound regression of brain metastases in a mouse model. DOI : 10.1021/acs.jmedchem.5b01073
GANAPLACIDE
Inhibition of CYP3A4 (unknown origin)
—
Ki = 156.0 nM
Reference
Title : The Development Process for Discovery and Clinical Advancement of Modern Antimalarials. Authors : Ashton TD, Devine SM, Möhrle JJ, Laleu B, Burrows JN, Charman SA, Creek DJ, Sleebs BE. Publication Year :
2019 Journal : J Med Chem Abstract : Malaria is a devastating disease caused by <i>Plasmodium</i> parasites, resulting in approximately 435000 deaths in 2018. The impact of malaria is compounded by the emergence of widespread resistance to current antimalarial therapies. Recently, a new strategy was initiated to screen small molecule collections against the <i>Plasmodium</i> parasite enabling the identification of new antimalarial chemotypes with novel modes of action. This initiative ushered in the modern era of antimalarial drug development, and as a result, numerous lead candidates are advancing toward or are currently in human clinical trials. In this Perspective, we describe the development pathway of four of the most clinically advanced modern antimalarials, KAE609, KAF156, DSM265, and MMV048. Additionally, the mechanism of action and life-cycle stage specificity of the four antimalarials is discussed in relation to aligning with global strategies to treat and eliminate malaria. This perspective serves as a guide to the expectations of modern antimalarial drug development. DOI : 10.1021/acs.jmedchem.9b00761
MK-8033
Inhibition of CYP3A4 in human liver microsomes using testosterone as substrate at 10 uM by LC-MS-MS analysis relative to control
Liver
Inhibition = 31.0 %
Reference
Title : Discovery of 1-[3-(1-methyl-1H-pyrazol-4-yl)-5-oxo-5H-benzo[4,5]cyclohepta[1,2-b]pyridin-7-yl]-N-(pyridin-2-ylmethyl)methanesulfonamide (MK-8033): A Specific c-Met/Ron dual kinase inhibitor with preferential affinity for the activated state of c-Met. Authors : Northrup AB, Katcher MH, Altman MD, Chenard M, Daniels MH, Deshmukh SV, Falcone D, Guerin DJ, Hatch H, Li C, Lu W, Lutterbach B, Allison TJ, Patel SB, Reilly JF, Reutershan M, Rickert KW, Rosenstein C, Soisson SM, Szewczak AA, Walker D, Wilson K, Young JR, Pan BS, Dinsmore CJ. Publication Year :
2013 Journal : J. Med. Chem. Abstract : This report documents the first example of a specific inhibitor of protein kinases with preferential binding to the activated kinase conformation: 5H-benzo[4,5]cyclohepta[1,2-b]pyridin-5-one 11r (MK-8033), a dual c-Met/Ron inhibitor under investigation as a treatment for cancer. The design of 11r was based on the desire to reduce time-dependent inhibition of CYP3A4 (TDI) by members of this structural class. A novel two-step protocol for the synthesis of benzylic sulfonamides was developed to access 11r and analogues. We provide a rationale for the observed selectivity based on X-ray crystallographic evidence and discuss selectivity trends with additional examples. Importantly, 11r provides full inhibition of tumor growth in a c-Met amplified (GTL-16) subcutaneous tumor xenograft model and may have an advantage over inactive form kinase inhibitors due to equal potency against a panel of oncogenic activating mutations of c-Met in contrast to c-Met inhibitors without preferential binding to the active kinase conformation. DOI : 10.1021/jm301619u
VACTOSERTIB
Inhibition of CYP3A4 in human liver microsomes using midazolam as substrate at 10 uM incubated for 5 mins prior to substrate addition measured after 10 mins by LC/MS/MS analysis
Liver
Inhibition = 44.8 %
Reference
Title : Discovery of N-((4-([1,2,4]triazolo[1,5-a]pyridin-6-yl)-5-(6-methylpyridin-2-yl)-1H-imidazol-2-yl)methyl)-2-fluoroaniline (EW-7197): a highly potent, selective, and orally bioavailable inhibitor of TGF-β type I receptor kinase as cancer immunotherapeutic/antifibrotic agent. Authors : Jin CH, Krishnaiah M, Sreenu D, Subrahmanyam VB, Rao KS, Lee HJ, Park SJ, Park HJ, Lee K, Sheen YY, Kim DK. Publication Year :
2014 Journal : J. Med. Chem. Abstract : A series of 2-substituted-4-([1,2,4]triazolo[1,5-a]pyridin-6-yl)-5-(6-methylpyridin-2-yl)imidazoles was synthesized and evaluated to optimize a prototype inhibitor of TGF-β type I receptor kinase (ALK5), 6. Combination of replacement of a quinoxalin-6-yl moiety of 6 with a [1,2,4]triazolo[1,5-a]pyridin-6-yl moiety, insertion of a methyleneamino linker, and a o-F substituent in the phenyl ring markedly increased ALK5 inhibitory activity, kinase selectivity, and oral bioavailability. The 12b (EW-7197) inhibited ALK5 with IC50 value of 0.013 μM in a kinase assay and with IC50 values of 0.0165 and 0.0121 μM in HaCaT (3TP-luc) stable cells and 4T1 (3TP-luc) stable cells, respectively, in a luciferase assay. Selectivity profiling of 12b using a panel of 320 protein kinases revealed that it is a highly selective ALK5/ALK4 inhibitor. Pharmacokinetic study with 12b·HCl in rats showed an oral bioavailability of 51% with high systemic exposure (AUC) of 1426 ng × h/mL and maximum plasma concentration (Cmax) of 1620 ng/mL. Rational optimization of 6 has led to the identification of a highly potent, selective, and orally bioavailable ALK5 inhibitor 12b. DOI : 10.1021/jm500115w
INDOMETHACIN
Time dependent inhibition of CYP3A4 (unknown origin) at 100 uM by LC/MS system
—
Inhibition < 10.0 %
Reference
Title : Combination of GSH trapping and time-dependent inhibition assays as a predictive method of drugs generating highly reactive metabolites. Authors : Nakayama S, Takakusa H, Watanabe A, Miyaji Y, Suzuki W, Sugiyama D, Shiosakai K, Honda K, Okudaira N, Izumi T, Okazaki O. Publication Year :
2011 Journal : Drug Metab. Dispos. Abstract : Covalent binding (CB) of reactive metabolites (RMs) is potentially involved in severe adverse drug reactions. Because the CB assay is of low throughput and costly, a qualitative trapping assay using agents such as [(35)S]GSH is often performed in the early stages of drug discovery. However, trapping methods alone cannot replace the CB assay. We hypothesized that the time-dependent inhibition (TDI) assay might be complementary to the [(35)S]GSH trapping assay in detecting RMs. We performed CB assays, [(35)S]GSH trapping assays, and TDI assays for 42 structurally diverse compounds. First, we showed that the [(35)S]GSH trapping assay alone does not correlate with the extent of CB. Four compounds that the [(35)S]GSH trapping assay failed to detect but that showed high extent of CB were inactivators of the enzyme in the TDI assay. There was a tendency for compounds judged as positive in the TDI assay to show a high degree of CB irrespective of the result of the [(35)S]GSH trapping assay. Finally, to combine parameters from the two assays, we introduced intrinsic clearance to describe the formation of RMs (CL(int, RMs)). The Spearman rank correlation coefficient between the extent of CB and CL(int, RMs) was 0.77 (p < 0.0001), which was better than that for the formation rates of [(35)S]GSH adducts. Therefore, we demonstrated that a combination of the [(35)S]GSH trapping and TDI assays is an effective method for detecting compounds potentially capable of generating highly reactive metabolites in the early stages of drug discovery. DOI : 10.1124/dmd.111.039180
PLUMBAGIN
Inhibition of recombinant human CYP3A4 expressed in baker's yeast-derived microsomes (Sacchrosomes) at 10 uM using dibenzylfluorescein substrate by EROD assay relative to control
—
Inhibition = 15.0 %
Reference
Title : Identification of karanjin isolated from the Indian beech tree as a potent CYP1 enzyme inhibitor with cellular efficacy <i>via</i> screening of a natural product repository. Authors : Joshi P, Sonawane VR, Williams IS, McCann GJP, Gatchie L, Sharma R, Satti N, Chaudhuri B, Bharate SB. Publication Year :
2018 Journal : MedChemComm Abstract : CYP1A1 is thought to mediate carcinogenesis in oral, lung and epithelial cancers. In order to identify a CYP1A1 inhibitor from an edible plant, 394 natural products in the IIIM's natural product repository were screened, at 10 μM concentration, using CYP1A1-Sacchrosomes™ (<i>i.e.</i> microsomal enzyme isolated from recombinant baker's yeast). Twenty-seven natural products were identified that inhibited 40-97% of CYP1A1's 7-ethoxyresorufin-<i>O</i>-deethylase activity. The IC<sub>50</sub> values of the 'hits', belonging to different chemical scaffolds, were determined. Their selectivity was studied against a panel of 8 CYP-Sacchrosomes™. In order to assess cellular efficacy, the 'hits' were screened for their capability to inhibit CYP enzymes expressed within live recombinant human embryonic kidney (HEK293) cells from plasmids encoding specific CYP genes (1A2, 1B1, 2C9, 2C19, 2D6, 3A4). Isopimpinellin (IN-475; IC<sub>50</sub>, 20 nM) and karanjin (IN-195; IC<sub>50</sub>, 30 nM) showed the most potent inhibition of CYP1A1 in human cells. Isopimpinellin is found in celery, parsnip, fruits and in the rind and pulp of limes whereas different parts of the Indian beech tree, which contain karanjin, have been used in traditional medicine. Both isopimpinellin and karanjin negate the cellular toxicity of CYP1A1-mediated benzo[<i>a</i>]pyrene. Molecular docking and molecular dynamic simulations with CYP isoforms rationalize the observed trends in the potency and selectivity of isopimpinellin and karanjin. DOI : 10.1039/C7MD00388A
MAVORIXAFOR
Inhibition of human CYP3A4 at 1 uM
—
Inhibition = 35.0 %
Reference
Title : Discovery of novel N-aryl piperazine CXCR4 antagonists. Authors : Zhao H, Prosser AR, Liotta DC, Wilson LJ. Publication Year :
2015 Journal : Bioorg. Med. Chem. Lett. Abstract : A novel series of CXCR4 antagonists with substituted piperazines as benzimidazole replacements is described. These compounds showed micromolar to nanomolar potency in CXCR4-mediated functional and HIV assays, namely inhibition of X4 HIV-1(IIIB) virus in MAGI-CCR5/CXCR4 cells and inhibition of SDF-1 induced calcium release in Chem-1 cells. Preliminary SAR investigations led to the identification of a series of N-aryl piperazines as the most potent compounds. Results show SAR that indicates type and position of the aromatic ring, as well as type of linker and stereochemistry are significant for activity. Profiling of several lead compounds showed that one (49b) reduced susceptibility towards CYP450 and hERG, and the best overall profile when considering both SDF-1 and HIV potencies (6-20 nM). DOI : 10.1016/j.bmcl.2015.04.036
LORATADINE
Inhibition of CYP3A4 in human microsomes at 25 uM
—
Inhibition = 30.0 %
Reference
Title : Synthesis and structure-activity relationships of novel histamine H1 antagonists: indolylpiperidinyl benzoic acid derivatives. Authors : Fonquerna S, Miralpeix M, Pagès L, Puig C, Cardús A, Antón F, Cárdenas A, Vilella D, Aparici M, Calaf E, Prieto J, Gras J, Huerta JM, Warrellow G, Beleta J, Ryder H. Publication Year :
2004 Journal : J. Med. Chem. Abstract : A series of indolylpiperidinyl derivatives were prepared and evaluated for their activity as histamine H(1) antagonists. Structure-activity relationship studies were directed toward improving in vivo activity and pharmacokinetic profile of our first lead (1). Substitution of fluorine in position 6 on the indolyl ring led to higher in vivo activity in the inhibition of histamine-induced cutaneous vascular permeability assay but lower selectivity toward 5HT(2) receptor. Extensive optimization was carried out within this series and a number of histamine H(1) antagonists showing potency and long duration of action in vivo and low brain penetration or cardiotoxic potential were identified. Within this novel series, indolylpiperidines 15, 20, 48,51 and 52 exhibited a long half-life in rat and have been selected for further preclinical evaluation. DOI : 10.1021/jm0498203
EXEMESTANE
Drug metabolism assessed as human recombinant CYP3A4-mediated 6-Hydroxymethylexemestane formation assessed as inhibition of metabolite formation at 10 uM after 10 mins by Michaelis-Menten saturation curve analysis in presence of 50 uM troleandomycin
—
Inhibition = 95.0 %
Reference
Title : In vitro cytochrome P450-mediated metabolism of exemestane. Authors : Kamdem LK, Flockhart DA, Desta Z. Publication Year :
2011 Journal : Drug Metab. Dispos. Abstract : Exemestane is a potent and irreversible steroidal aromatase inhibitor drug used for the treatment of estrogen receptor-positive breast cancer. Our aim was to identify and assess the contribution of the specific cytochromes P450 (P450s) responsible for exemestane primary in vitro metabolism. With the use of high-performance liquid chromatography and liquid chromatography-tandem mass spectrometry analytical techniques, 17-hydroexemestane (MI) formation and 6-hydroxymethylexemestane (MII) formation were found to be the predominant exemestane metabolic pathways. In a bank of 15 well characterized human liver microsomes with known P450 isoform-specific activities, the MI formation rate correlated significantly with CYP1A2 (Spearman r = 0.60, p = 0.02) and CYP4A11 (Spearman r = 0.67, p = 0.01) isoform-specific activities, whereas the MII production rate significantly correlated with CYP2B6 (Spearman r = 0.57, p = 0.03) and CYP3A (Spearman r = 0.76, p = 0.005) isoform-specific activities. Recombinant CYP1A1 metabolized exemestane to MI with a catalytic efficiency (Cl(int)) of 150 nl/pmol P450 × min that was at least 3.5-fold higher than those of other P450s investigated. Recombinant CYP3A4 catalyzed MII formation from exemestane with a catalytic efficiency of 840 nl/pmol P450 × min that was at least 4-fold higher than those of other P450s investigated. Among a panel of 10 chemical inhibitors tested, only ketoconazole and troleandomycin (CYP3A-specific chemical inhibitors) significantly inhibited the formation of MII by 45 and 95%, respectively. None of them markedly inhibited the formation of MI. In summary, exemestane seems to be metabolized to MI by multiple P450s that include CYP4A11 and CYP1A1/2, whereas its oxidation to MII is primarily mediated by CYP3A. DOI : 10.1124/dmd.110.032276
EXEMESTANE
Drug metabolism assessed as human recombinant CYP3A4-mediated 6-Hydroxymethylexemestane formation assessed as inhibition of metabolite formation at 10 uM after 10 mins by Michaelis-Menten saturation curve analysis in presence of 1 uM ketoconazole
—
Inhibition = 45.0 %
Reference
Title : In vitro cytochrome P450-mediated metabolism of exemestane. Authors : Kamdem LK, Flockhart DA, Desta Z. Publication Year :
2011 Journal : Drug Metab. Dispos. Abstract : Exemestane is a potent and irreversible steroidal aromatase inhibitor drug used for the treatment of estrogen receptor-positive breast cancer. Our aim was to identify and assess the contribution of the specific cytochromes P450 (P450s) responsible for exemestane primary in vitro metabolism. With the use of high-performance liquid chromatography and liquid chromatography-tandem mass spectrometry analytical techniques, 17-hydroexemestane (MI) formation and 6-hydroxymethylexemestane (MII) formation were found to be the predominant exemestane metabolic pathways. In a bank of 15 well characterized human liver microsomes with known P450 isoform-specific activities, the MI formation rate correlated significantly with CYP1A2 (Spearman r = 0.60, p = 0.02) and CYP4A11 (Spearman r = 0.67, p = 0.01) isoform-specific activities, whereas the MII production rate significantly correlated with CYP2B6 (Spearman r = 0.57, p = 0.03) and CYP3A (Spearman r = 0.76, p = 0.005) isoform-specific activities. Recombinant CYP1A1 metabolized exemestane to MI with a catalytic efficiency (Cl(int)) of 150 nl/pmol P450 × min that was at least 3.5-fold higher than those of other P450s investigated. Recombinant CYP3A4 catalyzed MII formation from exemestane with a catalytic efficiency of 840 nl/pmol P450 × min that was at least 4-fold higher than those of other P450s investigated. Among a panel of 10 chemical inhibitors tested, only ketoconazole and troleandomycin (CYP3A-specific chemical inhibitors) significantly inhibited the formation of MII by 45 and 95%, respectively. None of them markedly inhibited the formation of MI. In summary, exemestane seems to be metabolized to MI by multiple P450s that include CYP4A11 and CYP1A1/2, whereas its oxidation to MII is primarily mediated by CYP3A. DOI : 10.1124/dmd.110.032276
NEVIRAPINE
Time dependent inhibition of CYP3A4 (unknown origin) at 100 uM by LC/MS system
—
Inhibition < 10.0 %
Reference
Title : Combination of GSH trapping and time-dependent inhibition assays as a predictive method of drugs generating highly reactive metabolites. Authors : Nakayama S, Takakusa H, Watanabe A, Miyaji Y, Suzuki W, Sugiyama D, Shiosakai K, Honda K, Okudaira N, Izumi T, Okazaki O. Publication Year :
2011 Journal : Drug Metab. Dispos. Abstract : Covalent binding (CB) of reactive metabolites (RMs) is potentially involved in severe adverse drug reactions. Because the CB assay is of low throughput and costly, a qualitative trapping assay using agents such as [(35)S]GSH is often performed in the early stages of drug discovery. However, trapping methods alone cannot replace the CB assay. We hypothesized that the time-dependent inhibition (TDI) assay might be complementary to the [(35)S]GSH trapping assay in detecting RMs. We performed CB assays, [(35)S]GSH trapping assays, and TDI assays for 42 structurally diverse compounds. First, we showed that the [(35)S]GSH trapping assay alone does not correlate with the extent of CB. Four compounds that the [(35)S]GSH trapping assay failed to detect but that showed high extent of CB were inactivators of the enzyme in the TDI assay. There was a tendency for compounds judged as positive in the TDI assay to show a high degree of CB irrespective of the result of the [(35)S]GSH trapping assay. Finally, to combine parameters from the two assays, we introduced intrinsic clearance to describe the formation of RMs (CL(int, RMs)). The Spearman rank correlation coefficient between the extent of CB and CL(int, RMs) was 0.77 (p < 0.0001), which was better than that for the formation rates of [(35)S]GSH adducts. Therefore, we demonstrated that a combination of the [(35)S]GSH trapping and TDI assays is an effective method for detecting compounds potentially capable of generating highly reactive metabolites in the early stages of drug discovery. DOI : 10.1124/dmd.111.039180
SUVECALTAMIDE
Inhibition of recombinant human CYP3A4 in human liver microsomes at 1 uM relative to control
Liver
Inhibition > 80.0 %
Reference
Title : Authors : Publication Year : Journal :
ALDUMASTAT
Inhibition of CYP3A4 (unknown origin) using BFC as substrate at 10 uM
—
Inhibition < 50.0 %
Reference
Title : Authors : Publication Year : Journal :
ALDUMASTAT
Inhibition of CYP3A4 (unknown origin) using DBF as substrate at 10 uM
—
Inhibition < 50.0 %
Reference
Title : Authors : Publication Year : Journal :
ZIMLOVISERTIB
Inhibition of CYP3A4 in human liver microsomes at 3 uM
Liver
Inhibition < 5.0 %
Reference
Title : Discovery of Clinical Candidate 1-{[(2S,3S,4S)-3-Ethyl-4-fluoro-5-oxopyrrolidin-2-yl]methoxy}-7-methoxyisoquinoline-6-carboxamide (PF-06650833), a Potent, Selective Inhibitor of Interleukin-1 Receptor Associated Kinase 4 (IRAK4), by Fragment-Based Drug Design. Authors : Lee KL, Ambler CM, Anderson DR, Boscoe BP, Bree AG, Brodfuehrer JI, Chang JS, Choi C, Chung S, Curran KJ, Day JE, Dehnhardt CM, Dower K, Drozda SE, Frisbie RK, Gavrin LK, Goldberg JA, Han S, Hegen M, Hepworth D, Hope HR, Kamtekar S, Kilty IC, Lee A, Lin LL, Lovering FE, Lowe MD, Mathias JP, Morgan HM, Murphy EA, Papaioannou N, Patny A, Pierce BS, Rao VR, Saiah E, Samardjiev IJ, Samas BM, Shen MWH, Shin JH, Soutter HH, Strohbach JW, Symanowicz PT, Thomason JR, Trzupek JD, Vargas R, Vincent F, Yan J, Zapf CW, Wright SW. Publication Year :
2017 Journal : J Med Chem Abstract : Through fragment-based drug design focused on engaging the active site of IRAK4 and leveraging three-dimensional topology in a ligand-efficient manner, a micromolar hit identified from a screen of a Pfizer fragment library was optimized to afford IRAK4 inhibitors with nanomolar potency in cellular assays. The medicinal chemistry effort featured the judicious placement of lipophilicity, informed by co-crystal structures with IRAK4 and optimization of ADME properties to deliver clinical candidate PF-06650833 (compound 40). This compound displays a 5-unit increase in lipophilic efficiency from the fragment hit, excellent kinase selectivity, and pharmacokinetic properties suitable for oral administration. DOI : 10.1021/acs.jmedchem.7b00231
LUVIXASERTIB
Inhibition of CYP3A4 (unknown origin) preincubated for 10 mins followed by addition of BFC as substrate measured after 10 mins in presence of NADP by fluorescence analysis
—
IC50 = 510.0 nM
Reference
Title : Authors : Publication Year : Journal :
DS-1971
Inhibition of CYP3A4 (unknown origin) at 10 uM relative to control
—
Inhibition = 56.6 %
Reference
Title : Discovery of DS-1971a, a Potent, Selective Na<sub>V</sub>1.7 Inhibitor. Authors : Shinozuka T, Kobayashi H, Suzuki S, Tanaka K, Karanjule N, Hayashi N, Tsuda T, Tokumaru E, Inoue M, Ueda K, Kimoto H, Domon Y, Takahashi S, Kubota K, Yokoyama T, Shimizugawa A, Koishi R, Fujiwara C, Asano D, Sakakura T, Takasuna K, Abe Y, Watanabe T, Kitano Y. Publication Year :
2020 Journal : J Med Chem Abstract : A highly potent, selective Na<sub>V</sub>1.7 inhibitor, DS-1971a, has been discovered. Exploration of the left-hand phenyl ring of sulfonamide derivatives (<b>I</b> and <b>II</b>) led to the discovery of novel series of cycloalkane derivatives with high Na<sub>V</sub>1.7 inhibitory potency in vitro. As the right-hand heteroaromatic ring affected the mechanism-based inhibition liability of CYP3A4, replacement of this moiety resulted in the generation of 4-pyrimidyl derivatives. Additionally, GSH adducts formation, which can cause idiosyncratic drug toxicity, was successfully avoided by this modification. An additional optimization led to the discovery of DS-1971a. In preclinical studies, DS-1971a demonstrated highly potent selective in vitro profile with robust efficacy in vivo. DS-1971a exhibited a favorable toxicological profile, which enabled multiple-dose studies of up to 600 mg bid or 400 mg tid (1200 mg/day) administered for 14 days to healthy human males. DS-1971a is expected to exert potent efficacy in patients with peripheral neuropathic pain, with a favorable safety profile. DOI : 10.1021/acs.jmedchem.0c00259
DS-1971
Inhibition of CYP3A4 (unknown origin) at 10 uM relative to control
—
Inhibition = 59.5 %
Reference
Title : Discovery of DS-1971a, a Potent, Selective Na<sub>V</sub>1.7 Inhibitor. Authors : Shinozuka T, Kobayashi H, Suzuki S, Tanaka K, Karanjule N, Hayashi N, Tsuda T, Tokumaru E, Inoue M, Ueda K, Kimoto H, Domon Y, Takahashi S, Kubota K, Yokoyama T, Shimizugawa A, Koishi R, Fujiwara C, Asano D, Sakakura T, Takasuna K, Abe Y, Watanabe T, Kitano Y. Publication Year :
2020 Journal : J Med Chem Abstract : A highly potent, selective Na<sub>V</sub>1.7 inhibitor, DS-1971a, has been discovered. Exploration of the left-hand phenyl ring of sulfonamide derivatives (<b>I</b> and <b>II</b>) led to the discovery of novel series of cycloalkane derivatives with high Na<sub>V</sub>1.7 inhibitory potency in vitro. As the right-hand heteroaromatic ring affected the mechanism-based inhibition liability of CYP3A4, replacement of this moiety resulted in the generation of 4-pyrimidyl derivatives. Additionally, GSH adducts formation, which can cause idiosyncratic drug toxicity, was successfully avoided by this modification. An additional optimization led to the discovery of DS-1971a. In preclinical studies, DS-1971a demonstrated highly potent selective in vitro profile with robust efficacy in vivo. DS-1971a exhibited a favorable toxicological profile, which enabled multiple-dose studies of up to 600 mg bid or 400 mg tid (1200 mg/day) administered for 14 days to healthy human males. DS-1971a is expected to exert potent efficacy in patients with peripheral neuropathic pain, with a favorable safety profile. DOI : 10.1021/acs.jmedchem.0c00259
EMAVUSERTIB
Inhibition of CYP3A4 (unknown origin) at 1 uM relative to control
Mechanism based inhibition of human cytochrome P450 3A5 measured by testosterone hydroxylation
—
Ki = 200.0 nM
Reference
Title : Cytochrome p450 enzymes mechanism based inhibitors: common sub-structures and reactivity. Authors : Fontana E, Dansette PM, Poli SM. Publication Year :
2005 Journal : Curr. Drug Metab. Abstract : The inhibition of human cytochrome P450s (CYPs) is one of the most common mechanisms which can lead to drug-drug interactions. The inhibition of CYPs can be reversible (competitive or non-competitive) or irreversible. Irreversible inhibition usually derives from activation of a drug by CYPs into a reactive metabolite, which tightly binds to the enzyme active site, leading to a long lasting inactivation. This process is called "mechanism based inhibition" or "suicide inhibition". The irreversible inactivation usually implies the formation of a covalent bond between the metabolite and the enzyme, which can lead to hapten formation and can in some cases trigger an autoimmune-response. For these reasons it is of utmost importance to study the mechanism of the CYP inhibition of new potential drugs as early as possible during the drug discovery process. The literature on CYPs is vast and covers numerous aspects of their biology and biochemistry, however to our knowledge there is no general and systematic review focusing on mechanism-based inhibitors; we have reviewed the literature and compiled all the available data on chemical entities, which are known to be CYP suicide inhibitors. Each compound is reported together with its chemical structure, the CYP isoform and the parameters describing the inactivation. Literature references are reported together with their PMID (PubMed ID number) to allow a fast retrieval of the papers. This review offers a quick reference to help predict liabilities of new chemical entities without carrying out extensive in vitro work, and will hopefully help in designing safer drugs. DOI : 10.2174/138920005774330639
CANNABIDIOL
Competitive inhibition of human recombinant CYP3A5 expressed in baculovirus-infected insect cells using diltiazem as substrate incubated for 15 mins followed by NADPH-generating system addition by Michaelis-Menten plot analysis
—
Ki = 190.0 nM
Reference
Title : The Essential Medicinal Chemistry of Cannabidiol (CBD). Authors : Nelson KM,Bisson J,Singh G,Graham JG,Chen SN,Friesen JB,Dahlin JL,Niemitz M,Walters MA,Pauli GF Publication Year :
2020 Journal : J Med Chem Abstract : This Perspective of the published essential medicinal chemistry of cannabidiol (CBD) provides evidence that the popularization of CBD-fortified or CBD-labeled health products and CBD-associated health claims lacks a rigorous scientific foundation. CBD's reputation as a cure-all puts it in the same class as other "natural" panaceas, where valid ethnobotanicals are reduced to single, purportedly active ingredients. Such reductionist approaches oversimplify useful, chemically complex mixtures in an attempt to rationalize the commercial utility of natural compounds and exploit the "natural" label. Literature evidence associates CBD with certain semiubiquitous, broadly screened, primarily plant-based substances of undocumented purity that interfere with bioassays and have a low likelihood of becoming therapeutic agents. Widespread health challenges and pandemic crises such as SARS-CoV-2 create circumstances under which scientists must be particularly vigilant about healing claims that lack solid foundational data. Herein, we offer a critical review of the published medicinal chemistry properties of CBD, as well as precise definitions of CBD-containing substances and products, distilled to reveal the essential factors that impact its development as a therapeutic agent. DOI : 10.1021/acs.jmedchem.0c00724
LAPATINIB
Inhibition of human recombinant CYP3A5 assessed as reduction of enzyme-CO complex formation at 50 uM preincubated for 3 to 5 mins with NADPH B followed by NADPH A addition measured after 30 mins by CO difference spectroscopy relative to control
—
Inhibition = 22.0 %
Reference
Title : Interaction of lapatinib with cytochrome P450 3A5. Authors : Chan EC, New LS, Chua TB, Yap CW, Ho HK, Nelson SD. Publication Year :
2012 Journal : Drug Metab. Dispos. Abstract : Lapatinib, an oral tyrosine kinase inhibitor used for breast cancer, has been reported to cause idiosyncratic hepatotoxicity. Recently, it has been found that lapatinib forms a metabolite-inhibitor complex (MIC) with CYP3A4 via the formation of an alkylnitroso intermediate. Because CYP3A5 is highly polymorphic compared with CYP3A4 and also oxidizes lapatinib, we investigated the interactions of lapatinib with CYP3A5. Lapatinib inactivated CYP3A5 in a time-, concentration-, and NADPH-dependent manner using testosterone as a probe substrate with K(I) and k(inact) values of 0.0376 mM and 0.0226 min(-1), respectively. However, similar results were not obtained when midazolam was used as the probe substrate, suggesting that inactivation of CYP3A5 by lapatinib is site-specific. Poor recovery of CYP3A5 activity postdialysis and the lack of a Soret peak confirmed that lapatinib does not form a MIC with CYP3A5. The reduced CO difference spectrum further suggested that a large fraction of the reactive metabolite of lapatinib is covalently adducted to the apoprotein of CYP3A5. GSH trapping of a reactive metabolite of lapatinib formed by CYP3A5 confirmed the formation of a quinoneimine-GSH adduct derived from the O-dealkylated metabolite of lapatinib. In silico docking studies supported the preferential formation of an O-dealkylated metabolite of lapatinib by CYP3A5 compared with an N-hydroxylation reaction that is predominantly catalyzed by CYP3A4. In conclusion, lapatinib appears to be a mechanism-based inactivator of CYP3A5 via adduction of a quinoneimine metabolite. DOI : 10.1124/dmd.112.044958
LAPATINIB
Inhibition of recombinant CYP3A5 (unknown origin)-mediated midazolam 1-hydroxylation at 50 uM
—
Inhibition = 45.0 %
Reference
Title : Metabolism-dependent inhibition of CYP3A4 by lapatinib: evidence for formation of a metabolic intermediate complex with a nitroso/oxime metabolite formed via a nitrone intermediate. Authors : Barbara JE, Kazmi F, Parkinson A, Buckley DB. Publication Year :
2013 Journal : Drug Metab. Dispos. Abstract : Metabolism-dependent inhibition (MDI) of cytochrome P450 (P450) enzymes has the potential to cause clinically relevant drug-drug interactions. In the case of several alkylamine drugs, MDI of P450 involves formation of a metabolite that binds quasi-irreversibly to the ferrous heme iron to form a metabolic intermediate (MI) complex. The specific metabolites coordinately bound to ferrous iron and the pathways leading to MI complex formation are the subject of debate. We describe an approach combining heme iron oxidation with potassium ferricyanide and metabolite profiling to probe the mechanism of MI complex-based CYP3A4 inactivation by the secondary alkylamine drug lapatinib. Ten metabolites formed from lapatinib by CYP3A4-mediated heteroatom dealkylation, C-hydroxylation, N-oxygenation with or without further oxidation, or a combination thereof, were detected by accurate mass spectrometry. The abundance of one metabolite, the N-dealkylated nitroso/oxime lapatinib metabolite (M9), correlated directly with the prevalence or the disruption of the MI complex with CYP3A4. Nitroso/oxime metabolite formation from secondary alkylamines has been proposed to occur through two possible pathways: (1) sequential N-dealkylation, N-hydroxylation, and dehydrogenation (primary hydroxylamine pathway) or (2) N-hydroxylation with dehydrogenation to yield a nitrone followed by N-dealkylation (secondary hydroxylamine pathway). All intermediates for the secondary hydroxylamine pathway were detected but the primary N-hydroxylamine intermediate of the primary hydroxylamine pathway was not. Our findings support the mechanism of lapatinib CYP3A4 inactivation as MI complex formation with the nitroso metabolite formed through the secondary hydroxylamine and nitrone pathway, rather than by N-dealkylation to the primary amine followed by N-hydroxylation and dehydrogenation as is usually assumed. DOI : 10.1124/dmd.113.051151
RITONAVIR
Mechanism based inhibition of human cytochrome P450 3A5
—
Ki = 120.0 nM
Reference
Title : Cytochrome p450 enzymes mechanism based inhibitors: common sub-structures and reactivity. Authors : Fontana E, Dansette PM, Poli SM. Publication Year :
2005 Journal : Curr. Drug Metab. Abstract : The inhibition of human cytochrome P450s (CYPs) is one of the most common mechanisms which can lead to drug-drug interactions. The inhibition of CYPs can be reversible (competitive or non-competitive) or irreversible. Irreversible inhibition usually derives from activation of a drug by CYPs into a reactive metabolite, which tightly binds to the enzyme active site, leading to a long lasting inactivation. This process is called "mechanism based inhibition" or "suicide inhibition". The irreversible inactivation usually implies the formation of a covalent bond between the metabolite and the enzyme, which can lead to hapten formation and can in some cases trigger an autoimmune-response. For these reasons it is of utmost importance to study the mechanism of the CYP inhibition of new potential drugs as early as possible during the drug discovery process. The literature on CYPs is vast and covers numerous aspects of their biology and biochemistry, however to our knowledge there is no general and systematic review focusing on mechanism-based inhibitors; we have reviewed the literature and compiled all the available data on chemical entities, which are known to be CYP suicide inhibitors. Each compound is reported together with its chemical structure, the CYP isoform and the parameters describing the inactivation. Literature references are reported together with their PMID (PubMed ID number) to allow a fast retrieval of the papers. This review offers a quick reference to help predict liabilities of new chemical entities without carrying out extensive in vitro work, and will hopefully help in designing safer drugs. DOI : 10.2174/138920005774330639
TACROLIMUS
Inhibition of human CYP3A5 expressed in supersomes assessed inhibition of 1'-OH midazolam formation preincubated at 2 uM followed by addition of 20-fold diluted buffer with 20 uM MDZ by LC-MS method in presence of NADPH
—
Inhibition = 52.0 %
Reference
Title : Cyclosporine A- and tacrolimus-mediated inhibition of CYP3A4 and CYP3A5 in vitro. Authors : Amundsen R, Åsberg A, Ohm IK, Christensen H. Publication Year :
2012 Journal : Drug Metab. Dispos. Abstract : Cyclosporine A (CsA) and tacrolimus (Tac) are immunosuppressive drugs used in the majority of patients with solid organ transplants, generally in combination with a wide range of drugs. CsA and Tac seem not only to be substrates of CYP3A but have also been described as inhibitors of CYP3A. For CsA, in particular, inhibition of CYP3A has been suggested as the main mechanism of interactions seen clinically with various drugs. The aim of this study was to investigate the inhibitory effect and inhibition characteristics of CsA and Tac on CYP3A4 and CYP3A5 in vitro and to evaluate its clinical relevance. Inhibition by CsA and Tac was studied using midazolam as the probe substrate in coincubation and preincubation investigations using human liver microsomes (HLMs) as well as specific CYP3A4- and CYP3A5-expressing insect microsomes (Supersomes). In vitro-in vivo extrapolations (IVIVEs) were performed to evaluate the clinical relevance of the inhibition. Both CsA and Tac competitively inhibited CYP3A in HLMs, showing inhibition constants (K(i)) of 0.98 and 0.61 μM, respectively. Experiments in Supersomes revealed that Tac inhibited both CYP3A4 and CYP3A5, whereas CsA only inhibited CYP3A4. In contrast to the HLM experiments, studies in Supersomes showed inhibition by Tac to be NADPH- and time-dependent, with a 5-fold reduction in IC(50) after preincubation, supporting a time-dependent inhibition mechanism in recombinant microsomes. By application of HLM data, IVIVE estimated the area under the concentration versus time curve of midazolam to increase by 73 and 27% with CsA and Tac, respectively. The inhibitory effect was predominantly on the intestinal level, whereas hepatic intrinsic clearance seemed unaffected. DOI : 10.1124/dmd.111.043018
TACROLIMUS
Inhibition of human CYP3A5 expressed in supersomes assessed inhibition of 1'-OH midazolam formation by LC-MS method
—
IC50 = 630.0 nM
Reference
Title : Cyclosporine A- and tacrolimus-mediated inhibition of CYP3A4 and CYP3A5 in vitro. Authors : Amundsen R, Åsberg A, Ohm IK, Christensen H. Publication Year :
2012 Journal : Drug Metab. Dispos. Abstract : Cyclosporine A (CsA) and tacrolimus (Tac) are immunosuppressive drugs used in the majority of patients with solid organ transplants, generally in combination with a wide range of drugs. CsA and Tac seem not only to be substrates of CYP3A but have also been described as inhibitors of CYP3A. For CsA, in particular, inhibition of CYP3A has been suggested as the main mechanism of interactions seen clinically with various drugs. The aim of this study was to investigate the inhibitory effect and inhibition characteristics of CsA and Tac on CYP3A4 and CYP3A5 in vitro and to evaluate its clinical relevance. Inhibition by CsA and Tac was studied using midazolam as the probe substrate in coincubation and preincubation investigations using human liver microsomes (HLMs) as well as specific CYP3A4- and CYP3A5-expressing insect microsomes (Supersomes). In vitro-in vivo extrapolations (IVIVEs) were performed to evaluate the clinical relevance of the inhibition. Both CsA and Tac competitively inhibited CYP3A in HLMs, showing inhibition constants (K(i)) of 0.98 and 0.61 μM, respectively. Experiments in Supersomes revealed that Tac inhibited both CYP3A4 and CYP3A5, whereas CsA only inhibited CYP3A4. In contrast to the HLM experiments, studies in Supersomes showed inhibition by Tac to be NADPH- and time-dependent, with a 5-fold reduction in IC(50) after preincubation, supporting a time-dependent inhibition mechanism in recombinant microsomes. By application of HLM data, IVIVE estimated the area under the concentration versus time curve of midazolam to increase by 73 and 27% with CsA and Tac, respectively. The inhibitory effect was predominantly on the intestinal level, whereas hepatic intrinsic clearance seemed unaffected. DOI : 10.1124/dmd.111.043018
TACROLIMUS
Inhibition of human CYP3A5 expressed in supersomes assessed inhibition of 1'-OH midazolam formation preincubated with compound before substrate addition by LC-MS method
—
IC50 = 140.0 nM
Reference
Title : Cyclosporine A- and tacrolimus-mediated inhibition of CYP3A4 and CYP3A5 in vitro. Authors : Amundsen R, Åsberg A, Ohm IK, Christensen H. Publication Year :
2012 Journal : Drug Metab. Dispos. Abstract : Cyclosporine A (CsA) and tacrolimus (Tac) are immunosuppressive drugs used in the majority of patients with solid organ transplants, generally in combination with a wide range of drugs. CsA and Tac seem not only to be substrates of CYP3A but have also been described as inhibitors of CYP3A. For CsA, in particular, inhibition of CYP3A has been suggested as the main mechanism of interactions seen clinically with various drugs. The aim of this study was to investigate the inhibitory effect and inhibition characteristics of CsA and Tac on CYP3A4 and CYP3A5 in vitro and to evaluate its clinical relevance. Inhibition by CsA and Tac was studied using midazolam as the probe substrate in coincubation and preincubation investigations using human liver microsomes (HLMs) as well as specific CYP3A4- and CYP3A5-expressing insect microsomes (Supersomes). In vitro-in vivo extrapolations (IVIVEs) were performed to evaluate the clinical relevance of the inhibition. Both CsA and Tac competitively inhibited CYP3A in HLMs, showing inhibition constants (K(i)) of 0.98 and 0.61 μM, respectively. Experiments in Supersomes revealed that Tac inhibited both CYP3A4 and CYP3A5, whereas CsA only inhibited CYP3A4. In contrast to the HLM experiments, studies in Supersomes showed inhibition by Tac to be NADPH- and time-dependent, with a 5-fold reduction in IC(50) after preincubation, supporting a time-dependent inhibition mechanism in recombinant microsomes. By application of HLM data, IVIVE estimated the area under the concentration versus time curve of midazolam to increase by 73 and 27% with CsA and Tac, respectively. The inhibitory effect was predominantly on the intestinal level, whereas hepatic intrinsic clearance seemed unaffected. DOI : 10.1124/dmd.111.043018
BRIVANIB
Drug metabolism in human liver microsomes assessed as 1 uM ketaconazole-mediated inhibition of CYP3A5-mediated (R)-1-(4-(4-fluoro-2-(hydroxymethyl)-1H-indol-5-yloxy)-5-methylpyrrolo[1,2-f][1,2,4]triazin-6-yloxy)propan-2-ol formation at 2 and 20 uM for 30 mins by LC/MS analysis
Liver
Inhibition = 50.0 %
Reference
Title : Identification of the oxidative and conjugative enzymes involved in the biotransformation of brivanib. Authors : Gong J, Gan J, Iyer RA. Publication Year :
2012 Journal : Drug Metab. Dispos. Abstract : Brivanib alaninate, the L-alanine ester prodrug of brivanib, is currently being developed as an anticancer agent. In humans, brivanib alaninate is rapidly hydrolyzed to brivanib. Prominent biotransformation pathways of brivanib included oxidation and direct sulfate conjugation. A series of in vitro studies were conducted to identify the human esterases involved in the prodrug hydrolysis and to identify the primary human cytochrome P450 and sulfotransferase (SULT) enzymes involved in the metabolism of brivanib. Brivanib alaninate was efficiently converted to brivanib in the presence of either human carboxylesterase 1 or carboxylesterase 2. Because esterases are ubiquitous, it is likely that multiple esterases are involved in the hydrolysis. Oxidation of brivanib in human liver microsomes (HLM) primarily formed a hydroxylated metabolite (M7). Incubation of brivanib with human cDNA-expressed P450 enzymes and with HLM in the presence of selective chemical inhibitors and monoclonal P450 antibodies demonstrated that CYP1A2 and CYP3A4 were the major contributors for the formation of M7. Direct sulfation of brivanib was catalyzed by multiple SULT enzymes, including SULT1A1, SULT1B1, SULT2A1, SULT1A3, and SULT1E1. Because the primary in vitro oxidative metabolite (M7) was not detected in humans after oral doses of brivanib alaninate, further metabolism studies of M7 in HLM and human liver cytosol were performed. The data demonstrated that M7 was metabolized to the prominent metabolites observed in humans. Overall, multiple enzymes are involved in the metabolism of brivanib, suggesting a low potential for drug-drug interactions either through polymorphism or through inhibition of a particular drug-metabolizing enzyme. DOI : 10.1124/dmd.111.042457
BRIVANIB
Drug metabolism in human liver microsomes assessed as monoclonal antibody-mediated inhibition of CYP3A5-mediated (R)-1-(4-(4-fluoro-2-(hydroxymethyl)-1H-indol-5-yloxy)-5-methylpyrrolo[1,2-f][1,2,4]triazin-6-yloxy)propan-2-ol formation at 2 and 20 uM for 30 mins by LC/MS analysis
Liver
Inhibition = 50.0 %
Reference
Title : Identification of the oxidative and conjugative enzymes involved in the biotransformation of brivanib. Authors : Gong J, Gan J, Iyer RA. Publication Year :
2012 Journal : Drug Metab. Dispos. Abstract : Brivanib alaninate, the L-alanine ester prodrug of brivanib, is currently being developed as an anticancer agent. In humans, brivanib alaninate is rapidly hydrolyzed to brivanib. Prominent biotransformation pathways of brivanib included oxidation and direct sulfate conjugation. A series of in vitro studies were conducted to identify the human esterases involved in the prodrug hydrolysis and to identify the primary human cytochrome P450 and sulfotransferase (SULT) enzymes involved in the metabolism of brivanib. Brivanib alaninate was efficiently converted to brivanib in the presence of either human carboxylesterase 1 or carboxylesterase 2. Because esterases are ubiquitous, it is likely that multiple esterases are involved in the hydrolysis. Oxidation of brivanib in human liver microsomes (HLM) primarily formed a hydroxylated metabolite (M7). Incubation of brivanib with human cDNA-expressed P450 enzymes and with HLM in the presence of selective chemical inhibitors and monoclonal P450 antibodies demonstrated that CYP1A2 and CYP3A4 were the major contributors for the formation of M7. Direct sulfation of brivanib was catalyzed by multiple SULT enzymes, including SULT1A1, SULT1B1, SULT2A1, SULT1A3, and SULT1E1. Because the primary in vitro oxidative metabolite (M7) was not detected in humans after oral doses of brivanib alaninate, further metabolism studies of M7 in HLM and human liver cytosol were performed. The data demonstrated that M7 was metabolized to the prominent metabolites observed in humans. Overall, multiple enzymes are involved in the metabolism of brivanib, suggesting a low potential for drug-drug interactions either through polymorphism or through inhibition of a particular drug-metabolizing enzyme. DOI : 10.1124/dmd.111.042457
NELFINAVIR
Mechanism based inhibition of human cytochrome P450 3A5 measured by testosterone hydroxylation
—
Ki = 570.0 nM
Reference
Title : Cytochrome p450 enzymes mechanism based inhibitors: common sub-structures and reactivity. Authors : Fontana E, Dansette PM, Poli SM. Publication Year :
2005 Journal : Curr. Drug Metab. Abstract : The inhibition of human cytochrome P450s (CYPs) is one of the most common mechanisms which can lead to drug-drug interactions. The inhibition of CYPs can be reversible (competitive or non-competitive) or irreversible. Irreversible inhibition usually derives from activation of a drug by CYPs into a reactive metabolite, which tightly binds to the enzyme active site, leading to a long lasting inactivation. This process is called "mechanism based inhibition" or "suicide inhibition". The irreversible inactivation usually implies the formation of a covalent bond between the metabolite and the enzyme, which can lead to hapten formation and can in some cases trigger an autoimmune-response. For these reasons it is of utmost importance to study the mechanism of the CYP inhibition of new potential drugs as early as possible during the drug discovery process. The literature on CYPs is vast and covers numerous aspects of their biology and biochemistry, however to our knowledge there is no general and systematic review focusing on mechanism-based inhibitors; we have reviewed the literature and compiled all the available data on chemical entities, which are known to be CYP suicide inhibitors. Each compound is reported together with its chemical structure, the CYP isoform and the parameters describing the inactivation. Literature references are reported together with their PMID (PubMed ID number) to allow a fast retrieval of the papers. This review offers a quick reference to help predict liabilities of new chemical entities without carrying out extensive in vitro work, and will hopefully help in designing safer drugs. DOI : 10.2174/138920005774330639
AZD-3759
Inhibition of CYP3A5 (unknown origin)
—
IC50 > 50.0 nM
Reference
Title : Discovery and Evaluation of Clinical Candidate AZD3759, a Potent, Oral Active, Central Nervous System-Penetrant, Epidermal Growth Factor Receptor Tyrosine Kinase Inhibitor. Authors : Zeng Q, Wang J, Cheng Z, Chen K, Johnström P, Varnäs K, Li DY, Yang ZF, Zhang X. Publication Year :
2015 Journal : J. Med. Chem. Abstract : Recent reports suggest that an increasing number of patients with lung cancer, especially those with activating mutations of the epidermal growth factor receptor (EGFR), also present with brain metastases and leptomeningeal metastases. These patients have poor prognosis as there are no approved drugs for these indications. Available agents have poor efficacy for these patients even at well above their standard dose. Herein, we report the discovery of (4-[(3-chloro-2-fluorophenyl)amino]-7-methoxyquinazolin-6-yl (2R)-2,4-dimethylpiperazine-1-carboxylate 1m (AZD3759), an investigational drug currently in Phase 1 clinical trial, which has excellent central nervous system penetration and which induces profound regression of brain metastases in a mouse model. DOI : 10.1021/acs.jmedchem.5b01073
SUVECALTAMIDE
Inhibition of recombinant human CYP3A5 in human liver microsomes at 1 uM relative to control
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