ABSTRACT:The metabolism and disposition of [ 14 C]apixaban, an orally bioavailable, highly selective, and direct acting/reversible factor Xa inhibitor, was investigated in 10 healthy male subjects without (group 1, n ‫؍‬ 6) and with bile collection (group 2, n ‫؍‬ 4) after a single 20-mg oral dose. Urine, blood, and feces samples were collected from all subjects. Bile samples were also collected for 3 to 8 h after dosing from group 2 subjects. There were no serious adverse events or discontinuations due to adverse effects. In plasma, apixaban was the major circulating component and O-demethyl apixaban sulfate, a stable and water-soluble metabolite, was the significant metabolite. The exposure of apixaban (C max and area under the plasma concentration versus time curve) in subjects with bile collection was generally similar to that in subjects without bile collection. The administered dose was recovered in feces (group 1, 56.0%; group 2, 46.7%) and urine (group 1, 24.5%; group 2, 28.8%), with the parent drug representing approximately half of the recovered dose. Biliary excretion represented a minor elimination pathway (2.44% of the administered dose) from group 2 subjects within the limited collection period. Metabolic pathways identified for apixaban included O-demethylation, hydroxylation, and sulfation of hydroxylated O-demethyl apixaban. Thus, apixaban is an orally bioavailable inhibitor of factor Xa with elimination pathways that include metabolism and renal excretion.Thromboembolic events, including acute myocardial infarction, unstable angina, deep vein thrombosis, pulmonary embolism, and ischemic stroke continue to be the leading cause of morbidity and mortality in the United States and other Western countries (Heit et al., 2005;Rosamond et al., 2007). Current therapies for the treatment and prevention of thromboembolic events, such as vitamin K antagonists (e.g., warfarin), heparin, and low-molecular-weight heparin (e.g., enoxaparin), are suboptimal (O'Donnell and Weitz, 2004;Wittkowsky, 2004;Campbell, 2006). However, the requirement for intravenous or subcutaneous injection and/or the need for careful monitoring because of the risk of excessive bleeding or unpredictable/inconsistent pharmacokinetics (PK) can complicate administration and present barriers to the use of these agents (O'Brien and Caro, 2002;Wittkowsky, 2004;Campbell, 2006). Therefore, new, orally active anticoagulants with predictable pharmacokinetic profiles that can be administered with a reduced need for monitoring are needed.Factor Xa is a key serine protease in the coagulation cascade and is a promising target enzyme for new therapeutic agents for the treatment and prevention of arterial and venous thrombosis (Kaiser, 2002;Samama, 2002;Walenga et al., 2003). In particular, factor Xa plays a critical role in blood coagulation, serving as the juncture between the extrinsic (tissue factor initiated) and intrinsic (surface activation and amplification) systems (Mann et al., 2003). Factor Xa forms the prothrombinase complex with phospholi...

The C-aryl glucoside 6 (dapagliflozin) was identified as a potent and selective hSGLT2 inhibitor which reduced blood glucose levels in a dose-dependent manner by as much as 55% in hyperglycemic streptozotocin (STZ) rats. These findings, combined with a favorable ADME profile, have prompted clinical evaluation of dapagliflozin for the treatment of type 2 diabetes.

In the present study, an open-label, three-treatment, threeperiod clinical study of rosuvastatin (RSV) and rifampicin (RIF) when administered alone and in combination was conducted in 12 male healthy subjects to determine if coproporphyrin I (CP-I) and coproporphyrin III (CP-III) could serve as clinical biomarkers for organic anion transporting polypeptide 1B1 (OATP1B1) and 1B3 that belong to the solute carrier organic anion gene subfamily. Genotyping of the human OATP1B1 gene was performed in all 12 subjects and confirmed absence of OATP1B1*5 and OATP1B1*15 mutations. Average plasma concentrations of CP-I and CP-III prior to drug administration were 0.91 6 0.21 and 0.15 6 0.04 nM, respectively, with minimum fluctuation over the three periods. CP-I was passively eliminated, whereas CP-III was actively secreted from urine. Administration of RSV caused no significant changes in the plasma and urinary profiles of CP-I and CP-III. RIF markedly increased the maximum plasma concentration (C max ) of CP-I and CP-III by 5.7-and 5.4-fold (RIF) or 5.7-and 6.5-fold (RIF1RSV), respectively, as compared with the predose values. The area under the plasma concentration curves from time 0 to 24 h (AUC 0-24h ) of CP-I and CP-III with RIF and RSV increased by 4.0-and 3.3-fold, respectively, when compared with RSV alone. In agreement with this finding, C max and AUC 0-24h of RSV increased by 13.2-and 5.0-fold, respectively, when RIF was coadministered. Collectively, we conclude that CP-I and CP-III in plasma and urine can be appropriate endogenous biomarkers specifically and reliably reflecting OATP inhibition, and thus the measurement of these molecules can serve as a useful tool to assess OATP drug-drug interaction liabilities in early clinical studies.

ABSTRACT:Several human immunodeficiency virus (HIV) protease inhibitors, including atazanavir, indinavir, lopinavir, nelfinavir, ritonavir, and saquinavir, were tested for their potential to inhibit uridine 5-diphospho-glucuronosyltransferase (UGT) activity. Experiments were performed with human cDNA-expressed enzymes (UGT1A1, 1A3, 1A4, 1A6, 1A9, and 2B7) as well as human liver microsomes.All of the protease inhibitors tested were inhibitors of UGT1A1, UGT1A3, and UGT1A4 with IC 50 values that ranged from 2 to 87 M. The IC 50 values found for all compounds for UGT1A6, 1A9, and 2B7 were >100 M. The inhibition (IC 50 ) of UGT1A1 was similar when tested against the human cDNA-expressed enzyme or human liver microsomes for atazanavir, indinavir, and saquinavir (2.4, 87, and 7.3 M versus 2.5, 68, and 5.0 M, respectively). By analysis of the double-reciprocal plots of bilirubin glucuronidation activities at different bilirubin concentrations in the presence of fixed concentrations of inhibitors, the UGT1A1 inhibition by atazanavir and indinavir was demonstrated to follow a linear mixed-type inhibition mechanism (K i ‫؍‬ 1.9 and 47.9 M, respectively). These results suggest that a direct inhibition of UGT1A1-mediated bilirubin glucuronidation may provide a mechanism for the reversible hyperbilirubinemia associated with administration of atazanavir as well as indinavir. In vitro-in vivo scaling with [I]/K i predicts that atazanavir and indinavir are more likely to induce hyperbilirubinemia than other HIV protease inhibitors studied when a free C max drug concentration was used. Our current study provides a unique example of in vitro-in vivo correlation for an endogenous UGT-mediated metabolic pathway.The HIV protease is an essential enzyme that cuts the viral gag-pol polyprotein into its functional subunits. Atazanavir, indinavir, saquinavir, lopinavir, ritonavir, and nelfinavir are HIV protease inhibitors. The structures of these HIV protease inhibitors are shown in Fig. 1. As opposed to atazanavir, which is an azapeptide protease inhibitor (Goldsmith and Perry, 2003), other listed HIV protease inhibitors are peptidomimetics sharing the same structural determinant, i.e., a hydroxyethylene or a hydroxyethylamine moiety, which makes them nonscissile HIV protease substrate analogs. These HIV protease inhibitors are metabolized primarily by hepatic CYP3A enzymes, and they are also inhibitors of CYP3A enzymes (Flexner, 2000;de Maat et al., 2003;Goldsmith and Perry, 2003;Ernest et al., 2005). All of these HIV protease inhibitors have a high protein binding (Ͼ98%), except indinavir and atazanavir, which have a protein binding of 60 and 86%, respectively. Most of the HIV protease inhibitors are bound to ␣1-acid glycoprotein instead of albumin (de Maat et al., 2003). There are no literature reports indicating that HIV protease inhibitors are good substrates for human UGT enzymes, although there is evidence to support indinavir as a substrate of UGTs (Balani et al., 1996).Glucuronidation represents a major pathway for the elimin...

ABSTRACT:An improved mass defect filter (MDF) method employing both drug and core structure filter templates was applied to the processing of high resolution liquid chromatography/mass spectrometry (LC/ MS) data for the detection and structural characterization of oxidative metabolites with mass defects similar to or significantly different from those of the parent drugs. The effectiveness of this approach was investigated using nefazodone as a model compound, which is known to undergo multiple common and uncommon oxidative reactions. Through the selective removal of all ions that fall outside of the preset filter windows, the MDF process facilitated the detection of all 14 nefazodone metabolites presented in human liver microsomes in the MDF-filtered chromatograms. The capability of the MDF approach to remove endogenous interferences from more complex biological matrices was examined by analyzing omeprazole metabolites in human plasma. The unprocessed mass chromatogram showed no distinct indication of metabolite peaks; however, after MDF processing, the metabolite peaks were easily identified in the chromatogram. Compared with precursor ion scan and neutral loss scan techniques, the MDF approach was shown to be more effective for the detection of metabolites in a complex matrix. The comprehensive metabolite detection capability of the MDF approach, together with accurate mass determination, makes high resolution LC/MS a useful tool for the screening and identification of both common and uncommon drug metabolites.The identification of drug metabolites, particularly metabolites formed through oxidation, reduction, or hydrolysis reactions, has become an integral part of the drug discovery and development process. These metabolites may have intrinsic pharmacological activity or display specific toxicity (Parkinson, 1996;Guengerich, 2000). In addition, most clinical drug-drug interactions are associated with oxidative biotransformation mediated by cytochrome P450 (Bjornsson et al., 2003). Although analytical sensitivity and the processing of data for liquid chromatography/mass spectrometry (LC/MS) have been tremendously improved in the last decade (Clarke et al., 2001;Kostiainen et al., 2003;Liu and Hop, 2005), the detection and identification of drug metabolites in complex biological matrices continue to be a challenge.Traditionally, detection of common or expected metabolites has been conducted on LC/MS data by generating extracted or reconstructed ion chromatograms corresponding to the expected protonated molecules of drug metabolites (Plumb et al., 2003). Over the last decade, product ion scanning techniques that use rule-based algorithms to generate a list of potential metabolite masses have been developed and continuously improved for rapid screening for common metabolites (Yu et al., 1999;Gangl et al., 2002;Lafaye et al., 2003). The technique employs a survey mode to search for the metabolites that are listed in the acquisition method. Both the detection of expected metabolites and the acquisition of their pr...