Tsuyoshi Ooie

PREDICTION OF IN VIVO DRUG METABOLISM IN THE HUMAN LIVER FROM IN VITRO METABOLISM DATA

As a new approach to predicting in vivo drug metabolism in humans, scaling of in vivo metabolic clearance from in vitro data obtained using human liver microsomes or hepatocytes is described in this review, based on the large number of literature data. Successful predictions were obtained for verapamil, loxtidine (lavoltidine), diazepam, lidocaine, phenacetin and some other compounds where CLint,in vitro is comparable with CLint,in vivo. On the other hand, for some metabolic reactions, differences in CLint,in vitro and CLint,in vivo greater than 5-fold were observed. The following factors are considered to be the cause of the differences: (1) metabolism in tissues other than liver, (2) incorrect assumption of rapid equilibrium of drugs between blood and hepatocytes, (3) presence of active transport through the sinusoidal membrane, and (4) interindividual variability. Furthermore, the possibility of predicting in vivo drug metabolic clearance from results obtained using a recombinant system of human P450 isozyme was described for a model compound, YM796, where the predicted metabolic clearances obtained from the recombinant system, taking account of the content of the P450 isozyme CYP3A4 in the human microsomes, were comparable with the observed clearances using human liver microsomes containing different amounts of CYP3A4. Even in the case where the first-pass metabolism exhibits nonlinearity, it appears to be possible to predict in vivo metabolic clearance from in vitro metabolic data.

Prediction of in vivo drug disposition from in vitro data based on physiological pharmacokinetics

Because of the increasing availability of human liver samples we now have a greater ability to predict in vivo drug disposition and pharmacokinetics in man from in vitro metabolic and binding studies. Firstly, we review several successful attempts to predict in vivo metabolic clearances in experimental animals and humans from in vitro biochemical parameters such as plasma protein binding and hepatic metabolism, based on anatomically and physiologically realistic pharmacokinetic models. Despite the success of this approach, however, there are still some difficulties in predicting in vivo hepatic metabolism in man using in vitro human liver samples due to the large inter‐individual differences arising from polymorphism (intrinsic variability) or differences in enzyme activity (extrinsic variability) due to the conditions under which liver samples have been kept. We propose a possible method to overcome these errors resulting from inter‐individual differences by applying the concept of a scaling factor. In the kinetic models used in prediction, we often make a number of assumptions, e.g. rapid equilibrium between the blood and hepatocytes, availability of only the unbound drug for uptake and elimination, and homogeneous distribution of enzymes along the path taken by the blood in the liver. However, recent evidence suggests that these assumptions are not necessarily valid. As examples involving the first and second assumptions, respectively, there is the plasma‐membrane‐permeability‐limited metabolism of a high‐clearance drug, 4‐methylumbelliferone, and the albumin‐mediated uptake of amphiphatic drugs. The multiple‐indicator dilution method (MID) is useful for estimating the membrane permeability of drugs in liver perfusion systems where the spatial organization and cell polarity of the liver are maintained. If the aforementioned factors are taken into consideration and membrane permeabilities using human hepatocytes and/or subcellular fractions such as microsomes are measured under conditions close to those in vivo, much more reliable predictions of drug hepatic clearance in man may become possible.

Characterization of the Transport Properties of a Quinolone Antibiotic, Fleroxacin, in Rat Choroid Plexus

AbstractPurpose. It is reported that the cerebrospinal fluid (CSF) to plasma unbound concentration ratio of fleroxacin at steady-state is approximately 0.5 in experimental animals. These results can be accounted for by assuming the presence of an active transport system for the efflux of this compound across the choroid plexus. In the present study, the transport system for fleroxacin was characterized in isolated rat choroid plexus. Methods. Choroid plexus was isolated from the lateral ventricles of rats. The accumulation of [14C] fleroxacin or [3H] benzylpenicillin by the choroid plexus was examined by the centrifugal filtration method. Results. The accumulation of [14C] fleroxacin by the rat isolated choroid plexus was significantly inhibited by metabolic inhibitors (rotenone, 30 µM and carbonyl cyanide p-trifluorometh oxyphenylhydrazone (FCCP), 100 µM) and sulfhydryl reagent (p-chloromer-curibenzenesulfonic acid (PCMBS), 100 µM). This accumulation was composed of a saturable component (Vmax = 240 pmol·min−1·µl tissue−1, Km = 664 µM) and non-saturable one (P = 0.424 min−1·µl tissue−1). Accumulation of fleroxacin was competitively inhibited by benzylpenicillin and probenecid with Ki values of 29 µM and 51 µM, respectively. These values are comparable with the Km of benzylpenicillin transport and the Ki of probenecid for the benzylpenicillin transport at the choroid plexus, respectively. Furthermore, fleroxacin inhibited competitively the accumulation of [3H] benzylpenicillin with a Ki of 384 µM, a value comparable with the Km of [14C] fleroxacin transport. Conclusions. Fleroxacin and benzylpenicillin showed mutual competitive inhibition, suggesting that both are transported via a common transport system in the choroid plexus and are pumped out from CSF into the circulation.

Absorption, metabolism, and excretion of [14C]imidafenacin, a new compound for treatment of overactive bladder, after oral administration to healthy male subjects

The absorption, metabolism, and excretion of imidafenacin [KRP-197/ONO-8025, 4-(2-methyl-1H-imidazol-1-yl)-2,2-diphenylbutanamide], a new antimuscarinic drug developed for treatment of overactive bladder, were assessed in six healthy male subjects after a single oral administration of 0.25 mg of [14C]imidafenacin (approximately 46 μCi). The highest radioactivity in the plasma was observed at 1.5 h after administration. The apparent terminal elimination half-life of the total radioactivity was 72 h. Approximately 65.6 and 29.4% of the administered radioactivity were recovered in the urine and feces, respectively, within 192 h after administration. The metabolite profiling by high-performance liquid chromatography-radiodetector and liquid chromatography/tandem mass spectrometry demonstrated that the main component of radioactivity was unchanged imidafenacin in the 2-h plasma. The N-glucuronide conjugate (M-9) was found as the major metabolite and the oxidized form of the 2-methylimidazole moiety (M-2) and the ring-cleavage form (M-4) were detected as the minor metabolites in the 2-h plasma, but M-4 was found to be the main component in the 12-h plasma. Unchanged imidafenacin, M-9, M-2, and other oxidized metabolites were excreted in the urine, but the unchanged imidafenacin and M-9 were not found in the feces. Two unique metabolites were found in the urine and feces, which were identified as the interchangeable cis- and trans-isomers of 4,5-dihydrodiol forms of the 2-methylimidazole moiety. These findings indicate that imidafenacin is rapidly and well absorbed (at least 65% of dose recovered in urine) after oral administration, circulates in human plasma as the unchanged form, its glucuronide, and other metabolites, and is then excreted in urine and feces as the oxidized metabolites of 2-methylimidazole moiety.

Kinetics of Quinolone Antibiotics in Rats: Efflux from Cerebrospinal Fluid to the Circulation

AbstractPurpose. An active transport system, which pumps quinolone antimicrobial agents (quinolones) from cerebrospinal fluid (CSF) to systemic blood, exists at the choroid plexus, an epithelial tissue that forms the blood-CSF barrier (BCSFB). The present study was carried out to clarify the contribution of this transport system to the disposition of quinolones in the central nervous system. Methods. Six quinolones were administered intracerebroventricularly to rats and their elimination from the CSF was examined. The inhibitory effect of probenecid and quinolones on the efflux of fleroxacin from the CSF was also examined. Probenecid or two types of quinolone (AM-1155, pefloxacin) were co-administered intracerebroventricularly with fleroxacin. Results. The elimination clearance from the CSF for norfloxacin, AM-1155, fleroxacin, ofloxacin, sparfloxacin and pefloxacin was 14, 22, 21, 20, 47 and 35 µl/min/rat, respectively. An approximately 3.5-fold difference was thus observed between norfloxacin and sparfloxacin. These values were 4- to 14-fold larger than the [l4C]mannitol clearance. Furthermore, the elimination clearance of quinolones from the CSF was 7- to 60-fold larger than the active efflux clearance at the BCSFB estimated from our previous in vitro data. Co-administration of AM-1155, pefloxacin and probenecid did not inhibit the elimination of fleroxacin from the CSF. Conclusions. The active transport system at the BCSFB plays only a small part in the elimination of quinolones from the CSF. Passive diffusion via the BCSFB and diffusion across the ependymal surface into brain extracellular fluid, followed by efflux across the blood-brain barrier, may be the predominant pathway for quinolone elimination from the CSF.