For the purpose of clarifying the pharmacokinetics of esmolol in rats, the plasma level, tissue distribution, metabolism and excretion were investigated after intravenous administration of ¹⁴C-esmolol, and the following results were obtained:
1. After intravenous administration to rats, the radioactivity in plasma decreased biphasically, and disappeared almost completely by 24 hr. The elimination half-lives were dose independent (10, 20, 40 mg/kg), and the plasma levels and AUC increased in proportion to the administered doses. The unchanged drug was not detected in plasma at 2 min after administration, but ASL-8123, the hydrolyzed metabolite of esmolol, was accounted for more than 80% of radioactivity in plasma. This indicated esmolol to be rapidly metabolized by esterases in blood. During repeated administration (for 7 days), the radioactivity in plasma showed no accumulation, and pharmacokinetic parameters after the final administration were similar to those after a single administration.
2. The radioactivity distributed rapidly and widely to tissues, and the levels were highest in kidney and lung, followed by adrenal gland, thyroid, pancreas, harderian gland, pituitary, liver and small intestine. The time course profiles in radioactivity levels in the heart, a target organ, were about 2 times higher than those in blood. In whole body autoradiograms, the transfer of radioactivity to the urine was already observed at 5 min after administration, indicating a rapid excretion of radioactivity to the urine.
3. Plasma protein binding of radioactivity in vitro was about 23%, and that in vivo (from 5 min to 8 hr after administration) accounted for 22-37%.
4. The main metabolite in plasma and urine was ASL-8123, which accounted for more than 80% of radioactivity in both samples. This indicates that enzymatic hydrolyses of esmolol in blood and the excretion of ASL-8123 into urine are rapid. Hydroxylated ASL-8123 was detected in minor quantities, but conjugates were not observed in either sample. No sex-related difference was observed in metabolite composition.
5. Radioactivity was completely excreted into feces or urine within a day, and urinary excretion (about 95% of dose) was the major elimination route. After repeated administration (for 7 days), the urine/feces ratio was not different from that after a single administration, and radioactivity was not remained. The biliary excretion of radioactivity was slight (less than 2% of dose) and the transfer into milk was observed, but the elimination of radioactivity in milk correlated with that in maternal plasma.

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Esmolol, an ultra-short-acting beta-adrenergic blocker, is a racemate. In this study, to compare the stereoselective pharmacokinetics of enantiomers, we have investigated the blood level profile after a single intravenous administration (10 mg/kg) of ¹⁴C-esmolol hydrochloride (¹⁴C-esmolol) to dogs. The distribution into the heart, a target organ, was tested after a single intravenous administration (20 mg/kg) of ¹⁴C-esmolol to rats. Each enantiomer was tested for metabolic conversion by blood esterase (in vitro), and the protein binding. In addition, the chiral inversion was investigated using human (in vitro) and dog blood (in vitro and in vivo).
1. The blood concentration and the AUC of d-esmolol were 1.6-fold higher than those of l-esmolol, and the half-life of d-esmolol was 1.7-fold longer than that of l-esmolol after administration of ¹⁴C-esmolol to dogs.
2. The half-life of d-esmolol was 1.4-fold longer than that of l-esmolol after incubation with dog blood, whereas no stereoselective difference was found in human blood.
3. The concentration in the heart showed no significant difference between two enantiomers after administration of ¹⁴C-esmolol to rats.
4. The chiral inversion was found neither in blood after administration of ¹⁴C-d-esmolol or ¹⁴C-l-esmolol to dogs nor in human and dog blood after the in vitro incubation.
5. There was no significant difference in protein bindings of each enantiomer after the in vitro incubation with human, dog and rat serum, and the bound fraction was the highest in human, followed by dog and rat serum.
In conclusion, these findings suggest that pharmacokinetics of enantiomers slightly differ in dog, whereas there are no stereoselective differences in human blood kinetics.

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In this study, the distribution and accumulation of SK-896 ([Leu¹³]motilin-Hse), a new human motilin analogue, were assessed after single or repeated intravenous administration of rats with³H-SK-896.
Following intravenous bolus injection of male rats with ³H-SK-896, the percentage of plasma immunoreactive radioactivity to plasma total radioactivity was 88.9% at 2 min, 50.9% at 15 min and 17.2% at 30 min, respectively. The highest radioactivity was found in the kidney at 2 min after administration, when most of the plasma total radioactivity was immunoreactive. The levels of radioactivity in the cerebrum, cerebellum, and testis were less than 3% of that in the plasma, and the radioactivity in the other tissues except for kidney were less than 30% of that in the plasma. These results suggested that little ³H-SK-896 and its immunoreactive metabolites transferred to the tissues except for kidney.
In female rats, the distribution profiles of the radioactivity after intravenous injection were the same as those of male rats. Also there was no localization of radioactivity in the genital organs, indicating that there was no sex difference in tissue distribution of ³H-SK-896 after intravenous injection to rats.
After multiple intravenous bolus injections (SK-896 was given once a day for 6 days and ³H-SK-896 was given on the 7th day) to male rats, the percentage of plasma immunoreactive radioactivity to plasma total radioactivity was 92.5% at 2 min, 50.6% at 15 min and 17.6% at 30 min, respectively, the same as with a single administration. There were no differences in the tissue distribution profiles of radioactivity between single and repeated intravenous bolus injections.

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SK-896 ([Leu¹³]motilin-Hse) is a new human motilin analogue synthesized by Escherichia coli using a biotechnological method. In this study, the pharmacokinetic properties of SK-896 after a single intravenous injection or during and after a single constant rate intravenous infusion of rats with ³H-SK-896 were assessed. After intravenous bolus injection of male and female rats with ³H-SK-896 at the dose of 4 μg/kg, the plasma levels of immunoreactive radioactivity declined bi-exponentially. No sex difference in pharmacokinetic properties was observed after the intravenous bolus injection. During constant rate intravenous infusion of male rats at the dose of 4μg/kg/h for 20 min, the plasma level of immunoreactive radioactivity increased rapidly, and declined bi-exponentially after the completion of infusion. No significant differences in pharmacokinetic parameters were observed between intravenous bolus injection and intravenous infusion, suggesting that there was essentially no change in the pharmacokinetics by the administration rate. Within 120 h after intravenous bolus injection to male and female rats at the dose of 4μg/kg, 10.9%, 9.55% and 3.84% of administered radioactivity were excreted into urine, feces and expired air in males, against 22.7%, 13.1% and 5.38% in female rats, respectively. Serum protein binding of ³H-SK-896 was 91.4-94.2% in dog, 93.6-96.6% in rat and 96.2-97.0% in human. Binding ratio of ³H-SK-896 to human serum albumin was 84.2-87.0%. Concentration-dependency of ³H-SK-896 binding to serum proteins was not observed over the concentration range studied.

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Diclofenac sodium (DF-Na) is a phenylacetate non-steroidal anti-inflammatory agent, and has a good anti-inflammatory analgesic antipyretic effect. In this study, in consideration of daily variations in the severity of rheumatoid arthritis and osteoarthritis and to improve patient compliance, we developed sustained release preparations that are expected to be effective by a single daily administration. To select the most optimal preparation for humans, a pharmacokinetic study was performed in dogs. We employed multiple units for control of the drug release rather than a single sustained release system in the design of a sustained release preparation, and prepared four sustained release preparations with different sustained release mechanisms.
1. A preparation coated with a combination of enteric polymer (PMA) and water-insoluble polymer (EC) to delay the solubilization and water permeation of the film compared to those in the PMA-coated control preparation.
2. A preparation in which the drug granules are coated with an enteric film (PMA) combined with an organic acid (fumaric acid) to delay the release of DF-Na.
3. A preparation containing sustained release granules coated with a combination of water-insoluble polymer (EC) and water-soluble polymer (PVP) to make the drug release time-dependent.
4. A preparation double-coated with a water-insoluble film (EC: PVP = 2 : 1) and an enteric film (PMA) to delay the release of DF-Na.
The four sustained release preparations with different mechanisms were selected based on the pharmacokinetic study in beagles for the next clinical study.

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Molecular genetics of cytochrome P450 enzymes (CYPs) in relation to the drug development and drug treatment are reviewed. CYPs play a key role in the metabolism of various drugs. The capacity of CYPs varies from one person to another, leading to variable rates of drug metabolism. This variability is mainly due to the genetic polymorphism of CYPs. A number of different functional alleles of different CYPs have been identified. CYP2D6, CYP2C19 and CYP2C9 are the three most well studied and best characterized. On the other hand, CYP3A4 is the most important isoform of CYPs which metabolizes more than half of substrate drugs of CYPs. Although CYP3A4 is known to show wide inter-individual variation, it has not been attributed to genetic factor but to rather induction and inhibition of this enzyme. However, up to now the existence of 12 mutant alleles have been reported for this enzyme and variable capacity of CYP3A4 is now recognized to be explainable in part by the genetic factors. In this article, CYP2D6, CYP2C19, CYP2C9 and CYP3A4 will be dealt with in some detail but other CYPs will be covered in outline.

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More than 30% of the cases of discontinuation of the clinical development of new drugs. Unfavorable drug metabolism/pharmacokinetic properties (DMPK) are probably responsible for. Since the ideal medicine is blessed with a good balance of pharmacological potency/selectivity and DMPK properties, a compound with an inappropriate DMPK will never become an innovative patient-oriented drug. The ability to predict human DMPK early in the drug discovery process is, therefore, key in reducing the attrition rate of compounds entering development. Moreover, by maximizing early DMPK studies, useful information and guidance for drug discovery programs can be provided to medicinal chemists and pharmacologists.
Dramatic advances in technologies such as combinatorial chemistry and high throughput screening have been revolutionizing the drug discovery process over the past decade. This paradigm shift is having a great impact on traditional DMPK research and is driving the challenge for high throughput DMPK research. In vitro DMPK studies that use a variety of human tissues allow the screening of a large number of compounds in a relatively short period of time. The use of LC-MS/MS also greatly contributes to the reduction of the time required for in vivo animal PK studies.
This paper will focus on the successful management and strategies in early DMPK research from the perspectives of the pharmaceutical industry.

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In the aftermath of the Human Genome Project, research on the medical usage of the informations obtained through this Project is progressing rapidly, globally and most prominently in the developed countries. The post-genomics research, which is expected to bring fruits in health promotion and prevention, diagnosis and treatment of various disorders, can also raise various ethical, legal and social problems. In this review, contents of the guideline on the conduct of research on human genome research, which was recently publicized from Japanese government, are briefly introduced.
Various ethical and social problems raised by a clinical application of gene diagnosis and gene therapy are also discussed.

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Cytochrome P450s included in CYP3A subfamily are involved in metabolism of numbers of clinically relevant drugs as well as numerous endogenous compounds. CYP3A forms are induced by treatment with various drugs, but clear differences are observed on induction profiles between experimental animals and humans. Recently, a novel orphan receptor, pregnane X receptor (PXR) has been isolated and reported to make a heterodimer with retinoid X recepterα (RXRα). In our experiment, co-expression of human PXR (hPXR) with RXRα did not show the CYP3A1 gene activation by treatment with rifampicin. When hPXR was co-expressed with other orphan receptors, COUP-TFs, apolipoprotein AI regulatory protein 1 (ARP-1) or v-ErbA-related protein 3, the CYP3A1 gene expression was activated by rifampicin in HepG2 cells. On the contrary, neither co-expression of PXR with RXRα nor with COUP-TFs showed the CYP3A4 gene activation by the treatment, although the activation was observed on the single expression of hPXR in HepG2 cells. On the other hand, both CYP3A1 and CYP3A4 inductions through hPXR activation by rifampicin are basically similar at the level of their cis-elements. The transactivation of the CYP3A1 gene by the drug is mediated through B site binding with hPXR and enhanced through C site. The transactivation of the CYP3A4 gene is mainly mediated through dNR-1 locating around at its -7600 bp and enhance through ER-6. B site and dNR-1 or C site and ER-6 conserve high homologies in those nucleotide sequences. To further verify the in vivo activation of the CYP3A4 gene expression by drugs, adenovirous reporter vector, AdCYP3A4-364, was constructed with a proximal promoter region ( -362 to + 11) of the CYP3A4 gene and injected into mouse. Induction profile of the CYP3A4 gene expression agreed well with that of the mouse CYP3A activity. The mutation study of receptor binding region (ER-6) diminished the induction of the CYP3A4 gene expression, but not the mouse CYP3A activity, testosterone 6β-hydroxylase. These results strongly suggest that the in vivo study of the gene transcriptional activation is very useful to assess the induction profile of the CYP3A4 gene expression by drugs.

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