Combining high-throughput pharmacokinetic screens at the hits-to-leads stage of drug discovery.

Profound technological advances in the drug discovery process have led to the identification of an increasingly large number of promising compounds at the hits-to-leads stage. Higher-throughput pharmacokinetic screens have therefore been developed to enhance the tractability of selected leads and minimize the risk of failure in the later stages of drug development.

High-performance chromatographic assay for the sulphoxide metabolite of 2'-deoxy-3'-thiacytidine in human urine.

A high-performance liquid chromatographic method is described for the determination in human urine of GI138870X, the sulphoxide metabolite of a novel dideoxynucleoside analogue, 2'-deoxy-3'-thiacytidine (lamivudine). GI138870X was extracted from human urine using Empore SDB RPS solid-phase extraction disks prior to reversed-phase chromatography with UV detection. The method has shown to be valid over the concentration range 0.5-100 micrograms/ml using a 0.5-ml sample volume.

Pharmacokinetics of lamivudine and BCH-189 in plasma and cerebrospinal fluid of nonhuman primates.

2'-Deoxy-3'-thiacytidine is a dideoxycytidine analog with a sulfur in place of the 3' carbon of the ribose. There are two enantiomeric forms of the compound, both of which inhibit human immunodeficiency virus type 1 and 2 replication in vitro. However, the (-) enantiomer (lamivudine) appears to be significantly less cytotoxic to uninfected lymphocytes than is the (+) enantiomer. Lamivudine has entered initial clinical trials, and the present study was designed to describe the pharmacokinetic behavior of this compound in both plasma and cerebrospinal fluid (CSF) of primates. Lamivudine was administered as an intravenous bolus dose of 20 mg/kg for 24 h for administration. Urine samples were also obtained from two animals. The same dose of the racemate (BCH-189) was administered to one animal. The drug was quantitated in CSF and plasma with a reverse-phase high-pressure liquid chromatography technique. Elimination of lamivudine from plasma was biexponential, with a mean alpha phase half-life of 5.4 min, a mean beta phase half-life of 84 min, and a total clearance of 6.1 liters/h. The total clearance of the same dose of BCH-189 in a single animal was 11.0 liters/h. In two animals from which urine was obtained for 12 h postadministration, 32 and 59% of the drug was recovered unchanged. The deamination product of lamivudine was not detected. The CSF/plasma ratio of lamivudine was significantly higher when the drug was measured in the lumbar CSF (mean, 0.41) than when it was measured in the ventricular CSF (mean, 0.079). The measured CSF/plasma ratio for ventricular CSF is equivalent to that of other dideoxycytidine analogs, confirming the importance of the nucleobase in determining the degree of CSF penetration. The difference in lamivudine exposure in ventricular and lumbar CSF suggests that there is a transport mechanism for efflux of cytidine analogs from ventricular CSF.

Automated high-performance liquid chromatographic method for the determination of a neuraminidase inhibitor (GG167) in human serum by pre-column fluorescence derivatisation using benzoin.

A pre-column fluorescence derivatisation high-performance liquid chromatographic method for the analysis of a neuraminidase inhibitor, GG167, in human serum is described. GG167 was extracted from serum samples using Bond Elut SCX solid-phase extraction cartridges, followed by derivatisation with benzoin prior to reversed-phase chromatography with fluorescence detection. This method has been automated using a Zymark robot and used in the analysis of human serum samples from clinical studies. The method has been shown to be valid over a concentration range of 10-800 ng/ml using a 1-ml sample volume.

Interspecies scaling and pharmacokinetic parameters of 3TC in humans.

3TC is a dideoxy-nucleoside analogue that has demonstrated in-vitro activity against human immunodeficiency virus (HIV). 3TC concentrations in humans were predicted before the initiation of clinical trials by interspecies scaling of pharmacokinetic parameters observed in animal species. Clearance and volume of distribution were estimated for humans using linear regression on a log-log scale of each parameter versus body weight for rats and dogs. The concentration-time profile and the average serum concentration at steady state after various dosage regimens were estimated as a basis for initial dose selection for clinical trials. The predicted parameters (clearance of 16.3 L/hr and volume of distribution of 40 L for a 70-kg man) were compared with that observed (mean clearance of 24 L/hr and mean volume of distribution of 96 L, mean weight of 74 kg) in 20 asymptomatic, HIV positive, volunteers after single intravenous doses of 3TC. Interspecies scaling was applied prospectively as a rationale for dose selection of 3TC in clinical trials.

High-performance liquid chromatographic assay for 2'-deoxy-3'-thiacytidine in human serum.

A high-performance liquid chromatographic method for the determination of 2'-deoxy-3'-thiacytidine (3TC), a novel dideoxy-nucleoside analogue, in human serum is described. 3TC was extracted from serum samples using Bond Elut Certify solid-phase extraction cartridges prior to reversed-phase chromatography with UV detection. The method has been shown to be valid over the concentration range 10-5000 ng/ml using a 1-ml sample volume, both before and after heat treatment of the samples at 60 degrees C for 3 h. The method has been automated using a Zymark robot and used in the analysis of serum samples from HIV positive patients.

The safety and pharmacokinetics of a reverse transcriptase inhibitor, 3TC, in patients with HIV infection: a phase I study.

OBJECTIVE: To determine the safety and pharmacokinetics of the nucleoside analogue, 3TC. DESIGN: A Phase I, open-label, single-centre study. METHODS: Twenty asymptomatic, HIV-infected male patients with CD4 lymphocyte counts < 500 x 10(6)/l who had not received previous antiretroviral therapy completed the study. Each patient received a single intravenous dose followed by a single oral dose of 3TC. Four patients were dosed at each of five dose levels (0.25, 1.0, 2.0, 4.0 and 8.0 mg/kg). RESULTS: The most commonly reported adverse event was headache, which was generally reported to be mild. The mean bioavailability of 3TC was 82% following oral administration. The majority of the dose (approximately 70%) was excreted unchanged in the urine. CONCLUSIONS: Overall, 3TC was well tolerated following dosing, and there were no significant changes in the safety parameters measured. Phase I/II clinical trials with 3TC are ongoing to evaluate its safety, pharmacokinetics and preliminary activity.

Absorption, distribution and excretion of lacidipine, a dihydropyridine calcium antagonist, in rat and dog.

1. The absorption, distribution and excretion of lacidipine have been studied in rat and dog after i.v. (0.05 mg/kg for rat; 0.5 mg/kg for dog) and oral dosage (2.5 mg/kg for rat; 2.0 mg/kg for dog). 2. Lacidipine was rapidly and extensively absorbed after oral dosing, in both species. Oral bioavailability was up to 26% in rat and up to 32% in dog, due to extensive first-pass metabolism. 3. After oral administration, peak levels of radioactivity were reached at 4-8 h in rat and 1-2 h in dog. Unchanged lacidipine peaked at 1-2 h in both species. Plasma levels of radioactivity were higher in female rats than in males but there was no difference in levels of unchanged drug. 4. After i.v. dosing the terminal half-life of unchanged drug was 2.9 h in rat and 8.2 h in dog. The half-life of radioactivity in plasma was longer in both species. 5. After both routes of administration, radioactivity was rapidly distributed in rat tissues with the highest concentration in liver, fat and gastrointestinal tract. Only traces of radioactivity were detected in the CNS and in rat foetuses. 6. Extensive biliary elimination occurred, and most of the radioactivity (73-95%) was excreted in the faeces after i.v. or oral administration. 7. The compound was extensively metabolized, no significant amount of unchanged drug was excreted in bile or urine.