An LC-MS-MS method for quantitative determination of maraviroc (UK-427,857) in human plasma, urine and cerebrospinal fluid.

Maraviroc is a first-in-class CCR5 antagonist that shows potent anti-HIV-1 activity in vitro and in vivo and is well tolerated in both healthy volunteers and HIV-1-infected patients. The method for determination of maraviroc (UK-427,857) and its major metabolite (UK-408,027) in human plasma consists of a protein-precipitation procedure and analysis by liquid chromatography/tandem mass spectrometry using positive ion TurboIonSpray® ionization and multiple reaction monitoring. The assay has been validated over a concentration range of 0.500-500 ng/mL for both analytes. The determinations of maraviroc in human cerebrospinal fluid (0.500-500 ng/mL) and in urine (5.00-5000 ng/mL) have also been validated but do not include measurement of the metabolite. The validations included extraction recovery, intra-assay and inter-assay precision and accuracy, stability of stock and spiking solutions, freeze-thaw stability, matrix stability, processed-extract stability, and evaluation of potential interferences from selected medications in plasma or urine.

Determination of a potent urokinase-type plasminogen activator, UK-356,202, in plasma at pg/mL levels using column-switching HPLC and fluorescence detection.

A rapid, sensitive and selective method using column-switching HPLC with fluorescence detection has been developed for the determination of UK-356,202, a potent urokinase-type plasminogen activator, in human plasma. A structural isomer of UK-356,202 is used as an internal standard. The lower limit of quantification is 20 pg/mL and the method is linear over a 100-fold concentration range. UK-356,202 is extracted from plasma simply through the removal of proteins by precipitation with acetonitrile. The HPLC system comprises three columns and the cycle time is 9.5 min per sample. The eluate from the extraction column is heart-cut onto a trace enrichment cartridge which is then back-flushed onto a narrow-bore Supelco ABZ+ Plus analytical column. The method has been used to analyze many thousands of samples from clinical and toxicological studies support. Its ruggedness is demonstrated by the use of a single extraction column for the analysis of over 1200 clinical samples.

Nonlinear oral pharmacokinetics of the alpha-antagonist 4-amino-5-(4-fluorophenyl)-6,7-dimethoxy-2-[4-(morpholinocarbonyl)-perhydro-1,4-diazepin-1-yl]quinoline in humans: use of preclinical data to rationalize clinical observations.

4-amino-5-(4-fluorophenyl)-6,7-dimethoxy-2-[4-(morpholinocarbonyl)-perhydro-1,4-diazepin-1-yl]quinoline (UK-294,315) is an antagonist of the human alpha1-adrenoceptor and exhibits nonlinear oral pharmacokinetics in humans. Superproportional increases in Cmax occur (220-fold, over a 1- to 50-mg dose range), area under the curve increases linearly, but time to maximum concentration decreases with dose, suggesting variation in rate but not extent of absorption. Oral absorption in humans is extensive, with only 14% of an orally administered (20 mg) radiolabeled dose excreted unchanged in the feces. In rats and dogs, UK-294,315 is partially eliminated as unchanged drug in feces (29 and 14% of an intravenous dose, respectively). Oral bioavailability is low in rats (11%) and high in dogs (71%), in keeping with systemic clearance. Fecal elimination of unchanged drug was 60% after oral administration to rats, indicating incomplete absorption in this species, whereas absorption in dogs is complete. UK-294,315 is a P-glycoprotein (P-gp) substrate (Km, 15 microM) exhibiting polarized flux in Caco-2 cell monolayers, saturable across a concentration range of 5 to 200 microM. Furthermore, the observations in vitro occurred at similar concentrations to those estimated in the gut lumen in clinical trials (dose range, 1-100 mg). It is considered that P-gp acts as a saturable absorption barrier to UK-294,315, slowing the rate of absorption at low doses, and is responsible for the observed nonlinearity in oral disposition in humans. Rat and dog pharmacokinetic studies offered limited insight into the process(es) driving nonlinear pharmacokinetics in humans. Our current understanding of the functional effects of P-gp in the human intestine, in combination with in vitro studies at clinically relevant concentrations, has helped rationalize the clinical data for UK-294,315.