Andreas Greischel

Concentration‐dependent plasma protein binding of the novel dipeptidyl peptidase 4 inhibitor BI 1356 due to saturable binding to its target in plasma of mice, rats and humans

Objectives The purpose of this study was to characterise the plasma protein binding of BI 1356.

Tissue distribution of the novel DPP-4 inhibitor BI 1356 is dominated by saturable binding to its target in rats.

BI 1356 (INN: linagliptin) is an inhibitor of dipeptidyl peptidase‐4 (DPP‐4). This study investigated whether saturable binding of BI 1356 to its target DPP‐4 occurs in tissues and whether drug accumulation occurs at these sites in vivo. In order to test these hypotheses, the tissue distribution of BI 1356 was determined in wild‐type and DPP‐4 deficient rats at different dose levels by means of whole body autoradiography and measurement of tissue radioactivity concentrations after single i.v. dosing of [14C]‐radio labeled BI 1356. The accumulation behavior of drug‐related radioactivity in tissues was further explored in an oral repeat dose study. Tissue levels of [14C]BI 1356 related radioactivity were markedly lower in all investigated tissues of the DPP‐4 deficient rats and the difference of the dose‐dependent increase of radioactivity tissue levels between both rat strains indicates that tissue distribution at low doses of BI 1356 is dominated by binding of BI 1356 to DPP‐4 in tissues. As the binding to DPP‐4 is strong but reversible, the tissue binding results in a long terminal half‐life in several tissues including plasma. The binding capacity to DPP‐4 is, however, limited. In the rat, saturation of DPP‐4 binding is suggested at an intravenous dose above 0.01–0.1 mg/kg [14C]BI 1356. As the DPP‐4 binding capacity is saturated already at low doses, accumulation of BI 1356 in tissues is unlikely, despite the long persistence of low amounts in the body. Copyright

Binding to dipeptidyl peptidase-4 determines the disposition of linagliptin (BI 1356)--investigations in DPP-4 deficient and wildtype rats.

Linagliptin (BI 1356) is a novel dipeptidyl peptidase‐4 (DPP‐4) inhibitor in clinical development for the treatment of type 2 diabetes. It exhibits non‐linear pharmacokinetics and shows concentration‐dependent plasma protein binding to its target, DPP‐4. The aim of this study was to investigate the impact of saturable binding of linagliptin to plasma and tissue DPP‐4 by comparing the pharmacokinetics of linagliptin in wildtype and DPP‐4 deficient Fischer rats using non‐compartmental and model‐based data analysis. The non‐compartmental analysis revealed a significantly reduced AUC in DPP‐4 deficient rats compared with wildtype rats when single intravenous doses ⩽1 mg/kg were administered, but the exposure was similar in both strains at higher doses. The terminal half‐lives were significantly shorter in DPP‐4 deficient rats compared with wildtype rats. For doses ⩽1 mg/kg, DPP‐4 deficient rats exhibited linear pharmacokinetics, whereas the pharmacokinetics of wildtype rats was non‐linear. In the model‐based analysis these differences could be accounted for by assuming concentration‐dependent protein binding in the central and one peripheral compartment in wildtype rats. In the model, disposition parameters for unbound linagliptin were assumed to be identical in both rat strains. Simulations with different doses of linagliptin and different concentrations of binding sites further illustrated that the interdependence of linagliptin and DPP‐4 in plasma and in the periphery has a major influence on the disposition of linagliptin in wildtype rats. In conclusion, the study showed that the concentration‐dependent binding of linagliptin to its target DPP‐4 has a major impact on the plasma pharmacokinetics of linagliptin. Copyright

Pharmacokinetics of the M1-agonist talsaclidine in mouse, rat, rabbit and monkey, and extrapolation to man

1. Talsaclidine is an M1-agonist under development for the treatment of Alzheimers disease. The aim of the study was to investigate the absorption, distribution, metabolism and excretion (ADME) of single intravenous and oral doses of [14C]-talsaclidine in mouse, rat, rabbit and monkey. Previous data in humans showed that the drug was mainly excreted into the urine as the unchanged parent drug. The hypothesis was tested if animal data of drugs, which are mainly excreted renally, could be extrapolated to human. 2. The apparent volume of distribution at steady-state (Vss) was comparable in all animal species (2-5 l.kg-1

The Dipeptidyl Peptidase-4 Inhibitor Linagliptin Exhibits Time- and Dose-Dependent Localization in Kidney, Liver, and Intestine after Intravenous Dosing: Results from High Resolution Autoradiography in Rats

Linagliptin is an orally active dipeptidyl peptidase-4 (DPP-4) inhibitor that is under development for the treatment of type 2 diabetes and shows dose-dependent pharmacokinetics in rats and humans. With microscopic autoradiography, the dose dependence of cellular distribution of [3H]linagliptin-related radioactivity was investigated in kidney at 3 h after intravenous injection of 7.4, 100, and 2000 μg/kg [3H]linagliptin. Furthermore, distribution of radioactivity in kidney, liver, and small intestine was investigated in relation to time (2 min, 3 h, and 192 h) after intravenous injection of 7.4 μg/kg [3H]linagliptin. The localization of radioactivity in the kidney at 3 h after administration of 7.4, 100, and 2000 μg/kg [3H]linagliptin changed with increasing dose from cortical glomeruli and parts of proximal tubule parts to parts of medullar proximal tubule. In addition, the compound distribution in the kidney shifted with time after administration of 7.4 μg/kg [3H]linagliptin from glomeruli (2 min) to the lower parts of proximal tubules (192 h). The radioactivity within proximal tubules was located primarily in the brush border. In the liver, the radioactivity persisted mainly around the portal triads and in the bile duct from 2 min to 192 h. In the small intestine, the radioactivity shifted from the lamina propria (2 min) to the surface of the villi and/or intestinal lumen (192 h). In conclusion, the cellular distribution pattern of [3H]linagliptin-related radioactivity reflected the known distribution of DPP-4. Together with the persistence of binding, this result supports the high relevance of DPP-4 binding of linagliptin for its pharmacokinetics and pharmacodynamics.

Altered drug disposition of the platelet activating factor antagonist apafant in mdr1a knockout mice

The aim of the present study was to determine a potential impact of P-glycoprotein (P-gp) on the tissue distribution and disposition of apafant (WEB 2086, CAS 105219-56-5), a selective platelet-activating factor antagonist, and on digoxin in mdr1a(-/-) and wildtype mice. Transport experiments in Caco-2 monolayers at low concentrations (<10 microM) showed that the secretory flux of [(14)C]apafant and [(3)H]digoxin exceeded the absorptive flux nine times. This efflux was concentration dependent and subject to inhibition by the P-gp substrates verapamil and cyclosporin A. This indicates that active drug transporter P-gp was involved in apafant and digoxin absorption. Mdr1a(-/-) mice showed a more than 70-fold higher concentration of digoxin-related radioactivity (P<0.001) in the brain than wildtype mice after intravenous doses of 0.05 mg/kg [(3)H]digoxin. Differences were less pronounced in other tissues. Both liquid scintillation counting and whole body autoradiography yielded comparable results and they also matched recently published data. Apafant-related radioactivity was about ten-fold higher in the brain of mdr1a(-/-) mice compared to wildtype mice following intravenous doses of 2 mg/kg radiolabelled apafant. Only slight or negligible differences were observed in other tissues. In wildtype mice, intestinal excretion of [(14)C]apafant (54.9%) exceeded biliary excretion (26.5%). However, in mdr1a(-/-) mice biliary excretion (50.7%) exceeded intestinal excretion (6.8%). These differences were mirrored in the urinary and faecal excretion. Pharmacokinetic parameters of apafant and radioactivity did not differ between wildtype and mdr1a(-/-) mice. The conclusions were: (1) apafant and digoxin are P-gp substrates, and (2) absence of mdr1a encoded P-gp significantly alters tissue distribution (especially in brain) and excretion routes (biliary and intestinal) of apafant.

Quantitation of the new hypoglycaemic agent AG-EE 388 ZW in human plasma by automated high-performance liquid chromatography with electrochemical detection

A sensitive and selective high-performance liquid chromatographic assay has been developed for a new hypoglycaemic agent AG-EE 388 ZW (I) in human plasma. Plasma samples containing I were acidified with 0.2 M hydrochloric acid, directly injected into C2 reversed-phase pre-columns, and cleaned up on-line. After pre-column switching, the substance was separated isocratically in 15 min on a C18 reversed-phase column. Quantitation was performed after amperometric detection by external standard calibration curves. The linearity of the assay was demonstrated over the therapeutic concentration range 5-200 ng/ml. between-day coefficient of variation was 9.2% at 30 ng/ml. The limit of detection was 5 ng/ml in plasma. Determination of human plasma samples after intravenous and oral administration of 1 mg of 14C-labelled I demonstrated the applicability of the assay for pharmacokinetic studies in humans.

Toxicity assessment of pramipexole in juvenile rhesus monkeys

Pramipexole (PPX) is a dopamine agonist approved for the treatment of the signs and symptoms of idiopathic Parkinsons disease as well as restless leg syndrome. The objective of this study was to investigate the toxicity of PPX when administered orally to juvenile rhesus monkeys once daily for 30 weeks, and to assess the reversibility of toxicity during a 12-week recovery. Rhesus monkeys (N=4 males and 4 females/group; 22-24 months of age) were orally treated daily for 30 weeks with 0.0, 0.1, 0.5 or 2.0 mg/kg PPX, and subjects were assessed daily using the NCTR Operant Test Battery (OTB). Clinical chemistry, hematology, ophthalmology and other standard postmortem toxicological evaluations, including histopathology and neuropathology as well as toxicokinetics were performed. The systemic exposure to PPX was higher than that at therapeutic doses in man and AUC(0-24 h)-data increased proportionally to dose. Blood pressure significantly decreased over time in all groups including control. Near the end of treatment, there were statistically significant decreases in heart rate for the 0.5 and 2.0 mg/kg/day groups compared to control. After 4 weeks of dosing, serum prolactin was significantly decreased in all treatment groups compared to control. This decrease remained at the end of treatment in the 0.5 and 2.0 mg/kg/day groups. In summary, administration of PPX at doses of up to 2.0 mg/kg/day for 30 weeks to juvenile rhesus monkeys produced adverse findings which were attributable to its pharmacological properties, including hypoprolactinemia.