Structure-activity relationship and pharmacokinetic studies of sotrastaurin (AEB071), a promising novel medicine for prevention of graft rejection and treatment of psoriasis.

Protein kinase C (PKC) isotypes have emerged as key targets for the blockade of early T-cell activation. Herein, we report on the structure-activity relationship and the detailed physicochemical and in vivo pharmacokinetic properties of sotrastaurin (AEB071, 1), a novel maleimide-based PKC inhibitor currently in phase II clinical trials. Most notably, the preferred uptake of sotrastaurin into lymphoid tissues is an important feature, which is likely to contribute to its in vivo efficacy.

Pharmacokinetics, distribution, metabolism, and excretion of deferasirox and its iron complex in rats.

Deferasirox (Exjade, ICL670, CGP72670) is an iron-chelating drug for p.o. treatment of transfusional iron overload in patients with beta-thalassemia or sickle cell disease. The pharmacokinetics and disposition of deferasirox were investigated in rats. The animals received single intravenous (10 mg/kg) or p.o. (10 or 100 mg/kg) doses of 14C-radiolabeled deferasirox. Biological samples were analyzed for radioactivity (liquid scintillation counting, quantitative whole-body autoradioluminography), for deferasirox and its iron complex [high-performance liquid chromatography (HPLC)/UV], and for metabolites (HPLC with radiodetection, liquid chromatography/mass spectrometry, 1H and 13C NMR, and two-dimensional NMR techniques). At least 75% of p.o.-dosed deferasirox was absorbed. The p.o. bioavailability was 26% at the 10 mg/kg dose and showed an overproportional increase at the 100 mg/kg dose, probably because of saturation of elimination processes. Deferasirox-related radioactivity was distributed mainly to blood, excretory organs, and gastrointestinal tract. Enterohepatic recirculation of deferasirox was observed. No retention occurred in any tissue. The placental barrier was passed to a low extent. Approximately 3% of the dose was transferred into the breast milk. Excretion of deferasirox and metabolites was rapid and complete within 7 days. Key clearance processes were hepatic metabolism and biliary elimination via multidrug resistance protein 2. Deferasirox, iron complex, and metabolites were excreted largely via bile and feces (total > or = 90%). Metabolism included glucuronidation at the carboxylate group (acyl glucuronide M3) and at phenolic hydroxy groups, as well as, to a lower degree, cytochrome P450-catalyzed hydroxylations. Two hydroxylated metabolites (M1 and M2) were administered to rats and were shown not to contribute substantially to iron elimination in vivo.

Biodistribution and plasma protein binding of zoledronic acid.

The bisphosphonate zoledronic acid is a potent inhibitor of osteoclast-mediated bone resorption. To investigate drug biodistribution and elimination, (14)C-zoledronic acid was administered intravenously to rats and dogs in single or multiple doses and assessed for its in vitro blood distribution and plasma protein binding in rat, dog, and human. Drug exposure in plasma, bones, and noncalcified tissues was investigated up to 240 days in rats and 96 h in dogs using radiometry after dissection. Drug biodistribution in the rat and within selected bones from dog was assessed by autoradiography. Concentrations of radioactivity showed a rapid decline in plasma and noncalcified tissue but only a slow decline in bone, to approximately 50% of peak at 240 days post dose, whereas the terminal half-lives (50-200 days) were similar in bone and noncalcified tissues, suggesting redistribution of drug from the former rather than prolonged retention in the latter. Uptake was highest in cancellous bone and axial skeleton. At 96 h after dose, the fraction of dose excreted was 36% in rat and 60% in dog; 94 to 96% of the excreted radioactivity was found in urine. Blood/plasma concentration ratios were 0.52 to 0.59, and plasma protein binding of zoledronic acid was moderate to low in all species. The results suggest that a fraction of zoledronic acid is reversibly taken up by the skeleton, the elimination of drug is mainly by renal excretion, and the disposition in blood and noncalcified tissue is governed by extensive uptake into and slow release from bone.

Pharmacokinetic investigation of a 14C-labelled beta 3/alpha tetrapeptide in rats.

The solid-phase synthesis and an ADME investigation with albino and pigmented male rats of the doubly 14C-labelled beta/alpha-tetrapeptide derivative Ac-beta3 hTyr-(D)Trp-beta3 hLys-beta3 hThr-lactone (3; Fig. 3) are described. After intravenous (i.v.) and peroral (p.o.) administration of the peptide, its concentration in blood and plasma, its tissue distribution, and the metabolism and the excretion of the peptide were analyzed over a period of up to 7 days post dose. The tetrapeptide in its ring opened form, 5, has a bioavailability of ca. 25%; radioactivity is distributed in the animals in an organ-specific way, and the compound appears to pass the blood-brain barrier to a very small extent, if at all (Tables 1-3 and Figs. 2-6). Excretion (37% renal, 44% fecal, including biliary) of the tetrapeptide 4 days after i.v. administration is almost complete, with only 4.3% remaining in the carcass; 4 days after p.o. administration 97% of the dose has been excreted in the feces. Radiochromatograms taken of plasma (0.5 and 24 h after i.v. dosing) and of urine and feces extracts (0-48 h collected) reveal the presence of lactone 3 and/or the corresponding hydroxy acid 5 with essentially no or very minor other peaks, respectively, representing possible metabolites (Tables 4-6, and Fig. 7 and 8). A comparison with a previous ADME investigation of a beta-nonapeptide show that--except for the lack of metabolism--all aspects of exposure, distribution, and elimination are different (structure-specific properties). The investigated tetrapeptide 3 is a potent and highly specific agonist of the somatostatin receptor hsst4, rendering the results described herein promising for diagnostic and therapeutic applications of beta-peptides.

Development and validation of a highly sensitive RIA for zoledronic acid, a new potent heterocyclic bisphosphonate, in human serum, plasma and urine.

Zoledronic acid is a new, highly potent bisphosphonate drug under clinical evaluation. A radioimmunoassay has been developed to determine zoledronic acid concentration in human serum, plasma, and urine. The assay utilizes rabbit polyclonal antisera against a zoledronic acid-BSA conjugate and a [125I]zoledronic acid derivative as tracer in a competitive format adapted to microtiter plates. The assay shows a LLOQ 0.4 ng/ml in serum or plasma (interassay%CV=17%, accuracy 97%), 5 ng/ml in urine (21%, 98%). In 23 patients receiving 4, 8 or 16 mg of zoledronic acid, drug concentrations in plasma were dose proportional and showed a multiphasic profile, followed by a prolonged gradual decline to concentrations near the LLOQ. Zoledronic acid disposition in plasma and the recovery of only 40-50% of the dose in urine are consistent with the rapid and extensive uptake by and slow release from bone in parallel with renal clearance, typically shown by bisphosphonates.

The outstanding metabolic stability of a 14C-labeled beta-nonapeptide in rats--in vitro and in vivo pharmacokinetic studies.

IN VITRO STUDIES: In CaCo-2 cell monolayers the beta-nonapeptide H(beta-HAla-beta-HLys-beta-HPhe)(3)-OH.4HCl (1), (14)C-labeled on both C atoms of the CH(2)-CO moiety of the central beta-HPhe residue, showed a low intrinsic permeability (<1%) and is subject to a prominent efflux system. The beta-peptide (1) binds to human and rat plasma protein in vitro independent of the concentration of 1 and of the species (30-36% bound fraction at 50, 500, and 5000 ng/ml), and has only low affinity for the corresponding blood cells (less than 5% of compound 1 in blood cells). IN VIVO STUDIES: The in vivo pharmacokinetic characteristics after i.v. administration of 5 mg/kg (to male rats and to bile-duct-operated rats) were: (i) negligible in vivo biotransformation of 1 (in urine, plasma and feces unchanged 1 represented virtually the only compound-related molecule); (ii) rapid initial decline (0-8 h post dose) of levels of compound 1 in blood and plasma followed by a slower decline (8-96 h post dose); (iii) in non-operated animals after 96 h only 38% of the dose was excreted and after 168 h 49% of the dose was found remaining in the carcass; elimination through the intestine wall represented the major elimination pathway in non-operated animals while in bile-duct-cannulated animals biliary excretion was not found to contribute substantially to elimination (iv) quantitative whole-body autoradioluminography (QWBAL) investigations revealed that the kidney was by far the most important target organ of distribution; other tissues with high concentrations of compound-related radioactivity were cartilage, lymph nodes, and liver, whereas lowest levels were found in white fat and in the brain. After p.o. administration (10 mg/kg) negligible radioactivity was observed in the systemic circulation, indicating negligible absorption; essentially the entire oral dose was recovered unchanged in feces collected over a period of 96 h.