Lectin-directed enzyme activated prodrug therapy (LEAPT): Synthesis and evaluation of rhamnose-capped prodrugs.

The lectin-directed enzyme activated prodrug therapy (LEAPT) bipartite drug delivery system utilizes glycosylated enzyme, localized according to its sugar pattern, and capped prodrugs released by that enzyme. In this way, the sugar coat of a synthetic enzyme determines the site of release of a given drug. Here, prodrugs of doxorubicin and 5-fluorouracil capped by the nonmammalian l-rhamnosyl sugar unit have been efficiently synthesized and evaluated for use in the LEAPT system. Both are stable in blood, released by synthetically d-galactosylated rhamnosidase enzyme, and do not inhibit the uptake of the synthetic enzyme to its liver target. These results are consistent with their proposed mode of action and efficacy in models of liver cancer, and confirm modular flexibility in the drugs that may be used in LEAPT.

Esophageal transit of the weekly film-coated risedronate (Actonel) placebo tablet in subjects with Kyphosis.

Risedronate sodium is a pyridinyl bisphosphonate of proven effectiveness for the treatment and prevention of osteoporosis and Paget's disease of the bone. The aim of this study was to compare the esophageal transit and gastric emptying of the placebo film-coated risedronate tablet when taken with 50 or 120 mL of water in subjects with Kyphosis. A total of 23 patients with radiologically documented osteoporosis participated in a single-center, open-label, crossover gamma scintigraphy study. The mean esophageal transit times were 15.6 s (50 mL) and 12.0 s (120 mL) and the mean gastric emptying half-times were 20.5 min (50 mL) and 14.3 min (120 mL). There was no relationship between the degree of Kyphosis measured from lateral standing radiographs and the esophageal transit time. This study demonstrated that even when taken with a minimal volume of water the esophageal transit and gastric emptying of the film-coated 35 mg weekly risedronate placebo tablet was similar in kyphotic subjects to previously obtained results from healthy control subjects.

Hydrophilic and lipophilic radiopharmaceuticals as tracers in pharmaceutical development: in vitro--in vivo studies.

BACKGROUND: Scintigraphic studies have been performed to assess the release, both in vitro and in vivo, of radiotracers from tablet formulations. Four different tracers with differing physicochemical characteristics have been evaluated to assess their suitability as models for drug delivery. METHODS: In-vitro disintegration and dissolution studies have been performed at pH 1, 4 and 7. In-vivo studies have been performed by scintigraphic imaging in healthy volunteers. Two hydrophilic tracers, (99mTc-DTPA) and (99mTc-MDP), and two lipophilic tracers, (99mTc-ECD) and (99mTc-MIBI), were used as drug models. RESULTS: Dissolution and disintegration profiles, differed depending on the drug model chosen. In vitro dissolution velocity constants indicated a probable retention of the radiotracer in the formulation. In vivo disintegration velocity constants showed important variability for each radiopharmaceutical. Pearson statistical test showed no correlation between in vitro drug release, and in vivo behaviour, for 99mTc-DTPA, 99mTc-ECD and 99mTc-MIBI. High correlation coefficients were found for 99mTc-MDP not only for in vitro dissolution and disintegration studies but also for in vivo scintigraphic studies. CONCLUSION: Scintigraphic studies have made a significant contribution to the development of drug delivery systems. It is essential, however, to choose the appropriate radiotracers as models of drug behaviour. This study has demonstrated significant differences in release patterns, depending on the model chosen. It is likely that each formulation would require the development of a specific model, rather than being able to use a generic drug model on the basis of its physicochemical characteristics.

Radionuclide imaging in drug development.

Radioactive tracers have made an immense contribution to the understanding of human physiology and pathology. At the start of the 21st century nuclear imaging has emerged as the main metabolic imaging modality which is of growing importance in drug development and clinical pharmacology. Using techniques adapted from those undertaken in clinical radiopharmacy and nuclear medicine facilities drug molecules and carrier systems may be radiolabelled and their release, biodistribution and uptake may be visualized in human subjects. Imaging studies are capable of locating the uptake of specific receptors in the brain, the site of disintegration of a tablet in the GI tract, the penetration of a nebulized solution into the lung and the residence time of an eye drop on the cornea. The technology uses suitable gamma emitting radionuclides such as 99mTc, 111In, 123I and 153Sm, which may be imaged with a gamma camera or positron emitters such as 11C, 13N, 15O and 18F for positron emission tomography (PET). Positron emitters are more appropriate for the direct labeling of drug molecules rather than metals such a 99mTc or 111In. A particular asset of these techniques is that the in vivo distribution and kinetics of a radiolabelled pharmaceutical formulation may be quantified. In this way correlation between the observed pharmacological effects and the precise site of delivery may be made. A powerful feature of nuclear molecular imaging is the evaluation of drug delivery systems in patient groups for whom the treatment is intended. Such studies not only provide data on the nature and characteristics of a product, such as reliability and reproducibility, but can demonstrate proof of principle for the new generation of targeted therapeutics. Imaging data are increasingly being used in product registration dossiers for submission to Regulatory Authorities.

LEAPT: lectin-directed enzyme-activated prodrug therapy.

Targeted drug delivery to selected sites allows reduced toxicity, enhanced efficiency and interchangeable target potential [Langer, R. (2001) Science 293, 58-59 and Molema, G. & Meijer, D. K. F., eds. (2001) Drug Targeting (Wiley-VCH, Weinheim, Germany)]. We describe a bipartite drug-delivery system that exploits (I) endogenous carbohydrate-to-lectin binding to localize glycosylated enzyme conjugates to specific, predetermined cell types followed by (II) administration of a prodrug activated by that predelivered enzyme at the desired site. The carbohydrate structure of an alpha-L-rhamnopyranosidase enzyme was specifically engineered through enzymatic deglycosylation and chemical reglycosylation. Combined in vivo and in vitro techniques (gamma scintigraphy, microautoradiography and confocal microscopy) determined organ and cellular localization and demonstrated successful activation of alpha-L-rhamnopyranoside prodrug. Ligand competition experiments revealed enhanced, specific localization by endocytosis and a strongly carbohydrate-dependent, 60-fold increase in selectivity toward target cell hepatocytes that generated a >30-fold increase (from 0.02 to 0.66 mg) in protein delivered. Furthermore, glycosylation engineering enhanced the serum-uptake rate and enzyme stability. This created enzyme activity (0.2 units in hepatocytes) for prodrug therapy, the target of which was switched simply by sugar-type alteration. The therapeutic effectiveness of lectin-directed enzyme-activated prodrug therapy was shown through the construction of the prodrug of doxorubicin, Rha-DOX, and its application to reduce tumor burden in a hepatocellular carcinoma (HepG2) disease model.

Usage of radiopharmaceuticals in the development of pharmaceutical drug delivery systems: validation of [99mTc]DTPA and [99mTc]ECD.

Tablets containing drugs of different lipophilicity, ranitidine and cinarizine, and placebo were prepared and their in vitro behaviour was studied by dissolution and disintegration tests. [(99m)Tc]Diethylenetriamine-pentaacetic acid ([(99m)Tc]DTPA) and [(99m)Tc]ethyl cysteinate dimer ([(99m)Tc]ECD) were used as tracers of the process. Both of them were added to tablets during wet granulation. Dissolution and disintegration profiles were assessed at different pH values (1, 4 and 7). Radioactivity was evaluated in filtered samples and scintigraphic studies were carried out in gamma camera. Stability in dissolution media was confirmed for both tracers under these conditions. Dissolution and disintegration velocity constants were calculated. [(99m)Tc]DTPA proved to be an appropriate tracer for polar drugs such as ranitidine. Nevertheless, it was not a suitable tracer for lipophilic active drugs such as cinarizine. On the other hand, the most lipophilic tracer, [(99m)Tc]ECD, exhibited the opposite behaviour. Scintigraphic studies of the disintegration process did not show significant differences between placebos and tablets containing active drugs. As disintegration is a physical process it does not discriminate between chemical differences in tablet formulations. Both methods complement each other because the dissolution process can be followed when a suitable radiotracer is chosen according to the physicochemical characteristics of the active drug.

Rhenium-188 and copper-67 radiopharmaceuticals for the treatment of bladder cancer.

The favourable nuclear properties of copper-67 and rhenium-188 for therapeutic application are described, together with methods for the chemical synthesis of a number of derivatives. Survival from invasive bladder cancer has changed little over the past 20 years. The intravesicular administration of Cu-67 or Re-188 radiopharmaceuticals in the treatment of bladder cancer offers some promise for improvement in this situation.

Radiopharmaceuticals for sentinel node detection

Imaging of sentinel nodes is a valuable technique in the management of some types of cancer. Several different radiopharmaceuticals have been successfully employed in the technique, but there have been few systematic studies to examine the relationship between physicochemical properties of the radiopharmaceuticals used and in-vivo performance. The present study describes the preparation and characterisation of radiocolloids of similar chemical composition but differing in particle size distribution in an attempt to identify the role of variations in the single parameter of size

Antibody conjugate radioimmunotherapy of superficial bladder cancer

The administration of antibody conjugates for cancer therapy is now proving to be of clinical value. We are currently undertaking a programme of clinical studies using the monoclonal antibody C595 (IgG3) which reacts with the MUC1 glycoprotein antigen that is aberrantly expressed in a high proportion of bladder tumours. Radioimmunoconjugates of the C595 antibody have been produced with high radiolabelling efficiency and immunoreactivity using Tc-99m and In-111 for diagnostic imaging, and disease staging and the cytotoxic radionuclides Cu-67 and Re-188 for therapy of superficial bladder cancer. A Phase I/II therapeutic trail involving the intravesical administration of antibody directly into the bladder has now begun

Evaluation of Pulsincap to provide regional delivery of dofetilide to the human GI tract.

Pulsincap formulations designed to deliver a dose of drug following a 5-h delay were prepared to evaluate the capability of the formulation to deliver dofetilide to the lower gastrointestinal (GI) tract. By the expected 5-h release time, the preparations were well dispersed throughout the GI tract, from stomach to colon. Plasma analysis permitted drug absorption to be determined as a function of GI tract site of release. Dofetilide is a well-absorbed drug, but showed a reduction in observed bioavailability when delivered from the Pulsincap formulations, particularly at more distal GI tract sites. Dispersion of the drug from the soluble excipient used in this prototype formulation relies on a passive diffusion mechanism and the relevance of this factor to the reduced extent and consistency of absorption from the colon is discussed. In these studies the effects of the degree of dispersion versus the site of dispersion could not be ascertained; nevertheless the scintigraphic analysis demonstrated good in vitro-in vivo correlation for time of release from Pulsincap preparations. The combination of scintigraphic and pharmacokinetic analysis permits identification of the site of drug release from the dosage form and pharmacokinetic parameters to be studied in man in a non-invasive manner.