In vivo Distribution
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II.M.1 II.M.2
Quantitative Whole Body Autoradiography (QWBA) . . . . . 587 Quantitative Tissue Distribution (QTD) . . . . . . . . . . . . . . . . . . . . . . . . . . 590
II.M.1 Quantitative Whole Body Autoradiography (QWBA) PURPOSE AND RATIONALE
Distribution studies with radiolabeled test substances in animals are an important part of the drug development process. Traditional routine methods used for these studies are quantitative tissue distribution studies (QTD) and alternatively whole-body autoradiography (WBA) with detection of the radioactivity in wholebody sections on X-ray film (John R. J. Baker 1989). WBA is a qualitative detection method with a very high local resolution which includes all organs an many small substructures. Radioluminography (RLG) is an alternative method of radiation detection based on the phosphorus imaging technique. RLG is much more sensitive than the WBA technique, its exposure time is much shorter and it has a much wider linear measure range. Because of the latter property RLG enables a quantification of drug concentrations in whole-body sections. Quantitative Whole-Body Autoradiography (QWBA) is based on the RLG technique and the use of standards obtained from dilution series containing known concentrations of radioactivity. Isotopes used in QWBA are mainly 14 C and 3 H. These standards were cut together with the whole-body sections to ensure an identical thickness and used for the internal calibration. The information of the calibration curve allows the determination of the concentrations in the organs and tissues of interest which can be derived from the measured area and the section thickness. The results of distribution studies form the principle basis for the assessment of exposure and the elimination of residues in human organs and tissues.
Direct determinations of exposure in man are generally limited to measurements in blood and plasma, which are easily accessible. Distribution patterns are determined in animals, usually rats, instead. Apart from the standard study design described below all kind of animals can be used up to the size of rabbits. The results obtained with QWBA provide pharmacokinetic data on the test substance and/or its metabolites, and evidence for interpretations regarding potential toxicological and pharmacological target organs. Additionally, before the first study with radiolabeled test substance in man can be started, a risk assessment of a human radiokinetic study is mandatory. The estimation of the radiation exposure in humans given a radiolabeled dose is based on exposure data obtained typically from QWBA studies in animals. PROCEDURE
Male pigmented and non-pigmented rats, weighing approx. 200 g, are used. At pre-selected time points (e.g. 0.25 h, 1 h, 2 h, 4 h, 8 h, 24 h, 72 h, 168 h) after administration of the radiolabeled test compound, the animals are sacrificed by CO2 -overdose, fixed on a piece of cardboard, embedded in sodiumcarboxymethylcelulosis and immediately deep-frozen using liquid nitrogen (–197 °C). In these frozen blocks a cer
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