Radiolabel, preparation

Cl4-DBpD) may occur in trace amounts in the herbicide, 2,4,5-trichlorophenoxyacetic acid (2,4,5-T) (I, 2). Radiolabeled preparations of this dioxin are needed to facilitate studies of its degradation chemistry, metabolism, and mode of action. 

Recently, this method was adapted to label two commercially available liposomal formulations doxorubicin encapsulated in polyethylene glycol (PEG)-coated liposomes (Caelyx /Doxil ) (14) and daunorubicin encapsulated in small distearoyl-phosphatidyl-choline/cholesterol liposomes (Daunoxome ) (15). Although no DTPA was encapsulated in these liposomes, the labeling efficiency was typically between 70% and 80% and the radiolabeled preparations were stable in vivo during the time course of the experiment (four hours). Most likely, the lipophilic In-oxine avidly associates with the lipid bilayer and encapsulation of DTPA might not be necessary when the experimental observation period does not exceed four to six hours. 

Maksymowych and Simpson (1998) used transwell culture systems with various transformed epithelial cell lines to evaluate the fate of the HA components of type A progenitor toxin complex. Radiolabeled preparations of both BoNT/A and HA were taken up by cultured T-84 human colon carcinoma cells by bulk endocytosis. However, efficient delivery across the T-84 cells was only observed for the neurotoxin. 

Reports in the literature have proposed various animal models for preclinical studies of therapeutic radiopharmaceuticals in the laboratory, all of which pose serious limitations for investigations. In the present case, the efficacy of radiolabelled preparations was demonstrated in other models that more closely approximate the disease in humans. In this respect, it was envisaged that animals with spontaneous tumours could provide useful populations of pathological animal models for testing the new agents while also allowing their therapeutic efficacy and toxicity to be examined. 

The labelled compounds were purified on a SepPak CIS cartridge. The radiolabelled DOTATATE preparations were eluted with methanol, with greater than 9S% purity after evaporation of the methanol. To determine its stability, the radiolabelled preparation was stored at room temperature and at 37°C for 72 h for Lu. The stability of the preparation, described as the radiochemical purity, was determined by SepPak separation. 

The paper describes the procedure for labelhng the DOTATATE conjugate with and Lu, as well as the methods used for quality control. Parameters such as stability of the radiolabelled preparation, serum stability and protein binding were assessed by internalization and competition assays. The influence of chemical contaminants on the labelling yield of DOTATATE was investigated. Results of the first appUca-... 

The radiolabelled preparations were stored at room temperature in 0.4M sodium acetate or 0.3M ascorbic acid solution up to 72 h post-labelling for Lu-DOTATATE and up to 36 h post-labelling for " Y-DOTATATE, The influence of the radioactive concentration of the stored preparation on its stability was investigated using solutions of various radioactive concentrations 1.0, 20 and 50 mCi/mL for Y-DOTATATE. and 1.0, 15 and 35 mCi/mL for Lu-DOTATATE. The stability of the preparations (described as the radiochemical purity) was determined by HPLC and SepPak separation. 

For each radiolabelled preparation, after the radiochemical purity was ascertained, 5 gL of the preparation was added to a 50 gL portion of aliquoted human serum. Tlie mixture was then incubated at 37°C up to 24 h for... 

For the competition assays, AR42J cells were seeded at a density about 800 000 cells per well on six-well plates and incubated for 24-48 h. Cells were then washed twice with RPMI-1640 medium supplemented with 10% FCS and antibiotics. The cells were supplied with fresh medium, and the radiolabelled preparation, corresponding roughly to 200 fmol peptide in 150 gL PBS/0.5% BSA buffer, was added. The competitor (Sandostatin) was added in ten concentrations (0.01, 0.1, 1.0, 3.0,10, 30,100, 300,1 000,10 OOOnM) in a total volume of 150 gL of PBS/0.5% BSA buffer. The cells were incubated for 2 h at... 

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