Preparation of [66Ga]bleomycin complex as a possible PET radiopharmaceutical

Several radiolabeled bleomycin derivatives have been developed for imaging and/or therapy of neoplastic tissues. The most important imaging compounds contain indium-111, cobalt- 57, and rhodium-105. In our previous studies we have prepared radiolabeled bleomycin complexes such as gallium-67 as therapeutic and/or imaging agents. Our recent studies on the preparation and tumour imaging properties of [67Ga]bleomycin in normal and tumour-bearing mice showed a good tumour/blood and tumour/muscle ratio suggesting that it is an appropriate diagnostic agent. Due to the interesting properties and increasing importance of positron emission tomography, we investigated the possibility of incorporating 66Ga as a positron emitter with an antineoplastic compound, bleomycin, for use in tumour imaging. We optimized 66Ga complex formation conditions with bleomycin, to develop [66Ga]BLM. We hereby report the production of 66Ga, preparation, optimization, stability, and formulation studies of [66Ga]-bleomycin complex. Targetry: An electroplated 66Zn target on a copper backing plate was irradiated at an angle of 6 degrees toward the proton beam in order to achieve higher production yield. The target was cooled by a flow of 18 deg. C distilled water with a rate of 50 Lit/min. Chemical Separation: The irradiated target was dissolved by 10 N HCl (15 ml, H2O2 added) and the solution was passed through a cation exchange resin (Dowex 50 Wx8, H+ form) (h:10 cm, diameter:1 cm). The resulting high-purity [66Ga]GaCl3 solution was used directly in the labeling step. Labeling of bleomycin with [66Ga]GaCl3: [66Ga]GaCl3 (0.25-2.5 mCi) dissolved in acidic medium obtained above (0.5-2 ml) was transferred to a 2 ml-vial and pH was adjusted to various pHs (1-7) using 1M HCl and/or 1M NaOH. The mixture was evaporated by slight warming under a nitrogen flow. A mixture of BLM (0.25-2.5 mg) in normal saline (0.1 ml) was then added and was heated at different temperatures (25, 50, 80, and 100 deg. C) and was cooled in an ice bath, and rapidly sent for use. The active solution was checked for radiochemical purity by polymer-backed silica gel layer chromatography using a 1:1 mixture of 10% ammonium acetate and methanol as mobile phase. The final solution was then passed through a 0.22 μm filter and pH was adjusted to 5-7 by the addition of 1 M sodium acetate buffer. Radionuclide purity: The gamma spectroscopy of the final sample was carried out by HPGe detector, and showed a radionuclidic purity higher than 99 % showing the presence of 511, 834, and 1039 keV gamma energies, all of which are resulted from 66Ga. [66Ga]BLM complex in final product: Stability studies were based on the previous studies performed for other radiolabeled bleomycins (5). A sample of [66Ga]BLM (0.5 mCi) was kept at room temperature for 5 hrs while checked by RTLC every half an hour. A micropipet sample (50 μl) was taken from the shaking mixture and the ratio of free radiogallium to [66Ga]BLM was checked by radio thin layer chromatography (eluent: 10% NH4OAc buffer and methanol (1:1). Total labeling and formulation of [66Ga]BLM took about 60 min, with a yield of 97%. A suitable specific activity product was formed via insertion of [66Ga]gallium cation. No unlabelled and/or labeled by-products were observed upon RTLC analysis of the final preparations. The radio-labeled complex was stable in aqueous solutions for at least 24 h and no significant amount of other radioactive species were detected by HPLC 24 h after labeling. Trace amounts of [66Ga]gallium chloride (∼2%) were detected by TLC. RTLC showed that radiochemical purity of the [66Ga]labeled components was higher than 98%. In contrast to other labeled bleomycins, [66Ga]bleomycin, is a PET radiotracer with a rather long half life, and the high chemical stability of this radiopharmaceutical makes it a very suitable diagnostic agent