Abstract
242274
Introduction: 18F-labeled amino acid analogues are important Positron Emission Tomography (PET) tracers for early diagnosis and treatment follow up of many diseases, including cancer. Designing an optimized 18F-radiosynthesis method, including the proper prosthetic group and solvents for radiofluorination of precursor, purification mechanism, and choosing suitable deprotection process1 can improve the translational potential and subsequent utilization of radiotracers that might be otherwise disregarded due to cumbersome methods and exotic manipulations. For this study, we used CDP31, a small molecule inhibitor of ASCT2, to demonstrate a simple universal reduction method of commonly used ester and amine protecting groups in 18F-radiolabeling that effectively increases radiochemical yield and ease of handling, and advances adaptation to facile automation.
Methods: A prior 2-pot radiosynthesis of [18F]CDP31 with deprotection via TFA/anisole solution was performed on a GE TRACERlabTM module in which the azeotropically dried K222/[18F]KF complex was labelled to the tosylated CDP 31 precursor (7 – 12 mg) via nucleophilic substitution in DMSO at 160 °C for 20 minutes. The reaction mixture was injected into the HPLC for purification and the collected HPLC intermediate was passed through a pre-conditioned Oasis HLB light cartridge. The retained radiolabeled intermediate was rinsed with water and eluted with 1.0 mL of absolute ethanol to a second reactor and was dried under vacuum and nitrogen gas flow. The removal of the carboxylic t-Bu and N-Boc amines group after TFA/anisole mixture (1.0 mL/ 10 µL) was added to the dried residue and was heated at 60 °C for 5 minutes for hydrolysis2. After deprotection, the volatiles underwent a drying cycle using ethanol to remove the acidic volatiles. The dried activity was reconstituted in PBS buffer (pH 7.4). (Fig. 1a Reaction Scheme). For the new 1-pot [18F]CDP31 synthesis, the initial radiolabeling was similar, but diverged at the deprotection, where 1mL of aqueous 20% H2SO4 solution was added to the radiolabeled intermediate. Carboxylic t-Bu and N-Boc deprotection1 proceeded at 110 oC for 10 minutes followed by HPLC purification. The HPLC collection was passed through a tC18 SepPak cartridge and the retained [18F]CDP31 product was eluted with ethanol and buffered with PBS solution (pH 7). (Fig. 1b Reaction Scheme).
Results: The synthesis of [18F]CDP31 by sulfuric acid based deprotection took less than 60 minutes with non-decay corrected radiochemical yields greater than 25% (n=2), representing a major improvement from the original 2-pots synthesis that had averaged a radiochemical yield of less than 10% (n=3) and requires more than 85 minutes of production time. The radiochemical purity of the [18F]CDP31 increased to greater than 97%, up from the average of 92% (n=3) from the original synthesis method. Preliminary stability test of the [18F]CDP31 product by the new method indicated no radiolysis at 3 hours.
Conclusions: Our 1-pot deprotection method of the lipophilic [18F]CDP31 radiotracer using the GE TracerLab automated synthesizer has resulted in higher yield, more straightforward, and less time consuming than previously multi-pots synthesis methods. We envision that this process may serve as a universal strategy for preparing 18F-labeled lipophilic amino acid analogues, in which the precursors are protected with t-Bu, N-Boc, Tmob3, and tritylamines4.
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