Next Generation Copper Mediators for the Efficient Production of 18F‐Labeled Aromatics

Abstract Cu‐mediated radiofluorination is a versatile tool for the preparation of 18F‐labeled (hetero)aromatics. In this work, we systematically evaluated a series of complexes and identified several generally applicable mediators for highly efficient radiofluorination of aryl boronic and stannyl substrates. Utilization of these mediators in nBuOH/DMI or DMI significantly improved 18F‐labeling yields despite use of lower precursor amounts. Impressively, application of 2.5 μmol aryl boronic acids was sufficient to achieve 18F‐labeling yields of up to 75 %. The practicality of the novel mediators was demonstrated by efficient production of five PET‐tracers and transfer of the method to an automated radiosynthesis module. In addition, (S)‐3‐[18F]FPhe and 6‐[18F]FDOPA were prepared in activity yields of 23±1 % and 30±3 % using only 2.5 μmol of the corresponding boronic acid or trimethylstannyl precursor.


Introduction
Positron emission tomography (PET) is a non-invasive imaging technique that employs probes labeled with β + -emitting radionuclides to visualize biochemical processes for research or diagnostic purposes. The most widely used PET radionuclide is fluorine-18, which is readily accessible in the form of [ 18 18 F nuclear reaction. Fluorine-18 has advantageous decay properties (high β + -emission probability, relatively low β + energy, suitably long half-life of 110 min) compared to other common short-lived PET isotopes like 11 C, 13 N, 15 O or 68 Ga. [1] However, direct application of [ 18 F]F À in [ 18 O]H 2 O for radiofluorination is usually not feasible due to its high degree and strength of hydration. [2] As such, time consuming azeotropic drying steps and addition of bases and phase transfer catalysts are usually required to obtain highly nucleophilic [ 18 F]F À that can be further used for S n 2, S n Ar or transition metal-catalyzed reactions. [3] Owing to radioactive decay and inevitable heating-induced adsorption of [ 18 F]F À to the reactor walls, azeotropic drying and other pre-processing steps are invariably associated with radioactivity losses. However, these steps and the use of additives can be omitted, for example, by applying the "minimalist" [4] or related [5] protocols for S n 2, S n Ar or Cu-mediated radiofluorination.
Recently introduced approaches for Cu-mediated production of 18 F-labeled (hetero)arenes from different substrates, [6] including readily available aryl boronic acids, [7] pinacol boronates [8] and trialkylstannanes, [9] have granted access to emerging or established but otherwise hardly accessible radiotracers. Whereas the original procedures were rather inefficient and not well suited for radiotracer production on a preparative scale, several approaches to circumvent these limitations have since been proposed, [5b,10] which significantly contributed to the dramatic growth of PET imaging in the last 5-7 years. Main drawback of these procedures in their current form is that relatively large amounts of labeling precursor and Cu(Py) 4 (OTf) 2 (� 20 μmol of each) are often required to achieve high and reproducible radiochemical yields (RCYs), [11] especially when radiosyntheses are performed in remotely controlled synthesis units. This not only increases production costs, but in many cases also complicates purification of the resulting PET-tracers. Surprisingly, to date, no significant efforts have been made to find more efficient radiolabeling mediators, which could help to eliminate this bottleneck.
In this study, we first prepared a series of Cu(II) complexes and evaluated their efficiencies as mediators for radiofluorination of various model substrates bearing -B(OH) 2 or -Bpin groups (Scheme 1A). The most promising candidates were selected and used in subsequent experiments to optimize further reaction parameters like solvent, time and temperature (Scheme 1B). In addition, the developed radiofluorination protocol was adapted to radiolabeling of aryl trialkylstannanes (Scheme 1B) and to the use of low precursor amounts (Scheme 1C). Finally, we exemplified the suitability of the optimized protocol for the manual and automated preparation of several PET-tracers (Scheme 1D).

Results and Discussion
In order to assess the influence of different ligands on radiofluorination, we prepared a series of Cu(II) complexes with fifteen N-heterocycles and nine counter ions. For comparison, Co(II) perchlorate and Ni(II) triflate pyridine complexes were also synthesized. For three copper complexes, including the literature known Cu(Py) 4 (OTf) 2 (with known crystal structure) [12] as well as Cu(3,4-Me 2 Py) 4 (OTf) 2 and Cu(4-MeOPy) 4 (ClO 4 ) 2 , the crystal structures were determined (see Supporting Information for more details).
Although application of the alternative mediators significantly improved the radiolabeling yields in nBuOH/DMA, they remained moderate and did not exceed 58 %. Consequently, the influence of aprotic reaction solvents other than DMA was studied.
In the next step, the influence of reaction time and temperature on labeling efficacy was evaluated. Using Cu(4-PhPy) 4 (ClO 4 ) 2 as the Cu mediator, 4-Ph-Ph-B(OH) 2 was radiofluorinated in RCCs of 65-80 % over a broad range of reaction temperatures and times (5-20 min at 80-140°C; Figure S24A). At reaction temperatures of � 140°C, an increase in the variability of 18 F-incorporation was observed. Reaction times of less than 5 min at 110°C led to a drop of labeling efficacy (down to 57 � 4 % and 20 � 13 % for reaction times of three and  Tables S22 and S23. one min, respectively; Figure S24B), while RCCs of 54 � 2 % and 37 � 7 % were still observed after 10 min at lower temperatures of 70 and 60°C, respectively. A further increase of the incubation time at these reaction temperatures to 30 min led to RCCs of 82 � 1 % and 66 � 8 %, respectively ( Figure S24C).

Conclusion
In conclusion, our screening study led to the discovery of several highly efficient mediators for Cu-mediated radiofluorination like Cu(4-PhPy) 4 (ClO 4 ) 2 , Cu(3,4-Me 2 Py) 4 (OTf) 2 and Cu(3,4-Me 2 Py) 4 (ClO 4 ) 2 . Especially in DMI or nBuOH/DMI as reaction media, these copper complexes enabled highly efficient 18 Flabeling of different boronic and stannyl substrates. The optimized protocol worked well at reaction temperatures in the range of 60-90°C, indicating its suitability for radiofluorination of thermolabile substrates. The practicality of the method was demonstrated by preparation of several PET-tracers, including 6-[ 18 F]FDOPA, in improved radiochemical yields and/or using significantly lower precursor loadings. Furthermore, the procedure was successfully implemented into a remote-controlled synthesis unit, highlighting its applicability for GMP-compliant production of clinically relevant PET-tracers.

Experimental Section
Chemistry: Detailed information on the synthesis of metal complexes and radiolabeling precursors as well as the corresponding analytical data ( 1 H, 13  Optimized procedure for radiofluorination of boranyl precursors: [ 18 F]F À (500 μL, 20-5000 MBq) was loaded onto a QMA cartridge and eluted with a solution of Et 4 NOTf (1 mg, 3.6 μmol) in nBuOH (400 μL) into a solution of the respective Cu complex and precursor (10 μmol of each or 10 μmol of mediator and 2.5 μmol of boranyl substrate) in DMI (800 μL). The reaction mixture was heated at 110°C for 10 min in atmospheric or synthetic air, cooled to ambient temperature and diluted with H 2 O (2 mL). RCCs were determined by radio-HPLC. PET-tracers were isolated by preparative HPLC and formulated as ready-to-use solutions. At high atmospheric humidity (e. g., during the midsummer), a significant drop of the RCCs was sometimes observed, presumably owing to the hygroscopicity of Et 4 NOTf. In such cases, [ 18 F]F À should preferably be eluted with a solution of Et 4 NOTf in MeOH. MeOH was removed at 60°C for 2-3 min under reduced pressure in a stream of argon and the residue was taken up into a solution of the respective Cu mediator and precursor (10 μmol of each if not otherwise noted) in a mixture of the corresponding solvent and nBuOH (1200 μL; 2 : 1).
Optimized procedure for radiofluorination of stannyl precursors: [ 18 F]F À was loaded onto a QMA carbonate cartridge and eluted with Et 4 NOTf (1 mg, 3.6 μmol) in MeOH (500 μL) into a V-Vial as described above, followed by evaporation of MeOH at 60°C under reduced pressure in a stream of argon. The V-Vial was filled with argon, sealed with a silicon septum and a solution of the appropriate precursor and copper complex (10 μmol of each if not otherwise noted) in DMI (800 μL) was added via a cannula through the septum. The reaction mixture was heated at 90 or 110°C for 10 min, cooled to ambient temperature and diluted with H 2 O (2 mL). RCCs were determined by radio-HPLC. PET-tracers were isolated by preparative HPLC and formulated as ready-to-use solutions.