A synthesis of 2-deoxy-2-{1sF)fluoro-~-glucose using accelerator-produced "F-fluoride ion generated in a water target

PHILIP A. BEELEY, WALTER A. SZAREK, GEORGE W . HAY, AND MILTON M . PERLMUTTER Carbohydrate Research Institute and Department of Chemistry, Queen's University, Kingston, Ont. , Canada K7L 3N6

Received March 13, 1984

PHILIP A. BEELEY, WALTER A. SZAREK, GEORGE W. HAY, and MILTON M. P E R L M U T T E R . ~ ~ ~ . J . Chem. 62, 2709 (1984). A synthesis of 2-deo~y-2-{~~F}fluoro-~-g~ucose (2-{IXF}DFG) from 1,2-anhydro-3,4: 5,6-di-0-isopropylidene-I-C-nitro-D-

mannitol has been developed. The procedure employed IXF produced by the 160(%e,p)'XF reaction using a water target. The label was introduced using KH{"F}F~ dried by microwave heating; the "F-labeled derivative, upon treatment with 80% trifluoroacetic acid, afforded 2-{'"}DFG in a radiochemical yield of 10%. The total time of the synthesis was less than 110 min.

PHILIP A BEELEY, WALTER A. SZAREK, GEORGE W. HAY et MILTON M. PERLMUTTER. Can. J. Chem. 62, 2709 (1984). On develop@ une methode de synthese du dCoxy-2 {1nF}fluoro-2 D - ~ ~ u c o S ~ ({IHF}-2 DFG) a partir de I'anhydro-1,2

di-0-isopropylidene-3,4:5,6 C-nitro-] D-mannitol. La mCthode implique I'utilisation de IXF produit par une reaction 16 0 ( 3 ~ e , p ) ' s ~ faisant appel a une cible aqueuse. On a introduit le marqueur en utilisant du KH{"F}FZ qui avait CtC sCchC par micro-ondes; la rCaction du dCrivC marque au "F avec de I'acide trifluoroacCtique a 80% fournit le {lXF}-2 DFG avec un rendement radiochimique de 10%. Le temps total de la synthese est inferieur 110 min.

[Traduit par le journal]

Introduction The use of 2-deoxy-2-{'8F)fluoro-~-glucose (2-{I8F)DFG) for

the measurement of regional cerebral glucose metabolism by positron emission tomography (PET) is now well established (1 -3) and has generated a widespread interest in the applica- tion of this technique. Concomitantly, there has been expended a substantial effort to find alternative routes for the synthesis of this radiopharmaceutical. The first synthesis (4) of 2-{I8F)DFG, involving the addition of "F-labeled F2 to tri-o-acetyl- D-glucal, gave a radiochemical yield of 10%. Recently, this yield has been increased by the use of {18F)-acetyl hypofluorite (5) or {I8F)-xenon difluoride (6, 7). However, these syntheses employ molecular fluorine, thereby restricting the theoretical incorporation of "F to 50%; these methods involve the pro- duction of isotope by the 20N(d,a) 18F reaction. In contrast, synthetic procedures based on I8F-fluoride ion would, in prin- ciple, allow all of the radioisotope to be incorporated into the required product, and would allow a choice from a wider range of nuclear reactions. For example, Levy et al. (8) have syn- thesized 2-{I8F)DFG in a radiochemical yield of lo%, by the displacement of the trifluoromethanesulfonyloxy group in methyl 4,6-0-benzylidene-3-O-methyl-2-O-trifluoromethane- sulfonyl-P-D-mannopyranoside. The low yield obtained by Levy et al. (8) is attributable in part to the low yield of CsH{"F)F2 obtained by trapping H"F on CSF, and in part to the difficulties in effecting the quantitative hydrolysis of the 1-0- and 3-0-methyl groups of the substrate. More recently Tewson (9) has synthesized the same compound in a radiochemical yield of 40% by a nucleophilic reaction at C-2 in methyl 4,6-0-benzylidene-P-D-mannopyranoside 2,3-cyclic sulfate. Tewson (9) reported an excellent (>90%) incorporation of I8F into the required fluorosulfate, a value approaching the theo- retical maximum. However, hydrolysis of the glycoside resulted in a considerably reduced yield of 2-{I8F)~FG. Never- theless, the radiochemical yield obtained is the highest reported at present.

In this laboratory, a synthesis of 2-deoxy-2-fluoro-D-glucose has been achieved (10) in high yield by the reaction of KHF2 with the acyclic a-nitroepoxide, 1 ,2-anhydro- 1 -deoxy-3,4 : 5,6-di-0-isopropylidene- 1-C-nitro-D-mannitol (1). We now re- port the reaction of KH{I8F)F2 with 1 (see Fig. l) , followed by

the removal of the blocking groups to afford 2-{I8F)DFG in a radiochemical yield of 10%; the overall synthesis time was less than one half-life of "F.

Experimental Production of ''F and labeling of KHFz

The production of "F was achieved by the 160(%e ,p )18~ reaction using the Queen's University 4-MeV Van de Graaff accelerator. Beam degradation in this low energy accelerator was circumvented by the use of a windowless ice target. The target consisted of a gold-plated, liquid nitrogen-cooled, cold finger located in the vacuum beam line of the accelerator. An external reservoir containing high purity water' supplied a constant jet of water vapor to the cold finger, and the ice layer thus formed was bombarded with the 4-MeV 3He beam. This arrangement afforded approximately 0.5 mCi of '9 per FA, accom- panied by the production of a similar quantity of 150 from the 16 0(3He,u)150 reaction. Normally a 2-p,A beam was used and approx-

imately 1 mCi of "F could be recovered from the target at the end of bombardment. A stoichiometric quantity of "F-fluoride ion-carrier in the form of KHFz (0.5 mole per mole of 1) was added as a 0.1 M aqueous solution to the "F-H,O (-3 mL) in a Teflon vial and the solution evaporated to dryness in a microwave oven.'

Reaction of the a-nitroepoxide 1 with KH{"F)Fz The u-nitroepoxide 1 was synthesized by the method of Szarek

et al. (10). A weighed amount of 1 (40-60 p,mol) was dried over PZ05 in a vacuum desiccator for 1 h prior to reaction. The solvent employed in this reaction, namely ethylene glycol, was initially treated with CaO and distilled under reduced pressure; it was then heated at reflux temperature in the presence of 4A molecular sieves and distilled. The distillate was stored over 4A molecular sieves; immediately prior to its use it was treated with Na under N, and distilled under reduced pressure. 'The sample of 1 was added to the KH{'"}F2 (20-30 p,mol) and the vial was sealed with a septum. The freshly distilled ethylene glycol (1 mL) was added to the vial and the solution heated, under NZ, at 135°C for 20-25 rnin. Thin-layer chromatography3 (tlc)

'HPLC-grade water; Fisher W5. 'CEM Corporation; model MDS-8 1. 3Thin-layer chromatography was performed using Merck plates pre-

coated with silica gel 60 F-254. The developed plates were dried and compounds located by heating the plates at - 150°C after they had been sprayed with 10% aqueous sulfuric acid containing 1% cerium sulfate and 1.5% molybdic acid.

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2710 CAN. J . CHEM. VOL. 62. 1984

C H O I

HC-"F I

0 H I

HOCHzCHzOH O C H 80% CF3COOH %CMe2 + KH[18FIF2 H C O

135°C ' Y c M e 2 H C O room temperature ' I

HCO, I

HCO,

3

FIG. 1. Synthesis of 2-deo~y-2-{'~F}fluoro-D-g~ucose.

(toluene - ethyl acetate, 1 : 1 (v/v)) indicated the presence of a trace of starting material 1 (R, 0.8), and of a component (R, 0.42) con- taining 15% (mean value obtained from 8 runs) of the total activity on the plate, the remainder of the activity being at the origin. The reaction mixture was cooled rapidly, diluted with distilled water ( I 0 mL), and the solution was extracted with dichloromethane (3 X 5 mL). The activity distribution between the organic and aqueous layers was com- parable to that observed above by tlc; this thin-layer chromatographic examination indicated that more than 90% of the activity present in the organic extracts was attributable to the product having R f 0.42, the remainder arising from a minute amount of non-migrating material carried over in the extraction procedure. The combined organic extracts were dried over anhydrous Na2SO4 and concentrated under reduced pressure.

Deprotection of 2-de0~-2-{'~F}jluoro-3,4 : 5,6-di-0-isopropylidene- aldehydo-D-glucose (2)

Method ( a ) Trifluoroacetic acid (80% (v/v), 2 mL) was added to the residue

obtained from the organic extracts, and the solution was heated to 50°C for 5 min. The acid was removed under reduced pressure (bath temperature < 50°C) and the residue was dissolved in aqueous ace- toniirile (0.3% water (v/v), 2 mL). Thin-layer chromatography (acetonitrile- water. 95 : 5 (v/v)l indicated that the maior radioactive . . . product migrated at the same rate as an authenhc sample of 2-deoxy-2-fluoro-D-glucose4 (RI 0.43). The component giving rise to the most intense spot in tlc (R, 0.1 I) did not contain any radioactivity. The presence of a minor component having Rf 0.32 was observed occasionally; this component migrated at the same rate as the product resulting from the treatment of the a-nitroepoxide 1 with tri- fluoroacetic acid. Thin-layer chromatography indicated the following activity distribution: 2-deoxy-2-fluoro-D-glucose, 80%; solvent front, 15%; origin, 5%.

Method (b) Trifluoroacetic acid (80% (v/v), 2 mL) was added to the residue

obtained from the organic extracts, and the solution was stirred at room temperature for 10 min; the reaction mixture was processed as described in Method (a). Thin-layer chromatography indicated the following activity distribution: 2-deoxy-2-fluoro-D-glucose, -86%; solvent front, 8%; origin -6%.

Column chromatography and high perfortnance liquid chromato- graphy (hplc) analysis

The aqueous acetonitrile solution (0.3% water (v/v)) of 2-{InF}DFG was chromatographed on a column (0.7 X 10 cm) of dry silica gel 60 using the same solvent as eluent. The initial 6-mL portion of the eluate was discarded and the product was obtained from the subsequent 10-mL portion. The solvent was removed under reduced pressure; hplc-grade water (-2.5 mL) and washed alumina (-50 mg) were added and the solution filtered through a 0.2-pm Millipore membrane. The product was analyzed by hplc (Bio-Rad Carbohydrate Column HPX-87; Varian 5000 Liquid Chromatograph; 85OC, water, flow rate

'Calbiochem-~ehrin~ Corporation.

0.6 mL/min; differential refractometer and Nal (TI) well counter detectors). The retention time of the product, detected by the refrac- tometer, was 9.7 min, corresponding to that of an authentic sample4 of 2-deoxy-2-fluoro-D-glucose. Only one major radioactive com- ponent was detected; this had a retention time of 10.0 min and a radiochemical purity greater than 97%. The retention times indicated by the two detectors corresponded to those obtained using a sample of 2-{"F}DFG provided by the Montreal Neurological Institute. Chem- ical and radiochemical yields of 2-deoxy-2-fluoro-D-glucose were of a comparable value, namely -10% (mean value obtained from 8 runs).

Activity measurements Activity distribution on the plates was measured by fractional

counting after charring using 10% H2S04 in methanol. The activities in aqueous and organic solutions were measured by counting 100-pL samples at a fixed geometry to the detector system. For relative activity distributions, a 7.62 X 7.62 cm Nal (TI) detector coupled to a single-channel analyzer, or an intrinsic Ge detector coupled to a multichannel analyzer, was used. The absolute activity of "F-fluoride ion and of 2-{"F}DFG was measured by counting 100-pL samples at a fixed geometry to the calibrated Ge detector.

Results and discussion This article described a new synthesis of 2-{I8F)DFG in a

radiochemical (decay-corrected) yield of 10%. Although this yield is lower than some of those recently reported, never- theless the synthetic method is a viable alternative for the "in-house" production of 2-{I8F)DFG. Because the chemical yield of 2-deoxy-2-fluoro-D-glucose was comparable to the radiochemical yield, it was considered to be unnecessary to treat the accelerator-derived I8F-H20 with a cation-exchange resin, as was necessary in the method of Tewson (9). Such treatment may result in a loss of I8F, thereby reducing the radiochemical yield of the radiopharmaceutical when based on the starting activity of I8F. The starting material, 1,2-anhydro- 3,4 : 5,6-di-0-isopropylidene- 1 -C-nitro-D-mannitol (I), has been isolated (10) in crystalline form in 72% yield from a mixture of 1,2-anhydro-3,4 : 5,6-di-0-isopropylidene- 1 -C- nitro-D-mannitol and -D-glucitol which was obtained in 80% yield by treatment of 1,2-dideoxy-3,4:5,6-di-0-isopropyl- idene- 1 -C-nitro-D-arabino-hex- I -enit01 with 30% hydrogen peroxide; this nitroolefin can be obtained from D-arabinose in a good overall yield. The substrate 1 is a crystalline solid that is stable when stored at 0°C. Full details of the carbohydrate synthesis will be published separately.

In the reaction of the a-nitroepoxide 1 with KHF2 it has been found that a 12-fold molar-equivalent excess of KHF2 affords 2-deoxy-2-fluoro-D-glucose in a yield greater than 30% based on 1 and that further increasing the concentration of the fluoride

v

salt enhances the yield. However, reactions using up to 2.5

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BEELEY ET AL. 271 1

molar equivalents of KHFz did not afford yields of 2-deoxy- 2-fluoro-D-glucose beyond lo%, in experiments leading to either labeled or unlabeled product. A decrease in the amount of KHF, carrier to 5 pno l per 50 ~ m o l of 1 resulted in a 15% incorporation of I8F into the fluoro aldehyde 2; thus the use of less than 1 molar equivalent of KHFz allows the specific activity of the product to be increased, if so desired.

The use of ethylene glycol as the solvent in the reaction of the a-nitroepoxide with fluoride ion resulted in the formation of a component the Rf of which was similar to that of the product, fluoro aldehyde 2; however, treatment of the mixture

, of products with 80% trifluoroacetic acid converted this com- ponent into one (R, 0.1 1, acetonitrile-water, 95 :5 (v/v)) which was well separated from 2-deoxy-2-fluoro-D-glucose (R, 0.43). Competing reactions with ethylene glycol have been observed (1 1 - 13) previously with carbohydrate substrates. The use of a variety of other solvents, together with a number of different fluoride salts as well as crown-ether systems (14), has been investigated and the results will be reported in the separate publication.

In the present study the fluoro aldehyde 2 has also been obtained by treatment of the a-nitroepoxide 1 with 18~-labeled tetra-n-butylammonium fluoride in ethylene glycol; the reagent salt was dried by the repeated addition of acetonitrile and dis- tillation using a rotary evaporator. In these experiments the yields (- 10%) of 2 that were obtained were lower than was the case with KHFz and were less reproducible. The use of the technique of microwave drying (15) combined with the per- formance of the reactions in Teflon vials greatly reduce the loss of I8F prior to its reaction and give better reproducibility.

The deprotection of the fluoro aldehyde 2 was originally accomplished using boron trichloride in dichloromethane. However, the time required for the efficient removal of boron salts during processing prompted the use of 80% trifluoroacetic acid. On the small scales employed in the experiments no epimerization of the fluoro aldehyde to the manno epimer was observed with the use of this reagent. If the reaction were performed at room temperature for 10 min, rather than at 50°C for 5 min, less radioactivity was observed at the solvent front in tlc; however, there was slightly more radioactivity at the origin, possibly resulting from the liberation of I8F-fluoride ion. Nevertheless, the room-temperature reaction provided a higher radiochemical purity of the final product 3.

The use of the a-nitroepoxide substrate 1 is currently being adapted for the synthesis of 2-{I8F)DFG employing ' 8 ~

produced with the McMaster University nuclear reactor.

beam-energy constraints of the accelerator employed necessi- tated the use of an unconventional target, it seems reasonable to assume that the method is applicable to any PET facility employing a conventional target. The amounts of radioactivity incorporated, of course, depend upon the concentrations of "F achieved by the accelerator.

Acknowledgements The authors are grateful for financial support in the form of

grants from the Medical Research Council of Canada and the Natural Sciences and Engineering Research Council of Canada. The high performance liquid chromatograph em- ployed in this study was purchased by funds provided by the Medical Research Council of Canada. The authors also wish to thank the members of the Nuclear Physics Laboratory of Queen's University, especially Drs. W. McLatchie and H.-B. Mak, for their cooperation in the production of I8F.

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T. IDO, V. CASELLA, J. FOWLER, E. HOFFMAN, A. ALAVI, P. SOM, and L. SOKOLOFF. Circ. Res. 44, 127 (1979).

3. M. E. PHELPS, S. C. HUANG, E. J. HOFFMAN, C. SELIN, L. SOKOLOFF, and D. E. KUHL. Ann. Neurol. 6, 371 (1979).

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6. C.-Y. SHIUE, K.-C. TO, and A. P. WOLF. J. Labelled Compd. Radiopharm. 20, 157 (1983).

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9. T. J. TEWSON. J . Nucl. Med. 24, 718 (1983). 10. W. A. SZAREK, G. W. HAY, and M. M. PERLMUTTER. J. Chem.

Soc. Chem. Commun. 1253 (1982). 11. J. E. G. BARNETT. Adv. Carbohydr. Chem. 22, 177 (1967); see

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445 (1982).

Conclusion The methodology described in this article provides an alter-

native route to 2-{I8F}D~G using a water target. Although the Can

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