Abstract
This protocol describes the step-by-step procedure for the synthesis of N-succinimidyl 4-[18F]fluorobenzoate ([18F]SFB), an agent widely used for labeling proteins and peptides with the positron-emitting radionuclide 18F. The protocols for the synthesis of unlabeled SFB and the quaternary salt precursor 4-formyl-N,N,N-trimethyl benzenaminium trifluoromethane sulfonate also are described. For the [18F]SFB synthesis, the quaternary salt is first converted to 4-[18F]fluorobenzaldehyde. Oxidation of the latter provides 4-[18F]fluorobenzoic acid, which is converted to [18F]SFB by treatment with N,N-disuccinimidyl carbonate. Using this method, [18F]SFB can be synthesized in decay-corrected radiochemical yields of 30%–35% and a specific radioactivity of 11–12 GBq μmol−1. The total synthesis and purification time required is about 80 min, starting from delivery of the [18F]fluoride. [18F]SFB remains an optimal reagent for labeling proteins and peptides with 18F because of good conjugation yields and metabolic stability.
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Kelloff, G.J. et al. The progress and promise of molecular imaging probes in oncologic drug development. Clin. Cancer Res. 11, 7967–7985 (2005).
Weissleder, R. Molecular imaging in cancer. Science 312, 1168–1171 (2006).
Rohren, E.M., Turkington, T.G. & Coleman, R.E. Clinical applications of PET in oncology. Radiology 231, 305–332 (2004).
Okarvi, S.M. Recent progress in fluorine-18 labelled peptide radiopharmaceuticals. Eur. J. Nucl. Med. 28, 929–938 (2001).
Wilbur, D.S. Radiohalogenation of proteins: an overview of radionuclides, labeling methods, and reagents for conjugate labeling. Bioconjug. Chem. 3, 433–470 (1992).
Chang, Y.S. et al. Preparation of 18F-human serum albumin: a simple and efficient protein labeling method with 18F using a hydrazone-formation method. Bioconjug. Chem. 16, 1329–1333 (2005).
de Bruin, B. et al. 1-[3-(2-[18F]fluoropyridin-3-yloxy)propyl]pyrrole-2,5-dione: design, synthesis, and radiosynthesis of a new [18F]fluoropyridine-based maleimide reagent for the labeling of peptides and proteins. Bioconjug. Chem. 16, 406–420 (2005).
Garg, P.K., Garg, S. & Zalutsky, M.R. Fluorine-18 labeling of monoclonal antibodies and fragments with preservation of immunoreactivity. Bioconjug. Chem. 2, 44–49 (1991).
Glaser, M. et al. 18F-fluorothiols: a new approach to label peptides chemoselectively as potential tracers for positron emission tomography. Bioconjug. Chem. 15, 1447–1453 (2004).
Kilbourn, M.R., Dence, C.S., Welch, M.J. & Mathias, C.J. Fluorine-18 labeling of proteins. J. Nucl. Med. 28, 462–470 (1987).
Shai, Y. et al. 18F-labeled insulin: a prosthetic group methodology for incorporation of a positron emitter into peptides and proteins. Biochemistry 28, 4801–4806 (1989).
Toyokuni, T. et al. Synthesis of a new heterobifunctional linker, N-[4-(aminooxy)butyl]maleimide, for facile access to a thiol-reactive 18F-labeling agent. Bioconjug. Chem. 14, 1253–1259 (2003).
Wust, F., Muller, M. & Bergmann, R. Synthesis of 4-([F-18]fluoromethyl)-2-chlorophenylisothiocyanate: A novel bifunctional F-18-labelling agent. Radiochimica Acta 92, 349–353 (2004).
Vaidyanathan, G. & Zalutsky, M.R. Labeling proteins with fluorine-18 using N-succinimidyl 4-[18F]fluorobenzoate. Appl. Radiat. Isot. 19, 275–281 (1992).
Vaidyanathan, G. & Zalutsky, M.R. Improved synthesis of N-succinimidyl 4-[18F]fluorobenzoate and its application to the labeling of a monoclonal antibody fragment. Bioconjug. Chem. 5, 352–356 (1994).
Vaidyanathan, G. & Zalutsky, M.R. Preparation of N-succinimidyl 3-[*I]iodobenzoate: an agent for the indirect radioiodination of proteins. Nat. Protocols 1, 707–713 (2006).
Vaidyanathan, G. & Zalutsky, M.R. Fluorine-18-labeled [Nle4,D-Phe7]-α-MSH, an α-melanocyte stimulating hormone analogue. Nucl. Med. Biol. 24, 171–178 (1997).
Vaidyanathan, G. & Zalutsky, M.R. Fluorine-18 labeled chemotactic peptides: a potential approach for the PET imaging of bacterial infection. Nucl. Med. Biol. 22, 759–764 (1995).
Page, R.L., Garg, P.K., Vaidyanathan, G. & Zalutsky, M.R. Preclinical evaluation and PET imaging of 18F-labeled Mel-14 F(ab')2 fragment in normal dogs. Nucl. Med. Biol. 21, 911–919 (1994).
Vaidyanathan, G., Bigner, D.D. & Zalutsky, M.R. Fluorine-18-labeled monoclonal antibody fragments: a potential approach for combining radioimmunoscintigraphy and positron emission tomography. J. Nucl. Med. 33, 1535–1541 (1992).
Wester, H.J., Hamacher, K. & Stocklin, G. A comparative study of N.C.A. fluorine-18 labeling of proteins via acylation and photochemical conjugation. Nucl. Med. Biol. 23, 365–372 (1996).
Azarian, V. et al. Synthesis and liposome encapsulation of a novel 18F-conjugate of ω-conotoxin GVIA for the potential imaging of N-type Ca2+ channels in the brain by positron emission tomography. J. Labelled Comp. Radiopharm. 49, 269–283 (2006).
Chen, X. et al. 18F-labeled RGD peptide: initial evaluation for imaging brain tumor angiogenesis. Nucl. Med. Biol. 31, 179–189 (2004).
Grierson, J.R. et al. Production of [F-18]fluoroannexin for imaging apoptosis with PET. Bioconjug. Chem. 15, 373–379 (2004).
Guenther, K.J. et al. Synthesis and in vitro evaluation of 18F- and 19F-labeled insulin: a new radiotracer for PET-based molecular imaging studies. J. Med. Chem. 49, 1466–1474 (2006).
Kiesewetter, D.O. et al. Fluoro-, bromo-, and iodopaclitaxel derivatives: synthesis and biological evaluation. Nucl. Med. Biol. 30, 11–24 (2003).
Lang, L. et al. Development of fluorine-18-labeled 5–HT1A antagonists. J. Med. Chem. 42, 1576–1586 (1999).
Zhang, X. et al. 18F-labeled bombesin analogs for targeting GRP receptor-expressing prostate cancer. J. Nucl. Med. 47, 492–501 (2006).
Haka, M.S., Kilbourn, M.R., Watkins, G.L. & Toorongian, S.A. Aryltrimethylammonium trifluoromethanesulfonates as precursors to aryl [18F]fluorides: improved synthesis of [18F]GBR-13119. J. Labelled Comp. Radiopharm. 27, 823–833 (1989).
Pascali, G., Kiesewetter, D.O., Salvadori, P.A. & Eckelman, W.C. Use of 1,8-bis-(dimethylamino)naphthalene/H18F complex as a new fluorinating agent. J. Labelled Comp. Radiopharm. 47, 373–383 (2004).
Mading, P., Fuchtner, F. & Wust, F. Module-assisted synthesis of the bifunctional labelling agent N-succinimidyl 4-[18F]fluorobenzoate ([18F]SFB). Appl. Radiat. Isot. 63, 329–332 (2005).
Wust, F., Hultsch, C., Bergmann, R., Johannsen, B. & Henle, T. Radiolabelling of isopeptide N epsilon-(gamma-glutamyl)-L-lysine by conjugation with N-succinimidyl-4-[18F]fluorobenzoate. Appl. Radiat. Isot. 59, 43–48 (2003).
Zijlstra, S., Gunawan, J., Freytag, C. & Burchert, W. Synthesis and evaluation of fluorine-18 labelled compounds for imaging of bacterial infections with PET. Appl. Radiat. Isot. 64, 802–807 (2006).
Zijlstra, S., Gunawan, J. & Burchert, W. Synthesis and evaluation of a 18F-labelled recombinant annexin-V derivative, for identification and quantification of apoptotic cells with PET. Appl. Radiat. Isot. 58, 201–207 (2003).
Kilbourn, M.R. in Fluorine-18 Labeling of Radiopharmaceuticals 41–42 (National Academy Press, Washington, D.C., 1990).
Schlyer, D.J., Firouzbakht, M.L. & Wolf, A.P. Impurities in the [18O]water target and their effect on the yield of an aromatic displacement reaction with [18F]fluoride. Appl. Radiat. Isot. 44, 1459–1465 (1993).
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This work was supported by grants from US Department of Energy and the US National Institutes of Health.
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G.V. conceived the idea, designed and executed the experiments and contributed to the writing of the paper. M.R.Z. provided financial support and contributed to the writing of the paper.
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Vaidyanathan, G., Zalutsky, M. Synthesis of N-succinimidyl 4-[18F]fluorobenzoate, an agent for labeling proteins and peptides with 18F. Nat Protoc 1, 1655–1661 (2006). https://doi.org/10.1038/nprot.2006.264
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DOI: https://doi.org/10.1038/nprot.2006.264
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