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Molecular Imaging and Contrast Agent Database (MICAD) [Internet].

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2’-[18F]Fluorofolic acid

, PhD
National Center for Biotechnology Information, NLM, NIH

Created: ; Last Update: March 9, 2011.

Chemical name:2’-[18F]Fluorofolic acidimage 110322819 in the ncbi pubchem database
Abbreviated name:2’-[18F]FFA
Agent category:Compound
Target:Putative folate receptor
Target category:Receptor
Method of detection:Positron emission tomography (PET)
Source of signal:18F
  • Checkbox In vitro
  • Checkbox Rodents
Click on the above structure for additional information in PubChem.



Folic acid is a water-soluble vitamin B9 (1) that is essential for methylation and DNA synthesis. The primary pathway for entry of folate into cells is through the facilitated transporter, which has a low affinity for folate (Michaelis constant (Km) = 1–5 μM). Epithelial cells in the choroid plexus, kidney, lung, thyroid, spleen, placenta, and thymus also possess a receptor on the cell membrane with higher affinity (dissociation constant (Kd) = 0.5 nM), which allows folate uptake via receptor-mediated endocytosis. Some human epithelial tumor cells were found to overexpress folate-binding protein (2). More than 90% of human ovarian and endometrial cancers express the high-affinity receptor, which is absent in the corresponding normal tissues. Breast, colorectal, renal, and lung carcinomas also overexpress the folate receptor but at lower frequencies (20%–50%). Activated macrophages, but not resting macrophages, have also been found to have the folate receptor (3).

Several folate-based conjugates (111In-DTPA-folate, 99mTc-EC-folate, and 68/67/66Ga-DF-folate) have been studied in tumor imaging (4-7). Bettio et al. (8) reported a synthesis of 18F-labeled folate by reaction of [18F]4-fluorobenzylamine (FBA) with the α- and γ-carboxyl groups of folic acid. [18F]α/γ-FBA-folate has been evaluated as a positron emission tomography (PET) agent for detection of folate receptors in tumors in mice. However, the low radiochemical yield from multistep radiosynthesis limited its routine use. Ross et al. (9) introduced the 18F label at the 2’ position of the 4-amino-benzoyl moiety in folic acid to form 2’-[18F]fluorofolic acid (2’-[18F]FFA). 2’-[18F]FFA was found to be a specific, high-affinity PET agent for imaging folate receptor-positive tumors in mice.



Ross et al. (9) prepared 2’-[18F]FFA with a two-step synthesis using a standard nucleophilic radiofluorination ([18F]KF/Kryptofix 2.2.2) reaction of N2-(N,N-dimethylaminomethylene)-2'-nitrofolic acid di-tert-butylester (8 µmol) (140ºC, 20 min) and subsequent acid hydrolysis (60ºC, 12 min). Maximal overall decay-corrected yield was 4% with >95% radiochemical purity. The total synthesis time was 80 min, and the specific activity was 23.7 ± 12.2 GBq/µmol (0.64 ± 0.33 Ci/µmol). The relative lipophilicity (retention factor, k’) of 2’-FFA as determined with high-performance liquid chromatography was 0.53 as compared to 0.30 for folic acid.

In Vitro Studies: Testing in Cells and Tissues


The human nasopharyngeal carcinoma KB cell line has putative folate receptors as determined with [3H]folate binding studies in cultures (8). The mean 50% inhibition concentrations for 2’-FFA and folic acid were 3.4 ± 0.3 and 1.1 ± 0.4 nM (9), respectively. Therefore, 2’-FFA has a binding affinity comparable to that of native folic acid.

Animal Studies



Ross et al. (9) performed biodistribution studies of 1 MBq (0.027 mCi) 2’-[18F]FFA in nude mice (n = 4) bearing KB tumor xenografts. The organ with the highest accumulation at 75 min after injection was the kidney (46.06% injected dose/gram (ID/g)), followed by the gallbladder (17.69% ID/g), tumor (9.37% ID/g), liver (7.79% ID/g), and intestine (2.80% ID/g). Very high radioactivity was found in the urine. The radioactivity levels in the blood and bone were 0.40% ID/g and 1.59% ID/g, respectively. Accumulation in the brain, lung, heart, spleen, and stomach was <2% ID/g. There was ~33% intact 2’-[18F]FFA in the plasma and tumor at 15 min after injection with one major hydrophilic metabolite. Pretreatment with folic acid (200 μg/mouse) 10 min before 2’-[18F]FFA injection reduced 2’-[18F]FFA accumulation by 90% in the brain, 83% in the tumor, and 81% in the kidney. Less reduction was observed in the lung, heart, spleen, and stomach.

The whole-body distribution of 2’-[18F]FFA was also assessed with PET imaging 75–105 min after injection. High accumulation was visualized in the kidneys, gallbladder, urinary bladder, tumor, and intestines. Moderate accumulation was observed in the liver. The accumulation of 2’-[18F]FFA in the tumor and kidneys was reduced to near background level after folic acid pretreatment.

Other Non-Primate Mammals


No publication is currently available.

Non-Human Primates


No publication is currently available.

Human Studies


No publication is currently available.


Stanger O. Physiology of folic acid in health and disease. Curr Drug Metab. 2002;3(2):211–23. [PubMed: 12003352]
Ke C.Y., Mathias C.J., Green M.A. The folate receptor as a molecular target for tumor-selective radionuclide delivery. Nucl Med Biol. 2003;30(8):811–7. [PubMed: 14698784]
Nakashima-Matsushita N., Homma T., Yu S., Matsuda T., Sunahara N., Nakamura T., Tsukano M., Ratnam M., Matsuyama T. Selective expression of folate receptor beta and its possible role in methotrexate transport in synovial macrophages from patients with rheumatoid arthritis. Arthritis Rheum. 1999;42(8):1609–16. [PubMed: 10446858]
Mathias C.J., Hubers D., Low P.S., Green M.A. Synthesis of [(99m)Tc]DTPA-folate and its evaluation as a folate-receptor-targeted radiopharmaceutical. Bioconjug Chem. 2000;11(2):253–7. [PubMed: 10725102]
Mathias C.J., Lewis M.R., Reichert D.E., Laforest R., Sharp T.L., Lewis J.S., Yang Z.F., Waters D.J., Snyder P.W., Low P.S., Welch M.J., Green M.A. Preparation of 66Ga- and 68Ga-labeled Ga(III)-deferoxamine-folate as potential folate-receptor-targeted PET radiopharmaceuticals. Nucl Med Biol. 2003;30(7):725–31. [PubMed: 14499330]
Mathias C.J., Wang S., Low P.S., Waters D.J., Green M.A. Receptor-mediated targeting of 67Ga-deferoxamine-folate to folate-receptor-positive human KB tumor xenografts. Nucl Med Biol. 1999;26(1):23–5. [PubMed: 10096497]
Mathias C.J., Wang S., Waters D.J., Turek J.J., Low P.S., Green M.A. Indium-111-DTPA-folate as a potential folate-receptor-targeted radiopharmaceutical. J Nucl Med. 1998;39(9):1579–85. [PubMed: 9744347]
Bettio A., Honer M., Muller C., Bruhlmeier M., Muller U., Schibli R., Groehn V., Schubiger A.P., Ametamey S.M. Synthesis and Preclinical Evaluation of a Folic Acid Derivative Labeled with 18F for PET Imaging of Folate Receptor-Positive Tumors. J Nucl Med. 2006;47(7):1153–1160. [PubMed: 16818950]
Ross T.L., Honer M., Muller C., Groehn V., Schibli R., Ametamey S.M. A new 18F-labeled folic acid derivative with improved properties for the PET imaging of folate receptor-positive tumors. J Nucl Med. 2010;51(11):1756–62. [PubMed: 20956469]
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