Automated radiosynthesis of 18F-FBEM, a sulfhydryl site specific labeling agent for peptides and proteins

Objectives In the process of developing ¹⁸F-FBEM-(Cys⁴⁰)-exendin-4 for imaging the glucagon-like peptide 1 receptor (GLP-1R) in the pancreas, we have instituted a robust automated synthesis of ¹⁸F-FBEM, a sulfhydryl (-SH) site specific agent for radiolabeling of peptides and proteins.

Methods Radiosynthesis of ¹⁸F-FBEM was performed in a TRACERlab FXN-Pro module using a 3-step reaction sequence in two separate reactors. Each step of the automated process was optimized during a series of test runs. In reactor 1, the ammonium salt precursor 1 (2-3 mg) was reacted with dried ¹⁸F^-/K222 in DMSO (0.4 mL) at 105 °C for 10 min to generate 2, which was hydrolyzed to 3 by heating with 0.1 N NaOH (0.2 mL) at 95 °C for 5 min. After neutralization with 0.2 N HCl (0.2 mL), the reaction mixture was diluted with two portions of water (5 mL each) and passed separately through a Waters C18 Light SepPak. The intermediate 3 was then eluted with 1 mL of CH₃CN/DMSO (3/1, v/v) into the second reactor of the module, and a solution of N-(2-aminoethyl) maleimide (3-5 mg) in DMSO (0.2 mL) was added, followed by DECP and DIPEA in 0.1 mL of CH₃CN each. The reaction mixture was heated at 70 °C for 7 min, diluted with 1mM HCl and purified by preparative HPLC. Dilution of of the HPLC fraction, solid phase extraction on a second C18 Light SepPak catridge, and elution with CH₃CN (1 mL) gave pure ¹⁸F-FBEM (4).

Results The fully automated, two-pot, three-step radiosynthesis generated 1.67-3.89 GBq of of ¹⁸F-FBEM from 22.2 GBq of starting ¹⁸F-fluoride.The radiochemical yield ranged from 7.5 to 17.5% (non-decay corrected), with specific activity of 31.8±5.3 MBq/nmol (n=3) at the end of synthesis. Radiochemical purity of ¹⁸F-FBEM was greater than 98%, and total synthesis time was ~90 min.

Conclusions We have developed a robust automated method to synthesize ¹⁸F-FBEM using a FXN-Pro module. ¹⁸F-FBEM was then used in the radiolabeling of (Cys⁴⁰)-exendin-4 to produce ¹⁸F-FBEM-(Cys⁴⁰)-exendin-4 for imaging of GLP-1R in the pancreas of normal and diabetic rats. $$graphic_016BB236-D290-40FB-B2C9-F5FC83A07042$$