Synthesis of [18F]PS13 and Evaluation as a PET Radioligand for Cyclooxygenase-1 in Monkey

Abstract

Cyclooxygenase-1 (COX-1) and its isozyme COX-2 are key enzymes in the syntheses of prostanoids. Imaging of COX-1 and COX-2 selective radioligands with positron emission tomography (PET) may clarify how these enzymes are involved in inflammatory conditions and assist in the discovery of improved anti-inflammatory drugs. We have previously labeled the selective high-affinity COX-1 ligand, 1,5-bis(4-methoxyphenyl)-3-(2,2,2-trifluoroethoxy)-1H-1,2,4-triazole (PS13), with carbon-11 (t_1/2 = 20.4 min). This radioligand ([¹¹C]PS13) has been successful for PET imaging of COX-1 in monkey and human brain and in periphery. [¹¹C]PS13 is being used in clinical investigations. Alternative labeling of PS13 with fluorine-18 (t_1/2 = 109.8 min) is desirable to provide a longer-lived radioligand in high activity that might be readily distributed among imaging centers. However, labeling of PS13 in its 1,1,1-trifluoroethoxy group is a radiochemical challenge. Here we assess two labeling approaches based on nucleophilic addition of cyclotron-produced [¹⁸F]fluoride ion to gem-difluorovinyl precursors, either to label PS13 in one step or to produce [¹⁸F]2,2,2-trifluoroethyl p-toluenesulfonate for labeling a hydroxyl precursor. From the latter two-step approach, we obtained [¹⁸F]PS13 ready for intravenous injection in a decay-corrected radiochemical yield of 7.9% and with a molar activity of up to 7.9 GBq/μmol. PET imaging of monkey brain with [¹⁸F]PS13 shows that this radioligand can specifically image and quantify COX-1 without radiodefluorination but with some radioactivity uptake in skull, ascribed to red bone marrow. The development of a new procedure for labeling PS13 with fluorine-18 at a higher molar activity is, however, desirable to suppress occupancy of COX-1 by carrier at baseline.

Keywords: COX-1; Fluorine-18; PET; PS13; brain imaging; nucleophilic addition; radioligand.

Figure 1.

Chemical structures showing the labeling site for A [¹¹C]PS13 and B [¹⁸F]PS13.

Figure 2.

Strategies for ¹⁸F-labeling of a 1,1,1-trifluoroethoxy group: A, a one-step approach (Fawaz et al.); B, a two-step approach (Rafique et al.); and C, radiosynthesis approaches investigated in this work for the synthesis of [¹⁸F]PS13.

Figure 3.

One-step approach for the synthesis of [¹⁸F]PS13 including precursor 5 synthesis. Reagents and conditions: (i) 4-methoxybenzoyl chloride, toluene, pyrimidine, 110 °C, 2.5 h; (ii) KOH (10%, aq.), EtOH, 60 °C, 1.5 h; (iii) 1. K₂CO₃, DMF, RT, 10 min and 2. 1,1,1-trifluoro-2-iodoethane, 100 °C, 3 h; (iv) n-BuLi (1.6 M in hexanes), THF, −78 °C, 45 min; (v) [¹⁸F]F⁻, K 2.2.2, K₂CO₃, DMSO, proton source, thermal heating, 10–20 min.

Figure 4.

Representation of the ¹⁹F/¹⁸F addition–elimination process for 5 in the absence of an added proton source.

Figure 5.

Two-step approach for the synthesis of [¹⁸F]PS13.

Figure 6.

PET studies of brain in monkey A with [¹⁸F]PS13 (carrier PS13:6.6 nmol/kg at baseline). Panel A: regional V_T images from baseline scan (top row) and blocked scans (with PS13, 0.3 mg/kg, i.v.) (middle row), and corresponding MRI images (bottom row). Left column: coronal images; middle column: sagittal images; right column: horizontal images. Highest radioactivity uptake occurred in the prefrontal and parietal cortices. Panel B: whole brain time-activity curves at baseline and after preblock obtained with [¹⁸F]PS13. Corresponding curves obtained in a different monkey with [¹¹C]PS13 from the study in ref 17 with a much lower PS13 carrier dose (0.19 nmol/kg at baseline) are shown for comparison.

Figure 7.

Plasma clearance and metabolism of [¹⁸F]PS13 in monkey A. Panel A: reversed phase HPLC analysis of radiometabolites in plasma at 180 min after intravenous radioligand injection at baseline. Panel B: time-course of the percentage of radioactivity in plasma represented by unchanged [¹⁸F]PS13 under baseline and preblocked conditions. Panel C: time-course for unchanged [¹⁸F]PS13 in plasma under baseline and preblocked conditions.

Figure 8.

Analysis of PET data from monkey A under baseline and preblock conditions. Panel A: regional V_T values. Panel B: regional V_T values adjusted for normalized plasma free fraction (Supporting Information, Table S1). Panel C: V_T values determined from PET data for different durations of scan data from time of radioligand injection, normalized to the value at the end of scanning (180 min). Panel D: Lassen plot showing V_ND as the X-axis intercept and occupancy (slope of plot) of the available COX-1 (that not already occupied by carrier at baseline) by the blocking agent PS13 (0.3 mg/kg, i.v.).

Figure 9.

Summed (0–180 min) whole-body MIP PET image obtained with [¹⁸F]PS13 in monkey.