Development of ¹⁸F-Fluoroethylcholine for Cancer Imaging with PET: Synthesis, Biochemistry, and Prostate Cancer Imaging

(A) Automated apparatus for ¹⁸F-FECh synthesis. Reaction vessel is shown close up. Purification module is out of view. (B) Preparative HPLC of ¹⁸F-FECh in automated apparatus for ¹⁸F-FECh synthesis. Column used was ODS-silica gel column, 250 × 10 mm; solvent, 50 mmol/L phosphoric acid + 1 mmol/L 2-naphthalenesulfonic acid; flow rate, 5 mL/min. Radioactivity “halfway down the column” is reading of detector (detector 1) placed on side of column to watch approach of ¹⁸F-FECh. Radioactivity of effluent from column was monitored by another detector (detector 2).

(A) HPLC after incubation of ¹⁸F-FECh with choline kinase and with choline oxidase. No-carrier-added ¹⁸F-FECh chloride was incubated with yeast choline kinase and 15 μmol ATP or with bacterial choline oxidase. From incubated specimen, methanol-water–soluble component was separated and fractionated on cation- and anion-exchange HPLC. ¹⁸F radioactivity was measured in each fraction. (B) HPLC after incubation of ¹⁸F-FECh with choline kinase and γ-³²P-ATP. No-carrier-added ¹⁸F-FECh chloride (approximately 0.005 μmol) was incubated with yeast choline kinase and 0.01 μmol γ-³²P-ATP. From incubated specimen, methanol-water–soluble component was separated and fractionated on anion-exchange HPLC. ¹⁸F and ³²P radioactivities were measured in each fraction. kcpm = kilocounts per minute.

HPLC of methanol-water layer after incubation of ¹⁸F-FECh with Ehrlich ascites tumor cells. No-carrier-added ¹⁸F-FECh chloride was incubated with Ehrlich ascites tumor cells (approximately 10⁶ cells/mL) in glucose-fortified Hanks’ solution for 30 min. Methanol-water–soluble component was separated from incubated cells and divided into halves. One half was analyzed on HPLC using cation-exchange resin and other half was analyzed on HPLC using anion-exchange resin. ¹⁸F radioactivity was measured in every fraction.

TLC of chloroform layer after incubation of ¹⁸F-FECh with Ehrlich ascites tumor cells (autoradiography). No-carrier-added ¹⁸F-FECh chloride was incubated with Ehrlich ascites tumor cells (approximately 10⁶ cells/mL) in glucose-fortified Hanks’ solution for 30 min. Chloroform-soluble fraction of cell extract was concentrated and developed on TLC silica gel plates. Solvent was as follows: for alkaline system, chloroform, methanol, and 28% ammonia (65:35:5); for acidic system, benzene, pyridine, and formic acid (50:40:10). Autoradiography of TLC plates was performed using imaging plate detector system. Authentic choline-containing phospholipids were developed in same way as above and are indicated by arrows. PCh = phosphatidylcholine; Sph = sphingomyelin; Lyso = lysophosphatidylcholine.

Time course of uptake and metabolism of ¹⁸F-FECh in Ehrlich ascites tumor cells. No-carrier-added ¹⁸F-FECh chloride was incubated with Ehrlich ascites tumor cells (approximately 10⁶ cells/mL) in glucose-fortified Hanks’ solution for various periods up to 60 min. Total ¹⁸F uptake was measured after washing cells by centrifugation, and cell-to-medium ratio was calculated after hematocrit determination. After treatment of cells with methanol and chloroform, radioactivities in free ¹⁸F-FECh, phosphoryl-¹⁸F-FECh, ¹⁸F-labeled phospholipids (phosphatidyl-¹⁸F-FECh and so forth), and proteins were measured.