Syntheses of [¹¹C]2- and [¹¹C]3-trifluoromethyl-4-aminopyridine: potential PET radioligands for demyelinating diseases

[¹¹C]fluoroform was used to produce ¹¹C-trifluoromethylated derivatives of 4-aminopyridine with high molar activity for PET imaging of the brain.

Abstract

Trifluoromethyl groups are of great interest in PET radiopharmaceuticals. Radiolabelled 4-aminopyridine (4AP) derivatives have been proposed for imaging demyelinating diseases. Here, we describe methods for producing ¹¹C-trifluoromethylated derivatives of 4AP and present early imaging results with [¹¹C]3-trifluoromethyl-4AP in a rhesus macaque. This study shows the utility of [¹¹C]CuCF₃ for labelling pyridines and provides initial evidence for the potential use of [¹¹C]3-trifluoromethyl-4AP as a PET radioligand.

Introduction

The trifluoromethyl group (CF₃) has become prevalent in pharmaceuticals.1 Trifluoromethyl groups can alter physicochemical properties, such as pK_a, lipophilicity, and stability, and in turn enhance pharmacodynamics, metabolic stability, and bioavailability.2 Furthermore, the trifluoromethyl group plays an increasing role in the development of radiopharmaceuticals for imaging with positron emission tomography (PET) as it provides a chemical handle that is now amenable to labelling with either of the positron-emitters, carbon-11 (t_1/2 = 20.4 min)3 or fluorine-18 (t_1/2 = 109.8 min).4–10

The continual development of CF₃-containing radio-pharmaceuticals has driven the investigation of novel methods to radiolabel CF₃ groups. Most of the methods described to date have focused on labelling the CF₃ group with fluorine-18.4–10 These ¹⁸F-labelling reactions vary in their radiochemical yields, but their main limitation for broad application in producing useful PET radiotracers lies in the low to moderate molar activities (radioactivity per mole) that may be achieved (Fig. 1). To circumvent this limitation, Haskali and Pike recently developed the synthesis of [¹¹C]fluoroform from cyclotron-produced [¹¹C]methane, which allows the labelling of small molecules with molar activities exceeding 200 GBq μmol^–1.3 The shorter half-life of carbon-11 compared to that of fluorine-18 limits its use to sites that have ready access to a biomedical cyclotron and radiosynthesis capability. Nonetheless, carbon-11 allows scanning of the same subject more than once in a day. For example, baseline and blocking experiments can be done in the same animals on the same day, and in many institutions, opens the possibility to scan the same human subject twice in the same day either, with two ¹¹C-labelled tracers or with one ¹¹C- and one ¹⁸F-labelled tracer. This can be extremely valuable during radiotracer development and in PET imaging programs. Briefly, the process developed by Haskali and Pike uses cobalt(iii) fluoride (CoF₃) to convert [¹¹C]methane into [¹¹C]fluoroform. This method allows for the rapid and repetitively robust production of [¹¹C]fluoroform in high yield and molar activity. The [¹¹C]fluoroform can be used directly in trifluoromethylation reactions or converted into the reactive copper(i) derivative ([¹¹C]CuCF₃), an effective reagent for the trifluoromethylation of aryl halides and heteroaryl halides.11,12

Fig. 1. Methods for ¹⁸F- and ¹¹C-trifluoromethylation.

Aminopyridines feature in the development of radiotracers for PET imaging. For example, [¹⁸F]3-fluoro-4-aminopyridine, ([¹⁸F]3-F-4AP) is a radiofluorinated derivative of 4-aminopyridine (4AP; dalfampridine), an FDA-approved drug for multiple sclerosis (MS). [¹⁸F]3-F-4AP is able to detect demyelinated lesions in rodent models.13 Like 4AP, [¹⁸F]3-F-4AP binds to the voltage-gated family of K⁺ channels (K_v1) that are exposed and overexpressed in demyelinated axons. The inverse correlation between myelin and K⁺ channel expression allows for a resultant PET signal that is directly proportional to demyelination.13–15 More recently, it has been reported that other 4AP derivatives, including 3-trifluoromethyl-4-aminopyridine (3-CF₃-4AP),16 can also bind to the Shaker K⁺ channel in vitro, which shares high homology with several mamalian K_v1 channels.13 Investigation of these compounds as potential PET radiotracers for demyelination is therefore warranted.

Given the potential of trifluoromethylated 4AP derivatives as radiotracers, we decided to investigate the use of [¹¹C]fluoroform for making [¹¹C]2-CF₃-4AP and [¹¹C]3-CF₃-4AP. Specifically, we examined the labelling of pyridine precursors bearing a halo group (iodo or bromo) at 2- or 3-position and either an ester, amide, or amino group at 4-position. The final aminopyridine radiotracers might then be accessed easily from these precursors after amide deprotection or through a Curtius–Yamada rearrangement to introduce the amino group, as precedented in the radiochemical synthesis of [¹⁸F]3-F-4AP.17 To demonstrate the practicality of these reactions for radiotracer synthesis, we finally produced [¹¹C]3-CF₃-4AP and performed a preliminary brain PET imaging experiment with this radiotracer in a healthy rhesus macaque.

Results and discussion

Improvement in production of [¹¹C]fluoroform

The method originally reported for producing [¹¹C]fluoroform used cyclotron-produced [¹¹C]methane as starting radioactive material. To demonstrate how [¹¹C]fluoroform can be accessed by sites that do not routinely produce [¹¹C]methane, such as ours, we generated [¹¹C]methane from cyclotron-produced [¹¹C]carbon dioxide by reduction with hydrogen over heated nickel in a GE Tracerlab FX MeI synthesizer. Subsequently, [¹¹C]methane was converted into [¹¹C]fluoroform by passage through a heated reactor containing CoF₃ in a manner resembling, but modified from, that described previously (Fig. 2A). The time from the end of bombardment (EOB) to the trapping of [¹¹C]methane was about 6 minutes and the time from releasing the [¹¹C]methane to collecting the final radioactive product was about 7 minutes (consistent with the prior report). The overall decay-corrected yield of [¹¹C]fluoroform starting from [¹¹C]carbon dioxide (52 ± 8%; n = 15) was comparable to that reported from [¹¹C]methane. An improvement in the radiochemical purity of the collected [¹¹C]fluoroform from about 80% to 94 ± 4% (n = 15), based on HPLC analysis (Fig. 2C), was accomplished by decreasing the helium carrier gas flow to 5 mL min^–1 (from 20 mL min^–1) when radioactivity passed in front of a detector located before the reactor (radiodetector 2; Fig. 2A and B, S1 and S2, ESI†), and by capping the output valve for 60 seconds. These modifications beneficially increase the pressure and contact time between the [¹¹C]methane and CoF₃.

Fig. 2. Production of [¹¹C]fluoroform. A) Layout of the [¹¹C]fluoroform apparatus (see ESI† for construction and operation details). B) Monitoring of [¹¹C]fluoroform production with radiodetectors 1–4 (see ESI† for details). C) A typical HPLC chromatogram for crude [¹¹C]fluoroform (HPLC conditions: XBridge™ 3.5 μm, 100 × 4.6 mm C-18 column (Waters); flow rate, 1 mL min^–1; mobile phase: 0.1% trifluoroacetic acid solution of H₂O–MeCN (95/5)).

¹¹C-Trifluoromethylation of pyridine substrates

For exploration of the radiotrifluoromethylation of pyridine derivatives, we converted the [¹¹C]fluoroform into [¹¹C]CuCF₃ as described by Haskali and Pike,3 based on the cupration method of Lishchynskyi et al.11 This conversion is essentially quantitative. Test labelling reactions in non-automated apparatus were then performed to examine this ¹¹C-trifluoromethylation strategy to produce [¹¹C]2- or [¹¹C]3-CF₃-4AP radiotracers from halopyridine precursors with either an ester, amide or amino group as the 4-substituent (Fig. 3 and S3–S13, ESI†). Although this study focused on the trifluoromethylation step, it is worth noting that following introduction of the [¹¹C]CF₃ group, the final 4-aminopyridine tracers can be obtained after amide deprotection or a Curtius–Yamada rearrangement to introduce the amine (Fig. 3A).

Fig. 3. Copper-mediated labelling of trifluoromethyl pyridines with [¹¹C]fluoroform. A) Strategies for the radiosyntheses of [¹¹C]2- and [¹¹C]3-trifluoromethyl-4-aminopyridine, namely, CuCF₃-mediated trifluoromethylation of: (I) 2- and 3-bromo/iodo-4-aminopyridine; (II) 2- and 3-bromo/iodo-4-(Boc-amino)pyridine followed by acid-catalysed deprotection of the amine; and (III) 2- and 3-bromo/iodo methyl isonicotinate followed by ester hydrolysis and Curtius–Yamada rearrangement. B) Reaction schemes and substrate scope for ¹¹C-trifluoromethylation. Reported values represent the radiochemical conversion (RCC) determined by radio-HPLC. RCC was calculated based on the initial amount of [¹¹C]fluoroform, not the initial amount of radioactivity. ^aRCC values represent the combination of protected and free amine products upon labelling reaction.

Effect of the 4-substituent

We examined the effect of the 4-substituent on labelling efficacy, by comparing 4-CO₂Me, 4-NHBoc, and 4-NH₂ substituted 3-iodopyridines and 2-iodopyridines in 10-minute reactions with [¹¹C]CuCF₃ (Fig. 3). A high radiochemical yield (93%) of the corresponding [¹¹C]3-trifluoromethyl pyridine ([¹¹C]5) was obtained from methyl 3-iodoisonicotinate (1) at 130 °C, and only a somewhat lower yield (74%) at lower reaction temperature (80 °C). The 2-iodo isomer (2) gave a remarkably high yield (92%) of the corresponding [¹¹C]trifluoromethyl compound ([¹¹C]6) at this temperature. t-Butyl (3-iodopyridin-4-yl)carbamate; (7) underwent ¹¹C-trifluoromethyation at 130 °C to afford [¹¹C]11 in high yield (89%). The 2-iodo isomer (8) also produced the corresponding ¹¹C-trifluoromethylated pyridine ([¹¹C]12) in high yield (97%) at 130 °C. Finally, even the 4-amino-3-iodo precursor 13 afforded a moderate yield (78%) of the corresponding [¹¹C]3-trifluoromethyl pyridine ([¹¹C]17) at 130 °C. Thus, these reactions showed only low sensitivity to the electronic nature of the 4-substituent, with yield only moderately suppressed by an electron-donating amino substituent relative to a mildly electron-withdrawing carboxymethyl substituent. To our knowledge a CuCF₃-mediated trifluoromethylation of a heteroaryl halide containing a free amino group has not been observed previously.

Effects of the halo substituent and substitution pattern

In terms of the nature of the leaving group, in every case an iodo leaving group gave a higher yield than a bromo leaving group, independent of the 4-substituent. Thus, the 4-CO₂Me 3-iodo-substituted pyridine (1) gave much higher yield than the bromo counterpart (2) at 130 °C (for [¹¹C]5, 93% vs. 37%). Higher yields of [¹¹C]5 and [¹¹C]6 were obtained from iodo precursors at 80 °C than from bromo precursors at 130 °C. This trend was also observed in the 4-NHBoc substituted pyridines, where at 130 °C much higher yields were obtained from the iodo precursors 7 and 9 (89% and 97%, respectively) than from the bromo precursors 8 and 10 (29% and 7%, respectively). Similarly, the iodo-aminopyridine 13 gave significantly higher yields of the respective ¹¹C-trifluoromethyl product ([¹¹C]17) than did the bromo counterparts 14 and 15.

Among the 4-subsituted bromo precursors, a higher yield of ¹¹C-trifluoromethyl product was obtained with bromo in 3-position than with bromo in 2-position. This concurs with the ortho effect17 described for the non-radioactive trifluoromethylation of aryl halides and heteroaryl halides with CuCF₃ derived from fluoroform. On macro non-radioactive scale, precursors with electron-withdrawing groups ortho to the halo group have been found to have faster reaction rates than precursors with electron-withdrawing groups in meta position.11 In this study, the ortho effect was not observed for iodopyridine precursors. The high yields from these precursors were more aligned to the higher electrophilicity of 2-iodopyridines relative to 3-iodopyridines.

Production of [¹¹C]3-trifluoromethyl-4-aminopyridine and PET imaging of rhesus macaque brain

Following this radiolabelling approach, production of the candidate radiotracer [¹¹C]3-trifluoromethyl-4-aminopyridine ([¹¹C]3-CF₃-4AP) using methods II and III was achieved with retention of very high molar activity (438 GBq μmol^–1, 11.8 Ci μmol^–1 at EOB, Fig. S14 and S15, ESI†).3 In view of these encouraging results, [¹¹C]3-CF₃-4AP was produced with >98% radiochemical purity (Fig. S15, ESI†), formulated in saline, and injected intravenously into one healthy rhesus monkey who underwent a 2-hour dynamic brain PET scan. [¹¹C]3-CF₃-4AP showed high brain penetration with fast uptake (∼6 SUV at 5 min) and washout (∼2 SUV at 30 min) (Fig. 4). Radioactivity distribution across the brain was relatively homogeneous and resembled that reported previously for [¹⁸F]3-F-4AP.12,18 In addition, preliminary radiometabolite analysis of plasma samples obtained after [¹¹C]3-CF₃-4AP administration indicates that the tracer is moderately stable in vivo (>40% remaining 1 h post injection, Fig. S16, ESI†). These initial imaging results suggest that [¹¹C]3-CF₃-4AP may be a good ¹¹C-labelled alternative to [¹⁸F]3-F-4AP and therefore merits further investigation regarding its specificity, selectivity, and sensitivity for imaging demyelinated lesions.

Fig. 4. Initial PET imaging of brain in a healthy monkey with [¹¹C]3-CF₃-4AP and comparison with [¹⁸F]3-F-4AP. A. Structural MRI (MPRAGE) and PET images of monkey brain after intravenous injection of [¹¹C]3-trifluoromethyl-4-aminopyridine or [¹⁸F]3-fluoro-4-aminopyridine. PET images correspond to 15–60 min summed images. Normalized radioactivity concentration (SUV) range: 0–3. Time–activity curves for cerebral cortex, thalamus, cerebellum, white matter, and striatum for B. [¹⁸F]3-F-4AP and C. [¹¹C]3-CF₃-4AP.

Experimental methods

Materials and chemicals

Chemicals, including halopyridine precursors and reference trifluoromethyl pyridines, were obtained from Sigma Aldrich, Combi-blocks, or Astatech. Sources of materials used to build the [¹¹C]fluoroform apparatus are given below.

General methods

Safety and regulatory approval

All experiments involving nonhuman primates were performed in accordance with the U.S. Department of Agriculture (USDA) Animal Welfare Act and Animal Welfare Regulations (Animal Care Blue Book), Code of Federal Regulations (CFR), Title 9, Chapter 1, Subchapter A, Part 2, Subpart C, §2.31. 2017. Experiments were approved by the Institutional Animal Care and Use Committee (IACUC) at the Massachusetts General Hospital (MGH). Experiments involving radioactive materials were performed by trained personnel following all relevant regulations.

Radioactivity measurement

All values are decay corrected consistent with the nomenclature rules for radiochemistry.19 Decay-correction was done to time of HPLC injection for RCCs and for total synthesis time for production of PET tracer.

Statistics

Grouped data are reported as mean ± SE.

[¹¹C]Fluoroform synthesis

For experiments at NIH, [¹¹C]fluoroform was prepared in the apparatus described by Haskali and Pike.3 A scheme of the apparatus used at MGH is shown in Fig. 2A. This was operated as follows. The V₈ line (exhaust) of an FX MeI module is connected to the 3-way valve V_I (3-way miniature size solenoid valve; universal operation, cat # 8320G041; ASCO Valves®) with 1/16-inch PTFE tubing. V_I-down is connected to V_II-centervia 1/16-inch PTFE tubing. Helium gas from a tank set to about 80 psi is connected to V_II-leftvia 1/8-inch stainless steel (SS) tubing. The outlet of the 3-way valve V_II-right is connected via 1/16-inch PTFE tubing to a Sicapent (phosphorus pentoxide on a neutral indicator resin) column to remove trace water. The outlet of the Sicapent column is connected via 1/16-inch SS tubing to a SS column (ID = 10 mm, OD = 12 mm) packed with CoF₃ (∼20 g). The CoF₃ reactor column is heated with a temperature-controlled furnace (Thermo Scientific Lindberg/Blue M). The end of the CoF₃ column is connected via 1/16-inch SS tubing to another 3-way valve, V_III-left. V_III-up connects via 1/8-inch PTFE tubing to a 2-way valve, V_IV, that controls the exhaust, and V_III-right connects to the ‘HF trap’ (coiled 1/8-inch SS tubing submerged in a MeCN-dry ice bath) and the [¹¹C]fluoroform trap (consisting of a 5 mL septum-capped V-vial containing 0.5 mL of DMF cooled to about – 40 °C in a MeCN-dry ice bath). Radiodetectors 1 (before V_I), 2 (before CoF₃ column), 3 (under ¹¹CHF₃ trap), and 4 (after V_IV) are set up to monitor the reaction (Fig. 2B). These radiodetectors are 7.6 mm² silicon PIN diode detectors connected to a DC amplifier (Model 101HDC-3 multi-channel radiation detector; Carroll & Ramsey Associates). The amplifier is connected to a multichannel high-speed analog to digital acquisition system (DI-4128; DATAQ Instruments,) which is connected to a PC via USB and read using WinDaq software.

Production of [¹¹C]fluoroform

Before each [¹¹C]fluoroform production, the CoF₃ column oven was set to 275 °C and the column conditioned for 15 to 20 min under helium flow (approx. 20 mL min^–1) by opening V_II and V_IV. [¹¹C]carbon dioxide was produced with a cyclotron via the ¹⁴N(p,α)¹¹C nuclear reaction on nitrogen in the presence of oxygen (0.5–1%). After ∼5 min of proton bombardment, the [¹¹C]carbon dioxide (∼25 GBq; 675 mCi) was transferred into a TRACERlab FX MeI module (GE) and mixed with hydrogen. The mixture was passed over nickel at 350 °C to generate [¹¹C]methane. Unreacted [¹¹C]carbon dioxide was removed by a packed column of solid NaOH. The [¹¹C]methane was then trapped in the ‘CH₄ trap’ at – 75 °C. When [¹¹C]methane activity reached a plateau (based on the FX MeI detector reading), V_I was switched to V_I-down to connect the output of the FX MeI system to the [¹¹C]fluoroform synthesis apparatus. Before receiving [¹¹C]methane in this apparatus, V_II was set to V_II-up position, which stopped the helium flow from the tank. On the FX MeI system V₃, V₅ and V₈ were opened, the helium flow rate on the FX MeI module was set at 60 mL min^–1 and the temperature of the ‘CH₄ trap’ was increased gradually to 75 °C. Once the reading of radiodetector 2 peaked, the helium flow of the FX MeI module was decreased to 5 mL min^–1, and V_IV (exhaust) was closed for 60 s (to increase the residence time of the [¹¹C]methane in the column). After 60 s, V_III (¹¹CHF₃ trap) was switched to V_III-right to collect the product. The helium flow was then increased to 20 mL min^–1 until the product was collected (about 2 min). The [¹¹C]fluoroform (8.73 ± 1.18 GBq, 236 ± 32 mCi; n = 15) was trapped in a MeCN-dry ice cooled DMF solution for HPLC analysis (Fig. 2C) and further reactions. The time from EOB to collection of [¹¹C]fluoroform was 13 ± 1 min.

Preparation of Cu complex

In a glovebox, stock solutions of KO^tBu (33.6 mg; 0.3 mmol) and CuBr (7.2 mg; 50 mmol) were each prepared in anhydrous DMF (1 mL). For each reaction, portions of these solutions (KO^tBu: 50 μL, 15 μmol; CuBr: 100 μL, 5 μmol) were mixed to produce CuO^tBu. The vial was septum-sealed and removed from the glovebox. [¹¹C]Fluoroform (370–740 MBq, 10–20 mCi) in DMF (50–100 μL) was added to the vial, mixed, and left at RT for 1 min. A solution (50 μL; 5 μmol) of Et₃N·3HF (0.1 mM) in DMF was then added. The mixture was mixed and allowed to stand at RT for another minute before addition of the substrate to be used for the ¹¹C-trifluoromethylation reaction.

Radiolabelling reactions and HPLC analysis

After production of the [¹¹C]CuCF₃ complex, a solution of precursor (7–14 mg, 45 μmol) in DMF (0.1 mL) was added to the reaction vial. The reaction mixture was then heated to 80 or 130 °C (depending on substrate) for 10 min. The reaction vial was removed from heat and allowed to reach RT. Product formation was analysed with HPLC. Labelled product was identified by comparison of its retention time with that of the authentic trifluoromethyl pyridine derivative. Radiochemical conversion (RCC) was determined by HPLC, from the ratio of labelled product peak signal (integration of peak on chromatogram) to the remaining [¹¹C]fluoroform peak signal, corrected for decay from time of injection. HPLC analysis of reaction mixtures was done on a Gemini® C-18 column (5 μm, 100 × 4.6 mm; Phenomenex) eluted at 1 mL min^–1 with the following basic mobile phase gradient: 0–5 min 5% MeCN, 95% 10 mM NH₄HCO₃; 5–6 min ramp to 60% MeCN, 40% 10 mM NH₄HCO₃; 6–11 min 60% MeCN, 40% 10 mM NH₄HCO₃, in order to resolve starting materials and radiolabelled products from the [¹¹C]fluoroform.

Radiochemical synthesis of [¹¹C]3-CF₃-4AP ([¹¹C]17) from 13

After [¹¹C]fluoroform (16.0 GBq, 433 mCi) had been produced, a portion 7.70 GBq (208 mCi) was used to prepare [¹¹C]CuCF₃, as described above. A solution of precursor 13 (10 mg, 45 μmol) in DMF (0.1 mL) was then added to the reaction vial. This reaction mixture was heated to 130 °C for 10 min. The reaction vial was then removed from heat and allowed to reach RT. An aqueous solution of TFA (0.1%; 0.5 mL) was then added and the reaction mixture further diluted with HPLC mobile phase (2.0 mL). The product was purified using HPLC on an XBridge™ C-18 column (5 μm, 250 × 10 mm; Waters) eluted at 3 mL min^–1 with EtOH : 10 mM NH₄HCO₃ (20 : 80 v/v). 104 MBq (2.8 mCi) of radiochemically pure [¹¹C]17 was collected (t_R = 27 min) (Fig. S15†). Radiochemical yield (RCY) of isolated product (decay corrected, total synthesis time: 43 min) was ca. 6%. The product was diluted with sterile saline to total volume of 10 mL.

PET imaging in a rhesus macaque

One male rhesus macaque (animal weight: 14.5 kg) underwent PET/CT imaging as well as structural MRI. The macaque's brain was imaged on a Discovery MI (GE Healthcare) PET/CT scanner. CT images were acquired before the PET acquisition for attenuation correction. Dynamic PET data were then acquired for 120 min after a 3 min bolus injection of [¹¹C]3-CF₃-4AP (71.4 MBq, 1.93 mCi) followed by a 3 min saline flush. PET data were reconstructed using a 3D iterative reconstruction algorithm with 3 iterations and 34 subsets and were corrected for attenuation, scatter, random coincidences, dead-time and normalization. Dynamic images were framed into time-bins of 6 × 10, 8 × 15, 6 × 30, 8 × 60, 8 × 120 and 18 × 300 s. In a separate imaging session, a T1-weighted magnetization-prepared rapid gradient-echo (MPRAGE) was also acquired on a 3T Biograph mMR (Siemens Medical Systems) MRI system for anatomical reference.

Dynamic PET images were rigidly co-registered to the MR images. The MPRAGE was warped into an MR rhesus template space20 and the transformation matrix applied to the PET images. Rhesus atlases defined in the template space were then transformed into the original PET space for extraction of time–activity curves (TACs) in cerebral cortex, thalamus, white matter and striatum.21 Radioactivity was converted into standardized uptake value (SUV), which normalizes for radiotracer dose and subject body weight. All image processing was performed in MATLAB and used FSL for registration purposes.22

Conclusions

In summary, the apparatus reported by Haskali and Pike was successfully modified to produce [¹¹C]fluoroform from [¹¹C]carbon dioxide using a commercially available GE Tracerlab FX MeI module and an in-house built reactor. An appreciable improvement in the radiochemical purity of the [¹¹C]fluoroform was achieved by reducing the flow rate of [¹¹C]methane through the CoF₃ reactor. The [¹¹C]fluoroform was then readily converted into [¹¹C]CuCF₃ and tested in reactions with several [¹¹C]2-CF3-4AP and [¹¹C]3-CF₃-4AP precursors. [¹¹C]CuCF₃ labelled bromo- and iodo-substituted pyridine precursors containing electron-withdrawing (CO₂Me) and electron-donating groups (NHBoc and NH₂) in a wide range of yields (5–97%) in a short reaction time (10 minutes) under mild conditions (80 to 130 °C). Using one of the identified precursors (3-iodo-4-aminopyridine), we were able to produce a candidate radiotracer with high molar activity and high radiochemical purity for an initial PET imaging study in a healthy monkey. This radiotracer showed good brain penetration and suitable kinetics for a ¹¹C-labelled compound. This study illustrates the versatility of [¹¹C]CuCF₃ for labelling 4-subsituted pyridines with a [¹¹C]trifluoromethyl group. [¹¹C]3-CF₃-4AP is a potentially useful tracer for demyelinating diseases, warranting further investigation on the specificity and sensitivity of the radioligand to demyelinating brain injuries.

Conflicts of interest

The University of Chicago has obtained patents for the use of compounds described in this work in PET imaging where PB is listed as inventor. BYY, ST, and VWP are named as co-inventors on a patent application concerning [¹¹C]fluoroform synthesis.