In Vitro and In Vivo Evaluation of a 18F-Labeled High Affinity NOTA Conjugated Bombesin Antagonist as a PET Ligand for GRPR-Targeted Tumor Imaging

Expression of the gastrin-releasing peptide receptor (GRPR) in prostate cancer suggests that this receptor can be used as a potential molecular target to visualize and treat these tumors. We have previously investigated an antagonist analog of bombesin (D-Phe-Gln-Trp-Ala-Val-Gly-His-Sta-Leu-NH₂, RM26) conjugated to 1,4,7-triazacyclononane-N,N'',N''''-triacetic acid (NOTA) via a diethylene glycol (PEG₂) spacer (NOTA-P2-RM26) labeled with ⁶⁸Ga and ¹¹¹In. We found that this conjugate has favorable properties for in vivo imaging of GRPR-expression. The focus of this study was to develop a ¹⁸F-labelled PET agent to visualize GRPR. NOTA-P2-RM26 was labeled with ¹⁸F using aluminum-fluoride chelation. Stability, in vitro binding specificity and cellular processing tests were performed. The inhibition efficiency (IC₅₀) of the [^natF]AlF-NOTA-P2-RM26 was compared to that of the ^natGa-loaded peptide using ¹²⁵I-Tyr⁴-BBN as the displacement radioligand. The pharmacokinetics and in vivo binding specificity of the compound were studied. NOTA-P2-RM26 was labeled with ¹⁸F within 1 h (60-65% decay corrected radiochemical yield, 55 GBq/µmol). The radiopeptide was stable in murine serum and showed high specific binding to PC-3 cells. [^natF]AlF-NOTA-P2-RM26 showed a low nanomolar inhibition efficiency (IC₅₀=4.4±0.8 nM). The internalization rate of the tracer was low. Less than 14% of the cell-bound radioactivity was internalized after 4 h. The biodistribution of [¹⁸F]AlF-NOTA-P2-RM26 demonstrated rapid blood clearance, low liver uptake and low kidney retention. The tumor uptake at 3 h p.i. was 5.5±0.7 %ID/g, and the tumor-to-blood, -muscle and -bone ratios were 87±42, 159±47, 38±16, respectively. The uptake in tumors, pancreas and other GRPR-expressing organs was significantly reduced when excess amount of non-labeled peptide was co-injected. The low uptake in bone suggests a high in vivo stability of the Al-F bond. High contrast PET image was obtained 3 h p.i. The initial biological results suggest that [¹⁸F]AlF-NOTA-P2-RM26 is a promising candidate for PET imaging of GRPR in vivo.     

Labeling a TCO-functionalized single domain antibody fragment with 18F via inverse electron demand Diels Alder cycloaddition using a fluoronicotinyl moiety-bearing tetrazine derivative

Single domain antibody fragments (sdAbs) exhibit a rapid tumor uptake and fast blood clearance amenable for labeling with ¹⁸F (t_½ = 110 min) but suffer from high kidney accumulation. Previously, we developed a method for ¹⁸F-labeling of sdAbs via trans-cyclooctene (TCO)-tetrazine (Tz) inverse electron demand Diel’s Alder cycloaddition reaction (IEDDAR) that incorporated a renal brush border enzyme (RBBE)-cleavable linker. Although >15 fold reduction in kidney activity levels was achieved, tumor uptake was compromised. Here we investigate whether replacing the [¹⁸F]AlF-NOTA moiety with [¹⁸F]fluoronicotinyl would rectify this problem. Anti-HER2 sdAb 5F7 was first derivatized with a TCO-containing agent that included the RBBE-cleavable linker GlyLys (GK) and a PEG chain, and then subjected to IEDDAR with 6-[¹⁸F]fluoronicotinyl-PEG₄-methyltetrazine to provide [¹⁸F]FN-PEG₄-Tz-TCO-GK-PEG₄-5F7 ([¹⁸F]FN-GK-5F7). For comparisons, a control lacking GK linker and 5F7 labeled using residualizing N-succinimidyl 3-guanidinomethyl-5-[¹²⁵I]iodobenzoate (iso-[¹²⁵I]SGMIB) also were synthesized. Radiochemical purity, affinity (K_D) and immunoreactive fraction of [¹⁸F]FN-GK-5F7 were 99%, 5.4 ± 0.7 nM and 72.5 ± 4.3%, respectively. Tumor uptake of [¹⁸F]FN-GK-5F7 in athymic mice bearing subcutaneous SKOV3 xenografts (3.7 ± 1.2% ID/g and 3.4 ± 1.0% ID/g at 1 h and 3 h, respectively) was 2- to 3-fold lower than for co-injected iso-[¹²⁵I]SGMIB-5F7 (6.9 ± 1.9 %ID/g and 8.7 ± 3.0 %ID/g). However, due to its 6-fold lower kidney activity levels, tumor-to-kidney ratios for [¹⁸F]FN-GK-5F7 were 3–4 times higher than those for co-injected iso-[¹²⁵I]SGMIB-5F7 as well as those observed for the ¹⁸F conjugate lacking the RBBE-cleavable linker. Micro-PET/CT imaging of [¹⁸F]FN-GK-5F7 in mice with SKOV-3 subcutaneous xenografts clearly delineated tumor as early as 1 h with minimal activity in the kidneys; however, there was considerable activity in gallbladder and intestines. Although the tumor uptake of [¹⁸F]FN-GK-5F7 was unexpectedly disappointing, incorporating an alternative RBBE-cleavable linker into this labeling strategy may ameliorate this problem.     

Labeling proteins with fluorine-18 using N-succinimidyl 4-[18F]fluorobenzoate

Two methods were investigated for the no-carrier-added synthesis of N-succinimidyl 4-[¹⁸F]fluorobenzoate (S[¹⁸F]FB). The first, an attempted nucleophilic aromatic substitution by [¹⁸F]fluoride on N-succinimidyl 4-nitrobenzoate was unsuccessful. The second method involved three steps; [¹⁸F]fluoride for trimethylammonium substitution on 4-formyl-N,N,N-trimethylanilinium triflate, oxidation to 4-[¹⁸F]fluorobenzoic acid, followed by reaction with N-hydroxysuccinimide and dicyclohexylcarbodiimide to form S[¹⁸F]FB. Total synthesis and purification time was 100 min and the overall radiochemical yield was 25% (decay corrected). A monoclonal antibody F(ab′)₂ fragment could be labeled in 40–60% yield by reaction with S[¹⁸F]FB for 15–20 min. The tissue distribution in normal mice and in vitro tumor binding of the antibody F(ab′)₂ labeled by reaction with S[¹⁸F]FB were comparable to those observed for the fragment after radioiodination using N-succinimidyl 4-[¹²⁵I]iodobenzoate.     

Synthesis of a fluorinated fatty acid,dl-erythro-9,10-[18F]difluoropalmitic acid,and biodistribution studies in rats

9,10-Difluoropalmitic acid (DFPA) labeled with the cyclotron produced, positron emitting radionuclide ¹⁸F has been synthesized as a potential analogue of 9,10-[³H]palmitic acid, a fatty acid which has been used to study lipid metabolism in rat brain and pituitary. [¹⁸F]DFPA was prepared by the direct and stereoselective addition of [¹⁸F]F₂ to the double bond of cis-9,10-palmitoleic acid. The fluorination was carried out in FCCl₃ at −70 °C using a low concentration of F₂ (0.5%) in neon. [¹⁸F]DFPA has been obtained in radiochemical yields of 12–16% from end-of-bombardment (EOB) in approx. 2.5 h. Chemical and radiochemical purity exceeded 95%, and specific activities calculated to EOB ranged from 500 to 1000 mCi/mmol. [¹⁸F]DFPA crosses the blood-brain barrier and is incorporated into rat brain at about twice the level of that of 9,10-[³H]palmitic acid. The synthesis of [¹⁸F]DFPA permits us to study the biological disposition and metabolism of a vicinal-difluoro fatty acid.     

Fluorine-18 labeling of an anti-HER2 VHH using a residualizing prosthetic group via a strain-promoted click reaction: Chemistry and preliminary evaluation

In a previous study, we evaluated a HER2-specific single domain antibody fragment (sdAb) 2Rs15d labeled with ¹⁸F via conjugation of a residualizing prosthetic agent that was synthesized by copper-catalyzed azide-alkyne cycloaddition (CuAAC). In order to potentially increase overall efficiency and decrease the time required for labeling, we now investigate the use of a strain-promoted azide-alkyne cycloaddition (SPAAC) between the 2Rs15d sdAb, which had been pre-derivatized with an azide-containing residualizing moiety, and an ¹⁸F-labeled aza-dibenzocyclooctyne derivative. The HER2-targeted sdAb 2Rs15d and a nonspecific sdAb R3B23 were pre-conjugated with a moiety containing both azide- and guanidine functionalities. The thus derivatized sdAbs were radiolabeled with ¹⁸F using an ¹⁸F-labeled aza-dibenzocyclooctyne derivative ([¹⁸F]F-ADIBO) via SPAAC, generating the desired conjugate ([¹⁸F]RL-II-sdAb). For comparison, unmodified 2Rs15d was labeled with N-succinimidyl 4-guanidinomethyl-3-[¹²⁵I]iodobenzoate ([¹²⁵I]SGMIB), the prototypical residualizing agent for radioiodination. Radiochemical purity (RCP), immunoreactive fraction (IRF), HER2-binding affinity and cellular uptake of [¹⁸F]RL-II-2Rs15d were assessed in vitro. Paired label biodistribution of [¹⁸F]RL-II-2Rs15d and [¹²⁵I]SGMIB-2Rs15d, and microPET/CT imaging of [¹⁸F]RL-II-2Rs15d and the [¹⁸F]RL-II-R3B23 control sdAb were performed in nude mice bearing HER2-expressing SKOV-3 xenografts. A radiochemical yield of 23.9?±?6.9% (n?=?8) was achieved for the SPAAC reaction between [¹⁸F]F-ADIBO and azide-modified 2Rs15d and the RCP of the labeled sdAb was >95%. The affinity (K_d) and IRF for the binding of [¹⁸F]RL-II-2Rs15d to HER2 were 5.6?±?1.3?nM and 73.1?±?22.5% (n?=?3), respectively. The specific uptake of [¹⁸F]RL-II-2Rs15d by HER2-expressing BT474M1 breast carcinoma cells in vitro was 14–17% of the input dose at 1, 2, and 4?h, slightly higher than seen for co-incubated [¹²⁵I]SGMIB-2Rs15d. The uptake of [¹⁸F]RL-II-2Rs15d in SKOV-3 xenografts at 1?h and 2?h p.i. were 5.54?±?0.77% ID/g and 6.42?±?1.70% ID/g, respectively, slightly higher than those for co-administered [¹²⁵I]SGMIB-2Rs15d (4.80?±?0.78% ID/g and 4.78?±?1.39% ID/g). MicroPET/CT imaging with [¹⁸F]RL-II-2Rs15d at 1–3?h p.i. clearly delineated SKOV-3 tumors while no significant accumulation of activity in tumor was seen for [¹⁸F]RL-II-R3B23. With the exception of kidneys, normal tissue levels for [¹⁸F]RL-II-2Rs15d were low and cleared rapidly. To our knowledge, this is the first time SPAAC method has been used to label an sdAb with ¹⁸F, especially with residualizing functionality.     

Mechanistic Positron Emission Tomography Studies of 6-[18F]Fluorodopamine in Living Baboon Heart: Selective Imaging and Control of Radiotracer Metabolism Using the Deuterium Isotope Effect

Abstract: Mechanistic positron emission tomography (PET) studies using the deuterium isotope effect and specific pharmacological intervention were undertaken to examine the behavior of 6-[¹⁸F]fluorodopamine (6-[¹⁸F]FDA; 1 ) and (?)-6-[¹⁸F]fluoronorepinephrine {(?)-6-[¹⁸F]FNE; 2 } in the baboon heart. Two regiospecifically deuterated derivatives of 6-[¹⁸F]FDA [α,α-D₂(3 ) and β,β-D₂ (4 )] were used to assess the contributions of monoamine oxidase (MAO) and dopamine β-hydroxylase, respectively, to the clearance kinetics of 6-[¹⁸F]FDA. Compound 3 showed a reduced rate of clearance, consistent with MAO-catalyzed cleavage of the α C-D bond, whereas compound 4 showed no change, indicating that cleavage of the β C-D bond is not a rate-limiting step. Pretreatment with pargyline, an MAO inhibitor, also decreased the rate of clearance. Desipramine and tomoxetine [norepinephrine (NE) uptake inhibitors], but not GBR-12909 (a dopamine uptake inhibitor), blocked the uptake of both (?)-6-[¹⁸F]FNE and 6-[¹⁸F]FDA, with (?)-6-[¹⁸F]FNE showing a higher degree of blockade. Chiral HPLC demonstrated that 6-[¹⁸F]FDA is stereoselectively converted to (?)-6-[¹⁸F]FNE in vivo in the rat heart. These studies demonstrate that (a) the more rapid clearance of 6-[¹⁸F]FDA relative to (?)-6-[¹⁸F]FNE can be largely accounted for by metabolism by MAO; (b) selective deuterium substitution can be used to protect a radiotracer from metabolism in vivo and to favor a particular pathway; (c) 6-[¹⁸F]FDA and (?)-6-[¹⁸F]FNE share the NE transporter; (d) 6-[¹⁸F]FDA is stereoselectively converted to (?)-6-[¹⁸F]FNE in vivo; and (e) the profile of radioactivity in the heart for 6-[¹⁸F]FDA is complex, probably including labeled metabolites as well as neuronal and nonneuronal uptake.     

Synthesis and in vitro evaluation of three novel radiotracers for imaging of metabotropic glutamate receptor subtype 2 in rat brain

The purpose of this study was to develop three new radiotracers, 1-(cyclopropylmethyl)-4-([¹¹C/¹⁸F]substituted-phenyl)piperidin-1-yl-2-oxo-1,2-dihydropyridine-3-carbonitrile ([¹¹C]1, [¹¹C]2, and [¹⁸F]4), and to examine their specific bindings with metabotropic glutamate receptor subtype 2 (mGluR2) in rat brain sections by using in vitro autoradiography. These compounds were found to possess potent in vitro binding affinities (K_i: 8.0–34.1 nM) for mGluR2 in rat brain homogenate. [¹¹C]1, [¹¹C]2, and [¹⁸F]4 were synthesized by [¹¹C/¹⁸F]alkylation of the corresponding phenol precursors with [¹¹C]methyl iodide or [¹⁸F]fluoroethyl bromide with >98% radiochemical purity and 80–130 GBq/μmol specific activity at the end of synthesis. In vitro autoradiography indicated that these radiotracers showed heterogeneous specific bindings in mGluR2-rich brain regions, such as the cerebral cortex, striatum, hippocampus, and granular layer of the cerebellum.     

Synthesis and preliminary biological evaluation of a novel P2X7R radioligand [18F]IUR-1601

The reference standard IUR-1601 ((S)-N-(2-chloro-3-(trifluoromethyl)benzyl)-1-(2-fluoroethyl)-5-oxopyrrolidine-2-carboxamide) was synthesized from tert-butyl (S)-5-oxopyrrolidine-2-carboxylate, fluoroethylbromide, and 2-chloro-3-(trifluoromethyl)benzylamine with overall chemical yield 12% in three steps. The target tracer [¹⁸F]IUR-1601 ((S)-N-(2-chloro-3-(trifluoromethyl)benzyl)-1-(2-[¹⁸F]fluoroethyl)-5-oxopyrrolidine-2-carboxamide) was synthesized from desmethyl-GSK1482160 with 2-[¹⁸F]fluoroethyl tosylate, prepared from 1,2-ethylene glycol-bis-tosylate and K[¹⁸F]F/Kryptofix2.2.2, in two steps and isolated by HPLC combined with SPE in 1–3% decay corrected radiochemical yield. The radiochemical purity was >99%, and the molar activity at end of bombardment (EOB) was 74–370?GBq/μmol. The potency of IUR-1601 in comparison with GSK1482160 was determined by a radioligand competitive binding assay using [¹¹C]GSK1482160, and the binding affinity K_i values for IUR-1601 and GSK1482160 are 4.31 and 5.14?nM, respectively.     

Triaryl (Z)-olefins suitable for radiolabeling with carbon-11 or fluorine-18 radionuclides for positron emission tomography imaging of cyclooxygenase-2 expression in pathological disease

A group of (Z)-1,1-diphenyl-2-(4-methylsulfonylphenyl)alk-1-enes were synthesized using methodologies that will allow incorporation of a [¹¹C]OCH₃ substituent at the para-position of the C-1 phenyl ring, a [¹¹C]SO₂CH₃ substituent at the para-position of the C-2 phenyl ring, a [¹⁸F]OCH₂CH₂F substituent at the para-position of the C-1 phenyl ring, and a [¹⁸F]CH₂CH₂F substituent at the C-2 position of the olefinic bond. The [¹¹C] and [¹⁸F] radiotracers are designed as potential radiopharmaceuticals to image cyclooxygenase-2 (COX-2) expression in any organ where COX-2 is upregulated. The COX-1/COX-2 inhibition data acquired suggest that compounds having a [¹¹C]OMe or [¹⁸F]OCH₂CH₂F substituent at the para-position of the C-1 phenyl ring may be more suitable for imaging COX-2 expression in view of their ability to exclusively inhibit the COX-2 isozyme.     

Tumor diagnosis by pet: potential of seven tracers examined in five experimental tumors including an artificial metastasis model

The potential of seven tracers for the metabolic imaging of tumors by positron emission tomography was studied using five experimental tumor models. The tracers examined were 2-deoxy-2-[¹⁸F]fluoro-d-glucose ([¹⁸F]FDG), 2-deoxy-2-[¹⁸F]fluoro-d-galactose (2-[¹⁸F]FdGal) and 2-deoxy-2-[¹⁸F]fluoro-l-fucose (2-[¹⁸F]FdFuc) for investigating energy metabolism. l-[methyl-¹¹C]Methionine ([¹¹C]Met) and 6-[¹⁸F]fluoro-l-fucose (6-[¹⁸F]FFuc) were used for assessing protein and glycoprotein synthesis, while [³H]thymidine ([³ H]Thd) and 2-deoxy-5′-[¹⁸F]fluorouridine ([¹⁸F]FdUrd) were used to investigate nucleic acid metabolism. The highest mean uptake by the five different tumors was found for [³H]Thd, followed in order by [¹⁸F]FDG, [¹¹C]Met, 2-[¹⁸F]FdGal, [¹⁸F]FdUrd, 2-[¹⁸F]FdFuc and 6-[¹⁸F]FFuc. The tumor-to-tissue uptake ratios indicated that the nucleosides, [¹¹C]Met and 6-[¹⁸F]FFuc were better tracers in the brain region. All the tracers except for the fucose analogs were suitable for the thoracic region, while [¹¹C]Thd and [¹⁸ F]FDG were superior in the abdominal region. In comparison with the primary tumor model of Lewis lung carcinoma (3LL), [³H]Thd uptake in the artificial metastatic 3LL model showed the maximum enhancement, followed by [¹⁸F]FDG, [¹¹C]Met and the other tracers. The [¹⁸F]FDG uptake correlated with the [³H]Thd uptake. [¹⁸F]FdUrd, 6-[¹⁸F]FFuc and 2-[¹⁸F]FdGal could be used for distinguishing different types of tumors. The combined use of these radiotracers can possibly allow the assessment of tumor metabolism, and this indicates the viability of tumors.     

Efficient microwave-assisted direct radiosynthesis of [18F]PR04.MZ and [18F]LBT999: Selective dopamine transporter ligands for quantitative molecular imaging by means of PET

PR04.MZ 8-(4-fluoro-but-2-ynyl)-3-p-tolyl-8-aza-bicyclo[3.2.1]octane-2-carboxylic acid methyl ester (1) and LBT999 8-((E)-4-fluoro-but-2-enyl)-3b-p-tolyl-8-aza-bicyclo[3.2.1]octane-2β-carboxylic acid methyl ester (2) are selective dopamine reuptake inhibitors, derived from cocaine. Compounds 1 and 2 were labelled with fluorine-18 at their terminally fluorinated N-substituents employing microwave enhanced direct nucleophilic fluorination. K[¹⁸F]F^? Kryptofix^®222 cryptate, tetrabutyl ammonium [¹⁸F]fluoride and caesium [¹⁸F]fluoride were compared as fluoride sources under conventional and microwave enhanced conditions. Fluorination yields were remarkably increased under microwave irradiation for all three fluoride salts. Radiochemically pure (>98%) [¹⁸F]PR04.MZ (0.95–1.09 GBq, 42–135 GBq/μmol) was obtained within 34–40 min starting from 3.0 GBq [¹⁸F]fluoride ion in 32–36% non-decay-corrected overall yield using K[¹⁸F]F^?Kryptofix^®222 cryptate in MeCN.     

Non-β-oxidizable ω-[18F]fluoro long chain fatty acid analogs show cytochrome P-450-mediated defluorination: Implications for the design of PET tracers of myocardial fatty acid utilization

The nature of the in vivo defluorination of non-β-oxidizable no-carrier-added ω-[¹⁸F]fluoro long chain fatty acid (LCFA) analogs was studied with the aim of developing PET tracers of LCFA utilization. Extensive defluorination of 15-[¹⁸F]fluoro-3-thia-pentadecanoic acid (FTPA) in mouse was evidenced by radioactivity uptake by bone. [¹⁸F]Fluoride in the blood was verified analytically. Incubations of FTPA in rat-liver homogenates and subcellular fractions thereof showed a strong defluorination process in microsomes which was O₂- and NADPH-dependent. In contrast, defluorination of FTPA was relatively slow in Langendorff perfused rat heart. High bone uptake in mouse was also observed with 14-[¹⁸F]fluoro-13, 13-dimethyl-3-thia-tetradecanoic acid, where gem-dimethyl substitution precludes direct elimination of H¹⁸F. These data indicate that the defluorination of non-β-oxidizable ω-[¹⁸F]fluoro LCFA analogs is primarily governed by cytochrome P-450-mediated ω-oxidation.Therefore, labeling at the (ω-3) carbon was proposed to provide a more stabile ¹⁸F-label. Defluorination of the (ω-3)-labeled 13 (R,S)-[¹⁸F]fluoro-3-thia-hexadecanoic acid was lower than that of FTPA in mouse and was independent of O₂ and NADPH in vitro. Thus, (ω-3) labeling with ¹⁸F is preferable to ω labeling of non-β-oxidizable LCFA analogs.     

Synthesis and in vitro evaluation of 18F labeled tyrosine derivatives as potential positron emission tomography (PET) imaging agents

Three new ¹⁸F labeled fluoroalkyl tyrosine derivatives, O-(2-[¹⁸F]fluoroethyl)-α-methyltyrosine (FEMT, [¹⁸F]2), O-(2-[¹⁸F]fluoroethyl)-2-l-azatyrosine (FEAT, [¹⁸F]3), O-(2-[¹⁸F]fluoroethyl)-l-tyrosineamide (FETA, [¹⁸F]4) have been synthesized and radiofluorinated with 5–34% decay-corrected yield. In vitro studies were carried out in U-138 MG human glioblastoma. Cellular uptake of new tracers was compared to clinically utilized imaging agent O-(2-[¹⁸F]fluoroethyl)-l-tyrosine (FET, [¹⁸F]1). The uptake of tracers followed the order of FET ([¹⁸F]1) > FEAT([¹⁸F]3) > FEMT ([¹⁸F]2) ≈ FETA ([¹⁸F]4).     

Systematic comparison of two novel,thiol-reactive prosthetic groups for <Superscript>18</Superscript>F labeling of peptides and proteins with the acylation agent succinimidyl-4-[<Superscript>18</Superscript>F]fluorobenzoate ([<Superscript>18</Superscript>F]SFB)

A systematic comparison of 4-[¹⁸F]fluorobenzaldehyde-O-(2-{2-[2-(pyrrol-2,5-dione-1-yl)ethoxy]-ethoxy}-ethyl)oxime ([¹⁸F]FBOM) and 4-[¹⁸F]fluorobenzaldehyde-O-[6-(2,5-dioxo-2,5-dihydro-pyrrol-1-yl)-hexyl]oxime ([¹⁸F]FBAM) as prosthetic groups for the mild and efficient ¹⁸F labeling of cysteine-containing peptides and proteins with the amine-group reactive acylation agent, succinimidyl-4-[¹⁸F]fluorobenzoate ([¹⁸F]SFB), is described. All three prosthetic groups were prepared in a remotely controlled synthesis module. Synthesis of [¹⁸F]FBOM and [¹⁸F]FBAM was accomplished via oxime formation through reaction of appropriate aminooxy-functionalized labeling precursors with 4-[¹⁸F]fluorobenzaldehyde. The obtained radiochemical yields were 19% ([¹⁸F]FBOM) and 29% ([¹⁸F]FBAM), respectively. Radiolabeling involving [¹⁸F]FBAM and [¹⁸F]FBOM was exemplified by the reaction with cysteine-containing tripeptide glutathione (GSH), a cysteine-containing dimeric neurotensin derivative, and human native low-density lipoprotein (nLDL) as model compounds. Radiolabeling with the acylation agent [¹⁸F]SFB was carried out using a dimeric neurotensin derivative and nLDL. Both thiol-group reactive prosthetic groups show significantly better labeling efficiencies for the peptides in comparison with the acylation agent [¹⁸F]SFB. The obtained results demonstrate that [¹⁸F]FBOM is especially suited for the labeling of hydrophilic cysteine-containing peptides, whereas [¹⁸F]FBAM shows superior labeling performance for higher molecular weight compounds as exemplified for nLDL apolipoprotein constituents. However, the acylation agent [¹⁸F]SFB is the preferred prosthetic group for labeling nLDL under physiological conditions.     

Towards tropomyosin-related kinase B (TrkB) receptor ligands for brain imaging with PET: Radiosynthesis and evaluation of 2-(4-[18F]fluorophenyl)-7,8-dihydroxy-4H-chromen-4-one and 2-(4-([N-methyl-11C]-dimethylamino)phenyl)-7,8-dihydroxy-4H-chromen-4-one

The interaction of tropomyosin-related kinase B (TrkB) with the cognate ligand brain-derived neurotrophic factor (BDNF) mediates fundamental pathways in the development of the nervous system. TrkB signaling alterations are linked to numerous neurodegenerative diseases and conditions. Herein we report the synthesis, biological evaluation and radiosynthesis of the first TrkB radioligands based on the recently identified 7,8-dihydroxyflavone chemotype. 2-(4-[¹⁸F]fluorophenyl)-7,8-dihydroxy-4H-chromen-4-one ([¹⁸F]10b) was synthesized in high radiochemical yields via an efficient S_NAr radiofluorination involving a para-Michael acceptor substituted aryl followed by BBr₃-promoted double demethylation. Selective N-[¹¹C]methylation afforded 2-(4-([N-methyl-¹¹C]-dimethylamino)phenyl)-7,8-dihydroxy-4H-chromen-4-one ([¹¹C]10c) from the fully deprotected catechol-bearing normethyl precursor 13 with [¹¹C]MeOTf. In vitro autoradiography of [¹⁸F]10b with transverse rat brain sections revealed high specific binding in the cortex, striatum, hippocampus and thalamus in accordance with expected TrkB distribution. Blockade experiments with both 7,8-dihydroxyflavone (1a) and TrkB cognate ligand, BDNF, led to decreases of 80% and 85% of radioligand binding strongly supporting the hypothesis that 7,8-dihydroxyflavones exert their effect on TrkB phosphorylation via direct TrkB extracellular domain (ECD) binding. Positron emission tomography (PET) studies revealed that [¹⁸F]10b and [¹¹C]10c brain uptake is minimal and that they are rapidly eliminated from the plasma (effective plasma half-life 5–10 min) via hepatic secretion. Nevertheless, the high specific binding and TrkB specificity derived from in vitro experiments suggests that the 7,8-disubstituted flavone chemotype represents a promising scaffold for the development of TrkB radiotracers for PET.     

New approach for the synthesis of [F]fluoroethyltyrosine for cancer imaging: Simple, fast, and high yielding automated synthesis

O-(2-[¹⁸F]fluoroethyl)-l-tyrosine ([¹⁸F]FET) is one of the first ¹⁸F-labeled amino acids for imaging amino acid metabolism in tumors. This tracer overcomes the disadvantages of [¹⁸F]fluorodeoxyglucose, [¹⁸F]FDG, and [¹¹C]methionine, [¹¹C]MET. Nevertheless, the various synthetic methods providing ¹⁸F[FET] exhibit a big disadvantage concerning the necessity of two purification steps during the synthesis including HPLC purification, which causes difficulties in the automation, moderate yields, and long synthesis times >60 min.A new approach for the synthesis of [¹⁸F]FET is developed starting from 2-bromoethyl triflate as precursor. After optimization of the synthesis parameters including the distillation step of [¹⁸F]-FCH₂CH₂Br combined with the final purification of [¹⁸F]FET using a simple solid phase extraction instead of an HPLC run the synthesis [¹⁸F]FET could be significantly simplified, shortened, and improved. The radiochemical yield (RCY) was about 45% (not decay corrected and calculated relative to [¹⁸F]F⁻ activity that was delivered from the cyclotron). Synthesis time was only 35 min from the end of bombardment (EOB) and the radiochemical purity was >99% at the end of synthesis (EOS). Thus, this simplified synthesis for [¹⁸F]FET offers a very good option for routine clinical use.     

Synthesis and evaluation of 6-(3-[18F]fluoro-2-hydroxypropyl)-substituted 2-pyridylbenzothiophenes and 2-pyridylbenzothiazoles as potential PET tracers for imaging Aβ plaques

3-[¹⁸F]Fluoro-2-hydroxypropyl substituted compounds were synthesized and evaluated as novel ¹⁸F-labeled PET tracers for imaging Aβ plaque in a living brain. All compounds exhibited high binding affinities toward the synthetic Aβ_1–42 aggregate and/or Alzheimer’s disease brain homogenate. In the microPET study with normal mice, the 3-[¹⁸F]fluoro-2-hydroxypropyl substituted compounds resulted in fast brain washout by reducing the lipophilicities of the compounds. Intriguingly, (S)-configured PET tracers, (S)-[¹⁸F]1b and (S)-[¹⁸F]1c, exhibited a 2.8 and 4.0-fold faster brain washout rate at a peak/30 min in the mouse brain than the corresponding (R)-configured PET tracers despite there being no meaningful difference in binding affinities toward Aβ plaque. A further evaluation of (S)-[¹⁸F]1c with healthy rhesus monkeys also revealed excellent clearance from the frontal cortex with ratios of 7.0, 16.0, 30.0 and 49.0 at a peak/30, 60, 90, and 120 min, respectively. These results suggest that (S)-[¹⁸F]1c may be a potential PET tracer for imaging Aβ plaque in a living brain.     

Triaryl (Z)-olefins suitable for radiolabeling with iodine-124 or fluorine-18 radionuclides for positron emission tomography imaging of estrogen positive breast tumors

A group of (Z)-1,2-diphenyl-1-[4-[2-(4-methylpiperazin-1-yl)ethoxy]phenyl]but-1-enes were synthesized using methodologies that will allow incorporation of a [¹²⁴I]iodine substituent at the para-position of either the C-1 phenyl ring or the C-2 phenyl ring, or a [¹⁸F]OCH₂CH₂F substituent at the para-position of the C-2 phenyl ring. These [¹²⁴I] and [¹⁸F] radiotracers are designed as potential radiopharmaceuticals to image estrogen positive breast tumors using positron emission tomography (PET).     

Biodistribution of N-alkyl and N-fluoroalkyl derivatives of spiroperidol; Radiopharmaceuticals for PET studies of dopamine receptors

There is great interest in the application of positron labeled ligands to map the dopamine receptor in vivo. A series of fluorine-18-labeled N-alkyl and N-fluoroalkyl spiroperidol (SP) derivatives N-methyl[¹⁸F]SP; N-ethyl[¹⁸F]SP; N-(2-[¹⁸F]fluoroethyl)SP; N-propyl[¹⁸F]SP; N-(3-[¹⁸F]fluoropropyl)SP; N-(3-fluoropropyl) [¹⁸F]SP; N-(2-[¹⁸F]fluoropropyl)SP; N-(2-[¹⁸F]fluorobutyl)SP; N-(2-[¹⁸F]fluoropentyl)SP; and N-(2-[¹⁸F]fluorohexyl) SP were synthesized. The lipophilicity of these ligands (log octanol/water partition coefficient) varies from 2.67 to 5.56 and the initial brain uptake in rats, measured at 2 min, was greatest with the methyl, ethyl, propyl, fluoroethyl, and fluoropropyl derivatives. The highest striatum/cerebellum values 1 h after administration were obtained with the N-methyl, N-propyl, and N-3-fluoropropyl derivatives, while that of N-2-fluoroethyl showed the greatest uptake of total activity in the brain at this time. The uptake of all these ligands in the striatum could be blocked by cold SP showing the striatal uptake to be by the dopamine receptors.