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Abstract Background The Editorial Board of EJNMMI Radiopharmacy and Chemistry releases a biannual highlight commentary to update the readership on trends in the field of radiopharmaceutical development. Main body This selection of highlights provides commentary on 24 different topics selected by each coauthoring Editorial Board member addressing a variety of aspects ranging from novel radiochemistry to first-in-human application of novel radiopharmaceuticals. Conclusion Trends in radiochemistry and radiopharmacy are highlighted. Hot topics cover the entire scope of EJNMMI Radiopharmacy and Chemistry, demonstrating the progress in the research field in many aspects.

Harrington et al report their discovery of Nemacol, which is a small molecule inhibitor of the vesicular acetylcholine transporter (VAChT). VAChT loads synaptic vesicles with acetylcholine and is a key point of vulnerability in animals. Harrington et al show that Nemacol has nematode selectivity and potential utility against nematode parasites.

σ1 receptors play a crucial role in various neurological and neurodegenerative diseases including pain, psychosis, Alzheimer’s disease, and depression. Spirocyclic piperidines represent a promising class of potent σ1 receptor ligands. The relationship between structural modifications and σ1 receptor affinity and selectivity over σ2 receptors led to the 2-fluoroethyl derivative fluspidine (2, Ki = 0.59 nM). Enantiomerically pure (S)-configured fluspidine ((S)-2) was prepared by the enantioselective reduction of the α,β-unsaturated ester 23 with NaBH4 and the enantiomerically pure co-catalyst (S,S)-24. The pharmacokinetic properties of both fluspidine enantiomers (R)-2 and (S)-2 were analyzed in vitro. Molecular dynamics simulations revealed very similar interactions of both fluspidine enantiomers with the σ1 receptor protein, with a strong ionic interaction between the protonated amino moiety of the piperidine ring and the COO- moiety of glutamate 172. The 18F-labeled radiotracers (S)-[18F]2 and (R)-[18F]2 were synthesized in automated syntheses using a TRACERlab FX FN synthesis module. High radiochemical yields and radiochemical purity were achieved. Radiometabolites were not found in the brains of mice, piglets, and rhesus monkeys. While both enantiomers revealed similar initial brain uptake, the slow washout of (R)-[18F]2 indicated a kind of irreversible binding. In the first clinical trial, (S)-[18F]2 was used to visualize σ1 receptors in the brains of patients with major depressive disorder (MDD). This study revealed an increased density of σ1 receptors in cortico-striato-(para)limbic brain regions of MDD patients. The increased density of σ1 receptors correlated with the severity of the depressive symptoms. In an occupancy study with the PET tracer (S)-[18F]2, the selective binding of pridopidine at σ1 receptors in the brain of healthy volunteers and HD patients was shown.

Glioblastoma (GBM) is still an incurable tumor that is associated with high recurrence rate and poor survival despite the current treatment regimes. With the urgent need for novel therapeutic strategies, immunotherapies, especially chimeric antigen receptor (CAR)-expressing T cells, represent a promising approach for specific and effective targeting of GBM. However, CAR T cells can be associated with serious side effects. To overcome such limitation, we applied our switchable RevCAR system to target both the epidermal growth factor receptor (EGFR) and the disialoganglioside GD2, which are expressed in GBM. The RevCAR system is a modular platform that enables controllability, improves safety, specificity and flexibility. Briefly, it consists of RevCAR T cells having a peptide epitope as extracellular domain, and a bispecific target module (RevTM). The RevTM acts as a switch key that recognizes the RevCAR epitope and the tumor-associated antigen, and thereby activating the RevCAR T cells to kill the tumor cells. However, in the absence of the RevTM, the RevCAR T cells are switched off. In this study, we show that the novel EGFR/GD2-specific RevTMs can selectively activate RevCAR T cells to kill GBM cells. Moreover, we show that gated targeting of GBM is possible with our Dual-RevCAR T cells, which have their internal activation and co-stimulatory domains separated into two receptors. Therefore, a full activation of Dual-RevCAR T cells can only be achieved when both receptors recognize EGFR and GD2 simultaneously via RevTMs, leading to a significant killing of GBM cells both in vitro and in vivo.

Serotonin receptors modulate numerous behavioral and neuropsychological processes. Therefore, they are the target for the action of many drugs, such as antipsychotics, antidepressants, antiemetics, migraine remedies, and many others. The 5-HT1A receptors have been involved in the pathogenesis and treatment of anxiety and depression and represent a promising target for new drugs with reduced extrapyramidal side effects. In most antidepressants, a piperazine-based structural motif can be identified as a common moiety. Here we describe the synthesis, pharmacological, and in silico characterization of a novel arylpiperazines series with excellent 5-HT1A affinity. The final compounds, 4a, 8a, and 8b, were selected according to predictions of in silico pharmacokinetics, docking analysis, and molecular dynamics in conjunction with physical properties, and metabolic stability. The accentuated molecules could serve as a lead compound for developing 5-HT1A drug-like molecules for depression treatment.

Abstract Background The Editorial Board of EJNMMI Radiopharmacy and Chemistry releases a biyearly highlight commentary to update the readership on trends in the field of radiopharmaceutical development. Results This commentary of highlights has resulted in 23 different topics selected by each member of the Editorial Board addressing a variety of aspects ranging from novel radiochemistry to first in man application of novel radiopharmaceuticals and also a contribution in relation to MRI-agents is included. Conclusion Trends in (radio)chemistry and radiopharmacy are highlighted demonstrating the progress in the research field being the scope of EJNMMI Radiopharmacy and Chemistry.

We have discovered and synthesized a series of indole-based derivatives as novel sigma-2 (σ2) receptor ligands. Two ligands with high σ2 receptor affinity and subtype selectivity were then radiolabeled with F-18 in good radiochemical yields and purities, and evaluated in rodents. In biodistribution studies in male ICR mice, radioligand [18F]9, or 1-(4-(5,6-dimethoxyisoindolin-2-yl)butyl)-4-(2-[18F]fluoroethoxy)-1H-indole, was found to display high brain uptake and high brain-to-blood ratio. Pretreatment of animals with the selective σ2 receptor ligand CM398 led to significant reductions in both brain uptake (29%–54%) and brain-to-blood ratio (60%–88%) of the radioligand in a dose-dependent manner, indicating high and saturable specific binding of [18F]9 to σ2 receptors in the brain. Further, ex vivo autoradiography in male ICR mice demonstrated regionally heterogeneous specific binding of [18F]9 in the brain that is consistent with the distribution pattern of σ2 receptors. Dynamic positron emission tomography imaging confirmed regionally distinct distribution and high levels of specific binding for [18F]9 in the rat brain, along with appropriate tissue kinetics. Taken together, results from our current study indicated the novel radioligand [18F]9 as the first highly specific and promising imaging agent for σ2 receptors in the brain.

G protein-coupled receptors (GPCRs) transfer extracellular signals across cell membranes by activating intracellular heterotrimeric G proteins. Several studies suggested G proteins as novel drug targets for the treatment of complex diseases, e.g., asthma and cancer. Recently, we developed specific radiotracers, [³H]PSB-15900-FR and [³H]PSB-16254-YM, for the Gαq family of G proteins by tritiation of the macrocyclic natural products FR900359 (FR) and YM-254890 (YM). In the present study, we utilized these potent radioligands to perform autoradiography studies in tissues of healthy mice, mouse models of disease, and human tissues. Specific binding was high, while non-specific binding was extraordinarily low, giving nearly identical results for both radioligands. High expression levels of Gαq proteins were detected in healthy mouse organs showing the following rank order of potency: kidney > liver > brain > pancreas > lung > spleen, while expression in the heart was low. Organ sub-structures, e.g., of mouse brain and lung, were clearly distinguishable. Whereas an acute asthma model in mice did not result in altered Gαq protein expressions as compared to control animals, a cutaneous melanoma model displayed significantly increased expression in comparison to healthy skin. These results suggest the future development of Gαq-protein-binding radio-tracers as novel diagnostics.

The cyclic nucleotide phosphodiesterase 2A is an intracellular enzyme which hydrolyzes the secondary messengers cAMP and cGMP and therefore plays an important role in signaling cascades. A high expression in distinct brain areas as well as in cancer cells makes PDE2A an interesting therapeutic and diagnostic target for neurodegenerative and neuropsychiatric diseases as well as for cancer. Aiming at specific imaging of this enzyme in the brain with positron emission tomography (PET), a new triazolopyridopyrazine-based derivative (11) was identified as a potent PDE2A inhibitor (IC50, PDE2A = 1.99 nM; IC50, PDE10A ~2000 nM) and has been radiofluorinated for biological evaluation. In vitro autoradiographic studies revealed that [18F]11 binds with high affinity and excellent specificity towards PDE2A in the rat brain. For the PDE2A-rich region nucleus caudate and putamen an apparent KD value of 0.24 nM and an apparent Bmax value of 16 pmol/mg protein were estimated. In vivo PET-MR studies in rats showed a moderate brain uptake of [18F]11 with a highest standardized uptake value (SUV) of 0.97. However, no considerable enrichment in PDE2A-specific regions in comparison to a reference region was detectable (SUVcaudate putamen = 0.51 vs. SUVcerebellum = 0.40 at 15 min p.i.). Furthermore, metabolism studies revealed a considerable uptake of radiometabolites of [18F]11 in the brain (66% parent fraction at 30 min p.i.). Altogether, despite the low specificity and the blood–brain barrier crossing of radiometabolites observed in vivo, [18F]11 is a valuable imaging probe for the in vitro investigation of PDE2A in the brain and has potential as a lead compound for further development of a PDE2A-specific PET ligand for neuroimaging.

Isocitrate dehydrogenases (IDHs) are metabolic enzymes commonly mutated in human cancers (glioma, acute myeloid leukaemia, chondrosarcoma, and intrahepatic cholangiocarcinoma). These mutated variants of IDH (mIDH) acquire a neomorphic activity, namely, conversion of α-ketoglutarate to the oncometabolite D-2-hydroxyglutarate involved in tumourigenesis. Thus, mIDHs have emerged as highly promising therapeutic targets, and several mIDH specific inhibitors have been developed. However, the evaluation of mIDH status, currently performed by biopsy, is essential for patient stratification and thus treatment and follow-up. We report herein the development of new radioiodinated and radiofluorinated analogues of olutasidenib (FT-2102) as tools for noninvasive single photon emission computed tomography (SPECT) or positron emission tomography (PET) imaging of mIDH1 up- and dysregulation in tumours. Nonradiolabelled derivatives 2 and 3 halogenated at position 6 of the quinolinone scaffold were synthesised and tested in vitro for their inhibitory potencies and selectivities in comparison with the lead compound FT-2102. Using a common organotin precursor, (S)-[125I]2 and (S)-[18F]3 were efficiently synthesised by radio-iododemetallation and copper-mediated radiofluorination, respectively. Both radiotracers were stable at room temperature in saline or DPBS solution and at 37 °C in mouse serum, allowing future planning of their in vitro and in vivo evaluations in glioma and chondrosarcoma models.

The cerebral expression of the A2A adenosine receptor (A2AAR) is altered in neurodegenerative diseases such as Parkinson’s (PD) and Huntington’s (HD) diseases, making these receptors an attractive diagnostic and therapeutic target. We aimed to further investigate the pharmacokinetic properties in the brain of our recently developed A2AAR–specific antagonist radiotracer [18F]FLUDA. For this purpose, we retrospectively analysed dynamic PET studies of healthy mice and rotenone–treated mice, and conducted dynamic PET studies with healthy pigs. We performed analysis of mouse brain time–activity curves to calculate the mean residence time (MRT) by non–compartmental analysis, and the binding potential (BPND) of [18F]FLUDA using the simplified reference tissue model (SRTM). For the pig studies, we performed a Logan graphical analysis to calculate the radiotracer distribution volume (VT) at baseline and under blocking conditions with tozadenant. The MRT of [18F]FLUDA in the striatum of mice was decreased by 30% after treatment with the A2AAR antagonist istradefylline. Mouse results showed the highest BPND (3.9 to 5.9) in the striatum. SRTM analysis showed a 20% lower A2AAR availability in the rotenone–treated mice compared to the control–aged group. Tozadenant treatment significantly decreased the VT (14.6 vs. 8.5 mL · g−1) and BPND values (1.3 vs. 0.3) in pig striatum. This study confirms the target specificity and a high BPND of [18F]FLUDA in the striatum. We conclude that [18F]FLUDA is a suitable tool for the non–invasive quantitation of altered A2AAR expression in neurodegenerative diseases such as PD and HD, by PET.

The degree of acetylation of lysine residues on histones influences the accessibility of DNA and, furthermore, the gene expression. Histone deacetylases (HDACs) are overexpressed in various tumour diseases, resulting in the interest in HDAC inhibitors for cancer therapy. The aim of this work is the development of a novel 18F-labelled HDAC1/2-specific inhibitor with a benzamide-based zinc-binding group to visualize these enzymes in brain tumours by positron emission tomography (PET). BA3, exhibiting high inhibitory potency for HDAC1 (IC50 = 4.8 nM) and HDAC2 (IC50 = 39.9 nM), and specificity towards HDAC3 and HDAC6 (specificity ratios >230 and >2080, respectively), was selected for radiofluorination. The two-step one-pot radiosynthesis of [18F]BA3 was performed in a TRACERlab FX2 N radiosynthesizer by a nucleophilic aliphatic substitution reaction. The automated radiosynthesis of [18F]BA3 resulted in a radiochemical yield of 1%, a radiochemical purity of >96% and a molar activity between 21 and 51 GBq/µmol (n = 5, EOS). For the characterization of BA3, in vitro and in vivo experiments were carried out. The results of these pharmacological and pharmacokinetic studies indicate a suitable inhibitory potency of BA3, whereas the applicability for non-invasive imaging of HDAC1/2 by PET requires further optimization of the properties of this compound.

Convection-enhanced delivery (CED) has been introduced as a concept in cancer treatment to generate high local concentrations of anticancer therapeutics and overcome the limited diffusional distribution, e.g., in the brain. RNA interference provides interesting therapeutic options to fight cancer cells but requires nanoparticulate (NP) carriers with a size below 100 nm as well as a low zeta potential for CED application. In this study, we investigated calcium phosphate NPs (CaP-NPs) as siRNA carriers for CED application. Since CaP-NPs tend to aggregate, we introduced a new terpolymer (o14PEGMA(1:1:2.5) NH3) for stabilization of CaP-NPs intended for delivery of siRNA to brain cancer cells. This small terpolymer provides PEG chains for steric stabilization, and a fat alcohol to improve interfacial activity, as well as maleic anhydrides that allow for both labeling and high affinity to Ca(II) in the hydrolyzed state. In a systematic approach, we varied the Ca/P ratio as well as the terpolymer concentration and successfully stabilized NPs with the desired properties. Labeling of the terpolymer with the fluorescent dye Cy5 revealed the terpolymer’s high affinity to CaP. Importantly, we also determined a high efficiency of siRNA binding to the NPs that caused very effective survivin siRNA silencing in F98 rat brain cancer cells. Cytotoxicity investigations with a standard cell line resulted in minor and transient effects; no adverse effects were observed in organotypic brain slice cultures. However, more specific cytotoxicity investigations are required. This study provides a systematic and mechanistic analysis characterizing the effects of the first oligomer of a new class of stabilizers for siRNA-loaded CaP-NPs.

(S)-[18F]fluspidine ((S)-[18F]1) has recently been explored for positron emission tomography (PET) imaging of sigma-1 receptors in humans. In the current report, we have used plasma samples of healthy volunteers to investigate the radiometabolites of (S)-[18F]1 and elucidate their structures with LC-MS/MS. For the latter purpose additional in vitro studies were conducted by incubation of (S)-[18F]1 and (S)-1 with human liver microsomes (HLM). In vitro metabolites were characterized by interpretation of MS/MS fragmentation patterns from collision-induced dissociation or by use of reference compounds. Thereby, structures of corresponding radio-HPLC-detected radiometabolites, both in vitro and in vivo (human), could be identified. By incubation with HLM, mainly debenzylation and hydroxylation occurred, beside further mono- and di-oxygenations. The product hydroxylated at the fluoroethyl side chain was glucuronidated. Plasma samples (10, 20, 30 min p.i., n = 5-6), obtained from human subjects receiving 250–300 MBq (S)-[18F]1 showed 97.2, 95.4, and 91.0% of unchanged radioligand, respectively. In urine samples (90 min p.i.) the fraction of unchanged radioligand was only 2.6% and three major radiometabolites were detected. The one with the highest percentage, also found in plasma, matched the glucuronide formed in vitro. Only a small amount of debenzylated metabolite was detected. In conclusion, our metabolic study, in particular the high fractions of unchanged radioligand in plasma, confirms the suitability of (S)-[18F]1 as PET radioligand for sigma-1 receptor imaging.

The G protein-coupled adenosine A2B receptor is suggested to be involved in various pathological processes accompanied by increased levels of adenosine as found in inflammation, hypoxia, and cancer. Therefore, the adenosine A2B receptor is currently in focus as a novel target for cancer therapy as well as for noninvasive molecular imaging via positron emission tomography (PET). Aiming at the development of a radiotracer labeled with the PET radionuclide fluorine-18 for imaging the adenosine A2B receptor in brain tumors, one of the most potent and selective antagonists, the xanthine derivative PSB-603, was selected as a lead compound. As initial biodistribution studies in mice revealed a negligible brain uptake of [3H]PSB-603 (SUV3min: 0.2), structural modifications were performed to optimize the physicochemical properties regarding blood–brain barrier penetration. Two novel fluorinated derivatives bearing a 2-fluoropyridine (5) moiety and a 4-fluoro-piperidine (6) moiety were synthesized, and their affinity towards the four adenosine receptor subtypes was determined in competition binding assays. Both compounds showed high affinity towards the adenosine A2B receptor (Ki (5) = 9.97 ± 0.86 nM; Ki (6) = 12.3 ± 3.6 nM) with moderate selectivity versus the other adenosine receptor subtypes.

Abstract Background Both enantiomers of [18F]flubatine are new radioligands for neuroimaging of α4β2 nicotinic acetylcholine receptors with positron emission tomography (PET) exhibiting promising pharmacokinetics which makes them attractive for different clinical questions. In a previous preclinical study, the main advantage of (+)-[18F]flubatine compared to (−)-[18F]flubatine was its higher binding affinity suggesting that (+)-[18F]flubatine might be able to detect also slight reductions of α4β2 nAChRs and could be more sensitive than (−)-[18F]flubatine in early stages of Alzheimer’s disease. To support the clinical translation, we investigated a fully image-based internal dosimetry approach for (+)-[18F]flubatine, comparing mouse data collected on a preclinical PET/MRI system to piglet and first-in-human data acquired on a clinical PET/CT system. Time-activity curves (TACs) were obtained from the three species, the animal data extrapolated to human scale, exponentially fitted and the organ doses (OD), and effective dose (ED) calculated with OLINDA. Results The excreting organs (urinary bladder, kidneys, and liver) receive the highest organ doses in all species. Hence, a renal/hepatobiliary excretion pathway can be assumed. In addition, the ED conversion factors of 12.1 μSv/MBq (mice), 14.3 μSv/MBq (piglets), and 23.0 μSv/MBq (humans) were calculated which are well within the order of magnitude as known from other 18F-labeled radiotracers. Conclusions Although both enantiomers of [18F]flubatine exhibit different binding kinetics in the brain due to the respective affinities, the effective dose revealed no enantiomer-specific differences among the investigated species. The preclinical dosimetry and biodistribution of (+)-[18F]flubatine was shown and the feasibility of a dose assessment based on image data acquired on a small animal PET/MR and a clinical PET/CT was demonstrated. Additionally, the first-in-human study confirmed the tolerability of the radiation risk of (+)-[18F]flubatine imaging which is well within the range as caused by other 18F-labeled tracers. However, as shown in previous studies, the ED in humans is underestimated by up to 50 % using preclinical imaging for internal dosimetry. This fact needs to be considered when applying for first-in-human studies based on preclinical biokinetic data scaled to human anatomy.

Abstract Phosphodiesterase type 10A (PDE10A) is highly enriched in striatum and is under evaluation as a drug target for several psychiatric/neurodegenerative diseases. Preclinical studies implicate PDE10A in the regulation of energy homeostasis, but the mechanisms remain unclear. By utilizing small‐animal PET/MRI and the novel radioligand [18F]‐AQ28A, we found marked levels of PDE10A in interscapular brown adipose tissue (BAT) of mice. Pharmacological inactivation of PDE10A with the highly selective inhibitor MP‐10 recruited BAT and potentiated thermogenesis in vivo. In diet‐induced obese mice, chronic administration of MP‐10 caused weight loss associated with increased energy expenditure, browning of white adipose tissue, and improved insulin sensitivity. Analysis of human PET data further revealed marked levels of PDE10A in the supraclavicular region where brown/beige adipocytes are clustered in adults. Finally, the inhibition of PDE10A with MP‐10 stimulated thermogenic gene expression in human brown adipocytes and induced browning of human white adipocytes. Collectively, our findings highlight a novel thermoregulatory role for PDE10A in mouse and human adipocytes and promote PDE10A inhibitors as promising candidates for the treatment of obesity and diabetes.

Glioblastoma multiforme (GBM) is the most devastating primary brain tumour characterised by infiltrative growth and resistance to therapies. According to recent research, the sigma-1 receptor (sig1R), an endoplasmic reticulum chaperone protein, is involved in signaling pathways assumed to control the proliferation of cancer cells and thus could serve as candidate for molecular characterisation of GBM. To test this hypothesis, we used the clinically applied sig1R-ligand (S)-(−)-[18F]fluspidine in imaging studies in an orthotopic mouse model of GBM (U87-MG) as well as in human GBM tissue. A tumour-specific overexpression of sig1R in the U87-MG model was revealed in vitro by autoradiography. The binding parameters demonstrated target-selective binding according to identical KD values in the tumour area and the contralateral side, but a higher density of sig1R in the tumour. Different kinetic profiles were observed in both areas, with a slower washout in the tumour tissue compared to the contralateral side. The translational relevance of sig1R imaging in oncology is reflected by the autoradiographic detection of tumour-specific expression of sig1R in samples obtained from patients with glioblastoma. Thus, the herein presented data support further research on sig1R in neuro-oncology.

Monocarboxylate transporters 1-4 (MCT1-4) are involved in several metabolism-related diseases, especially cancer, providing the chance to be considered as relevant targets for diagnosis and therapy. [18F]FACH was recently developed and showed very promising preclinical results as a potential positron emission tomography (PET) radiotracer for imaging of MCTs. Given that [18F]FACH did not show high blood-brain barrier permeability, the current work is aimed to investigate whether more lipophilic analogs of FACH could improve brain uptake for imaging of gliomas, while retaining binding to MCTs. The 2-fluoropyridinyl-substituted analogs 1 and 2 were synthesized and their MCT1 inhibition was estimated by [14C]lactate uptake assay on rat brain endothelial-4 (RBE4) cells. While compounds 1 and 2 showed lower MCT1 inhibitory potencies than FACH (IC50 = 11 nM) by factors of 11 and 25, respectively, 1 (IC50 = 118 nM) could still be a suitable PET candidate. Therefore, 1 was selected for radiosynthesis of [18F]1 and subsequent biological evaluation for imaging of the MCT expression in mouse brain. Regarding lipophilicity, the experimental log D7.4 result for [18F]1 agrees pretty well with its predicted value. In vivo and in vitro studies revealed high uptake of the new radiotracer in kidney and other peripheral MCT-expressing organs together with significant reduction by using specific MCT1 inhibitor α-cyano-4-hydroxycinnamic acid. Despite a higher lipophilicity of [18F]1 compared to [18F]FACH, the in vivo brain uptake of [18F]1 was in a similar range, which is reflected by calculated BBB permeabilities as well through similar transport rates by MCTs on RBE4 cells. Further investigation is needed to clarify the MCT-mediated transport mechanism of these radiotracers in brain.

The adenosine A2A receptor (A2AR) is regarded as a particularly appropriate target for non-dopaminergic treatment of Parkinson’s disease (PD). An increased A2AR availability has been found in the human striatum at early stages of PD and in patients with PD and dyskinesias. The aim of this small animal positron emission tomography/magnetic resonance (PET/MR) imaging study was to investigate whether rotenone-treated mice reflect the aspect of striatal A2AR upregulation in PD. For that purpose, we selected the known A2AR-specific radiotracer [18F]FESCH and developed a simplified two-step one-pot radiosynthesis. PET images showed a high uptake of [18F]FESCH in the mouse striatum. Concomitantly, metabolism studies with [18F]FESCH revealed the presence of a brain-penetrant radiometabolite. In rotenone-treated mice, a slightly higher striatal A2AR binding of [18F]FESCH was found. Nonetheless, the correlation between the increased A2AR levels within the proposed PD animal model remains to be further investigated.

Overexpression of monocarboxylate transporters (MCTs) has been shown for a variety of human cancers (e.g., colon, brain, breast, and kidney) and inhibition resulted in intracellular lactate accumulation, acidosis, and cell death. Thus, MCTs are promising targets to investigate tumor cancer metabolism with positron emission tomography (PET). Here, the organ doses (ODs) and the effective dose (ED) of the first 18F-labeled MCT1/MCT4 inhibitor were estimated in juvenile pigs. Whole-body dosimetry was performed in three piglets (age: ~6 weeks, weight: ~13–15 kg). The animals were anesthetized and subjected to sequential hybrid Positron Emission Tomography and Computed Tomography (PET/CT) up to 5 h after an intravenous (iv) injection of 156 ± 54 MBq [18F]FACH. All relevant organs were defined by volumes of interest. Exponential curves were fitted to the time–activity data. Time and mass scales were adapted to the human order of magnitude and the ODs calculated using the ICRP 89 adult male phantom with OLINDA 2.1. The ED was calculated using tissue weighting factors as published in Publication 103 of the International Commission of Radiation Protection (ICRP103). The highest organ dose was received by the urinary bladder (62.6 ± 28.9 µSv/MBq), followed by the gall bladder (50.4 ± 37.5 µSv/MBq) and the pancreas (30.5 ± 27.3 µSv/MBq). The highest contribution to the ED was by the urinary bladder (2.5 ± 1.1 µSv/MBq), followed by the red marrow (1.7 ± 0.3 µSv/MBq) and the stomach (1.3 ± 0.4 µSv/MBq). According to this preclinical analysis, the ED to humans is 12.4 µSv/MBq when applying the ICRP103 tissue weighting factors. Taking into account that preclinical dosimetry underestimates the dose to humans by up to 40%, the conversion factor applied for estimation of the ED to humans would rise to 20.6 µSv/MBq. In this case, the ED to humans upon an iv application of ~300 MBq [18F]FACH would be about 6.2 mSv. This risk assessment encourages the translation of [18F]FACH into clinical study phases and the further investigation of its potential as a clinical tool for cancer imaging with PET.

Changes in the expression of α7 nicotinic acetylcholine receptors (α7 nAChRs) in the human brain are widely assumed to be associated with neurological and neurooncological processes. Investigation of these receptors in vivo depends on the availability of imaging agents such as radioactively labelled ligands applicable in positron emission tomography (PET). We report on a series of new ligands for α7 nAChRs designed by the combination of dibenzothiophene dioxide as a novel hydrogen bond acceptor functionality with diazabicyclononane as an established cationic center. To assess the structure-activity relationship (SAR) of this new basic structure, we further modified the cationic center systematically by introduction of three different piperazine-based scaffolds. Based on in vitro binding affinity and selectivity, assessed by radioligand displacement studies at different rat and human nAChR subtypes and at the structurally related human 5-HT3 receptor, we selected the compound 7-(1,4-diazabicyclo[3.2.2]nonan-4-yl)-2-fluorodibenzo-[b,d]thiophene 5,5-dioxide (10a) for radiolabeling and further evaluation in vivo. Radiosynthesis of [18F]10a was optimized and transferred to an automated module. Dynamic PET imaging studies with [18F]10a in piglets and a monkey demonstrated high uptake of radioactivity in the brain, followed by washout and target-region specific accumulation under baseline conditions. Kinetic analysis of [18F]10a in pig was performed using a two-tissue compartment model with arterial-derived input function. Our initial evaluation revealed that the dibenzothiophene-based PET radioligand [18F]10a ([18F]DBT-10) has high potential to provide clinically relevant information about the expression and availability of α7 nAChR in the brain.

Phosphodiesterase 2A (PDE2A) is highly and specifically expressed in particular brain regions that are affected by neurological disorders and in certain tumors. Development of a specific PDE2A radioligand would enable molecular imaging of the PDE2A protein via positron emission tomography (PET). Herein we report on the syntheses of three novel fluoroalkylated triazine derivatives (TA2–4) and on the evaluation of their effect on the enzymatic activity of human PDE2A. The most potent PDE2A inhibitors were 18F-radiolabelled ([18F]TA3 and [18F]TA4) and investigated regarding their potential as PET radioligands for imaging of PDE2A in mouse brain. In vitro autoradiography on rat brain displayed region-specific distribution of [18F]TA3 and [18F]TA4, which is consistent with the expression pattern of PDE2A protein. Metabolism studies of both [18F]TA3 and [18F]TA4 in mice showed a significant accumulation of two major radiometabolites of each radioligand in brain as investigated by micellar radio-chromatography. Small-animal PET/MR studies in mice using [18F]TA3 revealed a constantly increasing uptake of activity in the non-target region cerebellum, which may be caused by the accumulation of brain penetrating radiometabolites. Hence, [18F]TA3 and [18F]TA4 are exclusively suitable for in vitro investigation of PDE2A. Nevertheless, further structural modification of these promising radioligands might result in metabolically stable derivatives.

Sharpless asymmetric dihydroxylation of styrene derivative 6 afforded chiral triols (R)-7 and (S)-7, which were cyclized with tosyl chloride in the presence of Bu2SnO to provide 2-benzopyrans (R)-4 and (S)-4 with high regioselectivity. The additional hydroxy moiety in the 4-position was exploited for the introduction of various substituents. Williamson ether synthesis and replacement of the Boc protective group with a benzyl moiety led to potent σ1 ligands with high σ1/σ2-selectivity. With exception of the ethoxy derivative 16, the (R)-configured enantiomers represent eutomers with eudismic ratios of up to 29 for the ester (R)-18. The methyl ether (R)-15 represents the most potent σ1 ligand of this series of compounds, with a Ki value of 1.2 nM and an eudismic ratio of 7. Tosylate (R)-21 was used as precursor for the radiosynthesis of [18F]-(R)-20, which was available by nucleophilic substitution with K[18F]F K222 carbonate complex. The radiochemical yield of [18F]-(R)-20 was 18%–20%, the radiochemical purity greater than 97% and the specific radioactivity 175–300 GBq/µmol. Although radiometabolites were detected in plasma, urine and liver samples, radiometabolites were not found in brain samples. After 30 min, the uptake of the radiotracer in the brain was 3.4% of injected dose per gram of tissue and could be reduced by coadministration of the σ1 antagonist haloperidol. [18F]-(R)-20 was able to label those regions of the brain, which were reported to have high density of σ1 receptors.

A specific radioligand for the imaging of cyclic nucleotide phosphodiesterase 2A (PDE2A) via positron emission tomography (PET) would be helpful for research on the physiology and disease-related changes in the expression of this enzyme in the brain. In this report, the radiosynthesis of a novel PDE2A radioligand and the subsequent biological evaluation were described. Our prospective compound 1-(2-chloro-5-methoxy phenyl)-8-(2-fluoropyridin-4-yl)-3- methylbenzo[e]imidazo[5,1-c][1,2,4]triazine, benzoimidazotriazine (BIT1) (IC50 PDE2A = 3.33 nM; 16-fold selectivity over PDE10A) was fluorine-18 labeled via aromatic nucleophilic substitution of the corresponding nitro precursor using the K[18F]F-K2.2.2-carbonate complex system. The new radioligand [18F]BIT1 was obtained with a high radiochemical yield (54 ± 2%, n = 3), a high radiochemical purity (≥99%), and high molar activities (155−175 GBq/μmol, n = 3). In vitro autoradiography on pig brain cryosections exhibited a heterogeneous spatial distribution of [18F]BIT1 corresponding to the known pattern of expression of PDE2A. The investigation of in vivo metabolism of [18F]BIT1 in a mouse revealed sufficient metabolic stability. PET studies in mouse exhibited a moderate brain uptake of [18F]BIT1 with a maximum standardized uptake value of ~0.7 at 5 min p.i. However, in vivo blocking studies revealed a non-target specific binding of [18F]BIT1. Therefore, further structural modifications are needed to improve target selectivity.

Phosphodiesterase 10A (PDE10A) is a key enzyme of intracellular signal transduction which is involved in the regulation of neurotransmission. The molecular imaging of PDE10A by PET is expected to allow a better understanding of physiological and pathological processes related to PDE10A expression and function in the brain. The aim of this study was to develop a new 18F-labeled PDE10A ligand based on a 6,7-dimethoxy-4-pyrrolidinylquinazoline and to evaluate its properties in biodistribution studies. Nucleophilic substitution of the 7-tosyloxy-analogue led to the 7-[18F]fluoroethoxy-derivative [18F]IV with radiochemical yields of 25% ± 9% (n = 9), high radiochemical purity of ≥99% and specific activities of 110–1,100 GBq/μmol. [18F]IV showed moderate PDE10A affinity (KD,PDE10A = 14 nM) and high metabolic stability in the brain of female CD-1 mice, wherein the radioligand entered rapidly with a peak uptake of 2.3% ID/g in striatum at 5 min p.i. However, ex vivo autoradiographic and in vivo blocking studies revealed no target specific accumulation and demonstrated [18F]IV to be inapplicable for imaging PDE10A with PET.

Sigma-1 receptors (Sig1R) are highly expressed in various human cancer cells and hence imaging of this target with positron emission tomography (PET) can contribute to a better understanding of tumor pathophysiology and support the development of antineoplastic drugs. Two Sig1R-specific radiolabeled enantiomers (S)-(−)- and (R)-(+)-[18F]fluspidine were investigated in several tumor cell lines including melanoma, squamous cell/epidermoid carcinoma, prostate carcinoma, and glioblastoma. Dynamic PET scans were performed in mice to investigate the suitability of both radiotracers for tumor imaging. The Sig1R expression in the respective tumors was confirmed by Western blot. Rather low radiotracer uptake was found in heterotopically (subcutaneously) implanted tumors. Therefore, a brain tumor model (U87-MG) with orthotopic implantation was chosen to investigate the suitability of the two Sig1R radiotracers for brain tumor imaging. High tumor uptake as well as a favorable tumor-to-background ratio was found. These results suggest that Sig1R PET imaging of brain tumors with [18F]fluspidine could be possible. Further studies with this tumor model will be performed to confirm specific binding and the integrity of the blood-brain barrier (BBB).

Specific radioligands for in vivo visualization and quantification of cyclic nucleotide phosphodiesterase 2A (PDE2A) by positron emission tomography (PET) are increasingly gaining interest in brain research. Herein we describe the synthesis, the 18F-labelling as well as the biological evaluation of our latest PDE2A (radio-)ligand 9-(5-Butoxy-2-fluorophenyl)-2-(2-([18F])fluoroethoxy)-7-methylimidazo[5,1-c]pyrido[2,3-e][1,2,4]triazine (([18F])TA5). It is the most potent PDE2A ligand out of our series of imidazopyridotriazine-based derivatives so far (IC50 hPDE2A = 3.0 nM; IC50 hPDE10A > 1000 nM). Radiolabelling was performed in a one-step procedure starting from the corresponding tosylate precursor. In vitro autoradiography on rat and pig brain slices displayed a homogenous and non-specific binding of the radioligand. Investigation of stability in vivo by reversed-phase HPLC (RP-HPLC) and micellar liquid chromatography (MLC) analyses of plasma and brain samples obtained from mice revealed a high fraction of one main radiometabolite. Hence, we concluded that [18F]TA5 is not appropriate for molecular imaging of PDE2A neither in vitro nor in vivo. Our ongoing work is focusing on further structurally modified compounds with enhanced metabolic stability.

Both (+)-[18F]flubatine and its enantiomer (−)-[18F]flubatine are radioligands for the neuroimaging of α4β2 nicotinic acetylcholine receptors (nAChRs) by positron emission tomography (PET). In a clinical study in patients with early Alzheimer’s disease, (+)-[18F]flubatine ((+)-[18F]1) was examined regarding its metabolic fate, in particular by identification of degradation products detected in plasma and urine. The investigations included an in vivo study of (+)-flubatine ((+)-1) in pigs and structural elucidation of formed metabolites by LC-MS/MS. Incubations of (+)-1 and (+)-[18F]1 with human liver microsomes were performed to generate in vitro metabolites, as well as radiometabolites, which enabled an assignment of their structures by comparison of LC-MS/MS and radio-HPLC data. Plasma and urine samples taken after administration of (+)-[18F]1 in humans were examined by radio-HPLC and, on the basis of results obtained in vitro and in vivo, formed radiometabolites were identified. In pigs, (+)-1 was monohydroxylated at different sites of the azabicyclic ring system of the molecule. Additionally, one intermediate metabolite underwent glucuronidation, as also demonstrated in vitro. In humans, a fraction of 95.9 ± 1.9% (n = 10) of unchanged tracer remained in plasma, 30 min after injection. However, despite the low metabolic degradation, both radiometabolites formed in humans could be characterized as (i) a product of C-hydroxylation at the azabicyclic ring system, and (ii) a glucuronide conjugate of the precedingly-formed N8-hydroxylated (+)-[18F]1.

A small series of indol-3-yl-oxoacetamides was synthesized starting from the literature known N-(adamantan-1-yl)-2-(5-(furan-2-yl)-1-pentyl-1H-indol-3-yl)-2-oxoacetamide (5) by substituting the 1-pentyl-1H-indole subunit. Our preliminary biological evaluation showed that the fluorinated derivative 8 is a potent and selective CB2 ligand with Ki = 6.2 nM.

The enantiomers of [18F]fluspidine, recently developed for imaging of σ1 receptors, possess distinct pharmacokinetics facilitating their use in different clinical settings. To support their translational potential, we estimated the human radiation dose of (S)-(−)-[18F]fluspidine and (R)-(+)-[18F]fluspidine from ex vivo biodistribution and PET/MRI data in mice after extrapolation to the human scale. In addition, we validated the preclinical results by performing a first-in-human PET/CT study using (S)-(−)-[18F]fluspidine. Based on the respective time-activity curves, we calculated using OLINDA the particular organ doses (ODs) and effective doses (EDs). The ED values of (S)-(−)-[18F]fluspidine and (R)-(+)-[18F]fluspidine differed significantly with image-derived values obtained in mice with 12.9 μSv/MBq and 14.0 μSv/MBq (p < 0.025), respectively. A comparable ratio was estimated from the biodistribution data. In the human study, the ED of (S)-(−)-[18F]fluspidine was calculated as 21.0 μSv/MBq. Altogether, the ED values for both [18F]fluspidine enantiomers determined from the preclinical studies are comparable with other 18F-labeled PET imaging agents. In addition, the first-in-human study confirmed that the radiation risk of (S)-(−)-[18F]fluspidine imaging is within acceptable limits. However, as already shown for other PET tracers, the actual ED of (S)-(−)-[18F]fluspidine in humans was underestimated by preclinical imaging which needs to be considered in other first-in-human studies.

Cyclic nucleotide phosphodiesterases (PDEs) are a class of intracellular enzymes that inactivate the secondary messenger molecules, cyclic adenosine monophosphate (cAMP) and cyclic guanosine monophosphate (cGMP). Thus, PDEs regulate the signaling cascades mediated by these cyclic nucleotides and affect fundamental intracellular processes. Pharmacological inhibition of PDE activity is a promising strategy for treatment of several diseases. However, the role of the different PDEs in related pathologies is not completely clarified yet. PDE-specific radioligands enable non-invasive visualization and quantification of these enzymes by positron emission tomography (PET) in vivo and provide an important translational tool for elucidation of the relationship between altered expression of PDEs and pathophysiological effects as well as (pre-)clinical evaluation of novel PDE inhibitors developed as therapeutics. Herein we present an overview of novel PDE radioligands for PET published since 2012.

Phosphodiesterases (PDEs) are enzymes that play a major role in cell signalling by hydrolysing the secondary messengers cyclic adenosine monophosphate (cAMP) and/or cyclic guanosine monophosphate (cGMP) throughout the body and brain. Altered cyclic nucleotide-mediated signalling has been associated with a wide array of disorders, including neurodegenerative disorders. Recently, PDE5 has been shown to be involved in neurodegenerative disorders such as Alzheimer’s disease, but its precise role has not been elucidated yet. To visualize and quantify the expression of this enzyme in brain, we developed a radiotracer for specific PET imaging of PDE5. A quinoline-based lead compound has been structurally modified resulting in the fluoroethoxymethyl derivative ICF24027 with high inhibitory activity towards PDE5 (IC50 = 1.86 nM). Radiolabelling with fluorine-18 was performed by a one-step nucleophilic substitution reaction using a tosylate precursor (RCY(EOB) = 12.9% ± 1.8%; RCP > 99%; SA(EOS) = 70–126 GBq/μmol). In vitro autoradiographic studies of [18F]ICF24027 on different mouse tissue as well as on porcine brain slices demonstrated a moderate specific binding to PDE5. In vivo studies in mice revealed that [18F]ICF24027 was metabolized under formation of brain penetrable radiometabolites making the radiotracer unsuitable for PET imaging of PDE5 in brain.