Radiosynthesis and quality control of [11C]TASP457 as a clinically useful PET ligand for imaging of histamine H3 receptors in human brain.

INTRODUCTION: Recently, 6-[(1-cyclobutylpiperidin-4-yl)oxy]-1-(6-[11C]methoxypyridin-3-yl)-3,4-dihydroquinolin-2(1H)-one ([11C]TASP457, [11C]2) has been developed as a novel PET ligand for histamine H3 receptors in brain. [11C]2 is potentially suitable for imaging H3 receptors in rat and monkey brains, which has motivated us to perform first-in-human study of [11C]2 for qualifying H3 receptors in human brain. In this paper, we report an efficient radiosynthesis of [11C]2 to obtain sufficient radioactivity and high quality for clinical application. METHODS: In manual synthesis, we optimized the reaction conditions of desmethyl precursor 1, which contains a 2-hydroxypyridine moiety, with [11C]MeI or [11C]MeOTf. After optimization, we performed automated synthesis and quality control of [11C]2. RESULTS: Bubbling [11C]MeOTf into a heated mixture of precursor 1 and cesium carbonate in DMF at 100°C for 90s produced [11C]2 with decay-corrected radiochemical yields of (based on [11C]CO2) 7.9±1.8% (n=78). The specific activity of [11C]2 was 156±52GBq/µmol (n=78) at the end of synthesis. The total synthesis time was approximately 35min from the end of bombardment. All the quality control results of [11C]2 were in compliance with our in-house quality control/assurance specifications. CONCLUSION: We radiosynthesized [11C]TASP457 ([11C]2) with sufficient amounts of radioactivity and high quality for clinical usefulness. This radioligand is being used for PET assessment of H3 receptors in human brain in our facility.

Efficient radiosynthesis and non-clinical safety tests of the TSPO radioprobe [(18)F]FEDAC: Prerequisites for clinical application.

INTRODUCTION: [(18)F]FEDAC ([(18)F]1) has potent binding affinity and selectivity for translocator protein (18kDa, TSPO), and has been used to noninvasively visualize neuroinflammation, lung inflammation, acute liver damage, nonalcoholic fatty liver disease, and liver fibrosis. We had previously synthesized [(18)F]1 in two steps: (i) preparation of [(18)F]fluoroethyl bromide and (ii) coupling of [(18)F]fluoroethyl bromide with the appropriate precursor (2) for labeling. In this study, to clinically utilize [(18)F]1 as a PET radiopharmaceutical and to transfer the production technique of [(18)F]1 to other PET centers, we simplified its preparation by using a direct, one-step, tosyloxy-for-fluorine substitution. We also performed an acute toxicity study as a major non-clinical safety test, and determined radiometabolites using human liver microsomes. METHODS: [(18)F]1 was prepared via direct (18)F-fluorination by heating the corresponding tosylated derivative (3) with [(18)F]fluoride as its Kryptofix 222 complex in dimethyl sulfoxide at 110°C for 15min, following by HPLC purification. Non-clinical safety tests were performed for the extended single-dose toxicity study in rats, and for the in vitro metabolite analysis with human liver microsomal incubation. RESULTS: High quality batches of [(18)F]1, compatible with clinical applications, were obtained. At the end of irradiation, the decay-corrected radiochemical yield of [(18)F]1 using 1 and 5mg of precursor based on [(18)F]fluoride was 18.5±7.9% (n=10) and 52.0±5.8% (n=3), respectively. A single-dose of [(18)F]1 did not show toxicological effects for 14 days after the injection in male and female rats. In human liver microsomal incubations, [(18)F]1 was easily metabolized to [(18)F]desbenzyl-FEDAC ([(18)F]10) by CYPs (4.2% of parent compound left 60min after incubation). CONCLUSION: We successfully synthesized clinical grade batches of [(18)F]1 and verified the absence of innocuity of this radiotracer. [(18)F]1 will be used to first-in-human studies in our facility.

Dynamic Changes in Striatal mGluR1 But Not mGluR5 during Pathological Progression of Parkinson's Disease in Human Alpha-Synuclein A53T Transgenic Rats: A Multi-PET Imaging Study.

Parkinson's disease (PD) is a prevalent degenerative disorder affecting the CNS that is primarily characterized by resting tremor and movement deficits. Group I metabotropic glutamate receptor subtypes 1 and 5 (mGluR1 and mGluR5, respectively) are important targets for investigation in several CNS disorders. In the present study, we investigated the in vivo roles of mGluR1 and mGluR5 in chronic PD pathology by performing longitudinal positron emission tomography (PET) imaging in A53T transgenic (A53T-Tg) rats expressing an abnormal human α-synuclein (ASN) gene. A53T-Tg rats showed a dramatic decline in general motor activities with age, along with abnormal ASN aggregation and striatal neuron degeneration. In longitudinal PET imaging, striatal nondisplaceable binding potential (BPND) values for [(11)C]ITDM (N-[4-[6-(isopropylamino) pyrimidin-4-yl]-1,3-thiazol-2-yl]-N-methyl-4-[(11)C]methylbenzamide), a selective PET ligand for mGluR1, temporarily increased before PD symptom onset and dramatically decreased afterward with age. However, striatal BPND values for (E)-[(11)C]ABP688 [3-(6-methylpyridin-2-ylethynyl)-cyclohex-2-enone-(E)-O-[(11)C]methyloxime], a specific PET ligand for mGluR5, remained constant during experimental terms. The dynamic changes in striatal mGluR1 BPND values also showed a high correlation in pathological decreases in general motor activities. Furthermore, declines in mGluR1 BPND values were correlated with decreases in BPND values for [(18)F]FE-PE2I [(E)-N-(3-iodoprop-2E-enyl)-2ß-carbo-[(18)F]fluoroethoxy-3ß-(4-methylphenyl) nortropane], a specific PET ligand for the dopamine transporter, a biomarker for dopaminergic neurons. In conclusion, our results have demonstrated for the first time that dynamic changes occur in mGluR1, but not mGluR5, that accompany pathological progression in a PD animal model. SIGNIFICANCE STATEMENT: Synaptic signaling by glutamate, the principal excitatory neurotransmitter in the brain, is modulated by group I metabotropic glutamate receptors, including the mGluR1 and mGluR5 subtypes. In the brain, mGluR1 and mGluR5 have distinct functional roles and regional distributions. Their roles in brain pathology, however, are not well characterized. Using longitudinal PET imaging in a chronic rat model of PD, we demonstrated that expression of mGluR1, but not mGluR5, dynamically changed in the striatum accompanying pathological PD progression. These findings imply that monitoring mGluR1 in vivo may provide beneficial information to further understand central nervous system disorders.

Identification of a major radiometabolite of [11C]PBB3.

INTRODUCTION: [(11)C]PBB3 is a clinically used positron emission tomography (PET) probe for in vivo imaging of tau pathology in the brain. Our previous study showed that [(11)C]PBB3 was rapidly decomposed to a polar radiometabolite in the plasma of mice. For the pharmacokinetic evaluation of [(11)C]PBB3 it is important to elucidate the characteristics of radiometabolites. In this study, we identified the chemical structure of a major radiometabolite of [(11)C]PBB3 and proposed the metabolic pathway of [(11)C]PBB3. METHODS: Carrier-added [(11)C]PBB3 was injected into a mouse for in vivo metabolite analysis. The chemical structure of a major radiometabolite was identified using LC-MS. Mouse and human liver microsomes and liver S9 samples were incubated with [(11)C]PBB3 in vitro. In silico prediction software was used to assist in the determination of the metabolite and metabolic pathway of [(11)C]PBB3. RESULTS: In vivo analysis showed that the molecular weight of a major radiometabolite of [(11)C]PBB3, which was called as [(11)C]M2, was m/z 390 [M+H(+)]. In vitro analysis assisted by in silico prediction showed that [(11)C]M2, which was not generated by cytochrome P450 enzymes (CYPs), was generated by sulfated conjugation mediated by a sulfotransferase. CONCLUSION: The major radiometabolite, [(11)C]M2, was identified as a sulfated conjugate of [(11)C]PBB3. [(11)C]PBB3 was metabolized mainly by a sulfotransferase and subsidiarily by CYPs.

Radiosynthesis, photoisomerization, biodistribution, and metabolite analysis of 11C-PBB3 as a clinically useful PET probe for imaging of tau pathology.

UNLABELLED: 2-((1E,3E)-4-(6-((11)C-methylamino)pyridin-3-yl)buta-1,3-dienyl)benzo[d]thiazol-6-ol ((11)C-PBB3) is a clinically useful PET probe that we developed for in vivo imaging of tau pathology in the human brain. To ensure the availability of this probe among multiple PET facilities, in the present study we established protocols for the radiosynthesis and quality control of (11)C-PBB3 and for the characterization of its photoisomerization, biodistribution, and metabolism. METHODS: (11)C-PBB3 was synthesized by reaction of the tert-butyldimethylsilyl desmethyl precursor ( 1: ) with (11)C-methyl iodide using potassium hydroxide as a base, followed by deprotection. Photoisomerization of (11)C-PBB3 under fluorescent light was determined. The biodistribution and metabolite analysis of (11)C-PBB3 was determined in mice using the dissection method. RESULTS: (11)C-PBB3 was synthesized with 15.4% ± 2.8% radiochemical yield (decay-corrected, n = 50) based on the cyclotron-produced (11)C-CO2 and showed an averaged synthesis time of 35 min from the end of bombardment. The radiochemical purity and specific activity of (11)C-PBB3 were 98.0% ± 2.3% and 180.2 ± 44.3 GBq/µmol, respectively, at the end of synthesis (n = 50). (11)C-PBB3 showed rapid photoisomerization, and its radiochemical purity decreased to approximately 50% at 10 min after exposure to fluorescent light. After the fluorescent light was switched off, (11)C-PBB3 retained more than 95% radiochemical purity over 60 min. A suitable brain uptake (1.92% injected dose/g tissue) of radioactivity was observed at 1 min after the probe injection, which was followed by rapid washout from the brain tissue. More than 70% of total radioactivity in the mouse brain homogenate at 5 min after injection represented the unchanged (11)C-PBB3, despite its rapid metabolism in the plasma. CONCLUSION: (11)C-PBB3 was produced with sufficient radioactivity and high quality, demonstrating its clinical utility. The present results of radiosynthesis, photoisomerization, biodistribution, and metabolite analysis could be helpful for the reliable production and application of (11)C-PBB3 in diverse PET facilities.

Levels of interleukin-18 are markedly increased in Helicobacter pylori-infected gastric mucosa among patients with specific IL18 genotypes.

BACKGROUND: The cellular immune response in gastric mucosa infected with Helicobacter pylori is proposed to be predominantly of the T helper cell type 1 type. METHODS: Interleukin (IL)-18, IL-12, and interferon (IFN)-gamma levels were measured in gastric mucosal biopsy specimens by reverse-transcription polymerase chain reaction (PCR) and by enzyme-linked immunosorbent assay; IL18 polymorphisms were determined by PCR. RESULTS: Biopsy specimens from 128 patients (56 with nonulcer dyspepsia, 28 with gastric ulcers, 28 with duodenal ulcers, and 16 with gastric cancer) were examined; 96 patients had H. pylori infection. IL-18 levels were markedly up-regulated in mucosa infected with H. pylori (P < .001), whereas IL-12 and IFN-gamma levels were independent of H. pylori status. IL-18 levels correlated with IFN-gamma levels only in infected patients (R = 0.31 to R = 0.51). IL-18 levels were the determining factor for monocyte infiltration in H. pylori-infected mucosa (P < .001). H. pylori-infected patients displaying IL18 -607C/C and -137G/G had higher IL-18 levels than did those with other genotypes and were more likely to experience treatment failure. CONCLUSION: H. pylori infection induces IL-18 in the gastric mucosa. H. pylori-infected patients with IL18 -607C/C and -137G/G have higher IL-18 levels, which causes severe gastric inflammation. IL18 genotype might be a marker for predicting the effects of eradication therapy.

IL-17 is involved in Helicobacter pylori-induced gastric inflammatory responses in a mouse model.

BACKGROUND: Helicobacter pylori (H. pylori) is the major cause of chronic active gastritis and peptic ulcer disease. Recent studies have shown that H. pylori produces various cytokines that are related to neutrophil or mononuclear cell accumulation. Interleukin-17 (IL-17) is the founding member of an emerging family of inflammatory cytokines whose biological activities remain incompletely defined. In this study, the contributions of IL-17 to the induction of gastric inflammation and to the protection from H. pylori infection were investigated using IL-17 gene-knockout (IL-17(-/-)) mice. MATERIALS AND METHODS: IL-17(-/-)and wild-type C57BL/6 mice were challenged with H. pylori CPY2052 (2 x 10(8) CFU/mL) and the histological and microbiological evaluation were carried out at specified times. IL-17 and myeloperoxidase (MPO) protein levels in tissues were assayed in duplicate using ELISA kits. RESULTS: In wild-type mice, IL-17 was undetected at baseline; however, the protein expression of IL-17 was induced after infection with H. pylori. A severe infiltration of neutrophils appeared in the submucosa and the lamina propria in wild-type mice. In contrast, the degree of neutrophil infiltration in IL-17(-/-) mice was significantly lower than that in wild-type mice. Although wild-type mice infected with H. pylori showed drastically higher MPO activity compared with uninfected wild-type mice, any significant increase in the enzyme activity was not revealed in infected IL-17(-/-) mice. The number of H. pylori colonized in the stomach of IL-17(-/-) mice was significantly lower than that of wild-type mice from 1 to 6 months after infection. CONCLUSIONS: These results suggest that IL-17 may play an important role in the inflammatory response to the H. pylori infection and ultimately influence the outcome of the H. pylori-associated disease.