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The 18 kDa translocator protein (TSPO), previously known as the peripheral benzodiazepine receptor, is predominately localized to the outer mitochondrial membrane in steroidogenic cells. Brain TSPO expression is relatively low under physiological conditions, but is upregulated in response to glial cell activation. As the primary index of neuroinflammation, TSPO is implicated in the pathogenesis and progression of numerous neuropsychiatric disorders and neurodegenerative diseases, including Alzheimer’s disease (AD), amyotrophic lateral sclerosis (ALS), Parkinson’s disease (PD), multiple sclerosis (MS), major depressive disorder (MDD) and obsessive compulsive disorder (OCD). In this context, numerous TSPO-targeted positron emission tomography (PET) tracers have been developed. Among them, several radioligands have advanced to clinical research studies. In this review, we will overview the recent development of TSPO PET tracers, focusing on the radioligand design, radioisotope labeling, pharmacokinetics, and PET imaging evaluation. Additionally, we will consider current limitations, as well as translational potential for future application of TSPO radiopharmaceuticals. This review aims to not only present the challenges in current TSPO PET imaging, but to also provide a new perspective on TSPO targeted PET tracer discovery efforts. Addressing these challenges will facilitate the translation of TSPO in clinical studies of neuroinflammation associated with central nervous system diseases.

Monoacylglycerol lipase (MAGL) is a serine hydrolase, which metabolizes 2-arachidonoylglycerol (2-AG) to arachidonic acid (AA) in the central nervous system (CNS). Dysfunction of MAGL has been associated with multiple CNS disorders and symptoms, including neuroinflammation, cognitive impairment, epileptogenesis, nociception and neurodegenerative diseases. Inhibition of MAGL provides a promising therapeutic direction for the treatment of these conditions, and a MAGL positron emission tomography (PET) probe would greatly facilitate preclinical and clinical development of MAGL inhibitors. Herein, we design and synthesize a small library of fluoropyridyl-containing MAGL inhibitor candidates. Pharmacological evaluation of these candidates by activity-based protein profiling identified 14 as a lead compound, which was then radiolabeled with fluorine-18 via a facile SNAr reaction to form 2-[18F]fluoropyridinyl scaffold. Excellent blood−brain barrier penetration and high in vivo binding specificity was validated for radioligand [18F]14 (also named as [18F]MAGL-1902). This work may serve as a roadmap for clinical translation and further design of potent 18F-labeled MAGL PET tracers. KEY WORDS Monoacylglycerol lipase (MAGL); Central nervous system (CNS); 2-Arachidonylglycerol (2-AG); Arachidonic acid (AA); Positron emission tomography (PET); Fluorine-18

Purpose This study aimed to develop a chelate-free radiolabeled nanoparticle platform for simultaneous positron emission tomography (PET) and magnetic resonance (MR) imaging that provides contrast-enhanced diagnostic imaging and significant image quality gain by integrating the high spatial resolution of MR with the high sensitivity of PET. Methods A commercially available super-paramagnetic iron oxide nanoparticle (SPION) (Feraheme®, FH) was labeled with the [89Zr]Zr using a novel chelate-free radiolabeling technique, heat-induced radiolabeling (HIR). Radiochemical yield (RCY) and purity (RCP) were measured using size exclusion chromatography (SEC) and radio-thin layer chromatography (radio-TLC). Characterization of the non-radioactive isotope 90Zr-labeled FH was performed by transmission electron microscopy (TEM). Simultaneous PET-MR phantom imaging was performed with different 89Zr-FH concentrations. The MR quantitative image analysis determined the contrast-enhancing properties of FH. The signal-to-noise ratio (SNR) and full-width half-maximum (FWHM) of the line spread function (LSF) were calculated before and after co-registering the PET and MR image data. Results High RCY (92%) and RCP (98%) of the [89Zr]Zr-FH product was achieved. TEM analysis confirmed the 90Zr atoms adsorption onto the SPION surface (≈ 10% average radial increase). Simultaneous PET-MR scans confirmed the capability of the [89Zr]Zr-FH nano-platform for this multi-modal imaging technique. Relative contrast image analysis showed that [89Zr]Zr-FH can act as a dual-mode T1/T2 contrast agent. For co-registered PET-MR images, higher spatial resolution (FWHM enhancement ≈ 3) and SNR (enhancement ≈ 8) was achieved at a clinical dose of radio-isotope and Fe. Conclusion Our results demonstrate FH is a highly suitable SPION-based platform for chelate-free labeling of PET tracers for hybrid PET-MR. The high RCY and RCP confirmed the robustness of the chelate-free HIR technique. An overall image quality gain was achieved compared to PET- or MR-alone imaging with a relatively low dosage of [89Zr]Zr-FH. Additionally, FH is suitable as a dual-mode T1/T2 MR image contrast agent., Video abstract Point your SmartPhone at the code above. If you have a QR code reader the video abstract will appear. Or use: http://youtu.be/Me_QBfX7I3s

Monoacylglycerol lipase (MAGL) is a serine hydrolase that degrades 2-arachidonoylglycerol (2-AG) in the endocannabinoid system (eCB). Selective inhibition of MAGL has emerged as a potential therapeutic approach for the treatment of diverse pathological conditions, including chronic pain, inflammation, cancer, and neurodegeneration. Herein, we disclose a novel array of reversible and irreversible MAGL inhibitors by means of “tail switching” on a piperazinyl azetidine scaffold. We developed a lead irreversible-binding MAGL inhibitor 8 and reversible-binding compounds 17 and 37, which are amenable for radiolabeling with 11C or18F. [11C]8 ([11C]MAGL-2-11) exhibited high brain uptake and excellent binding specificity in the brain toward MAGL. Reversible radioligands [11C]17 ([11C]PAD) and [18F]37 ([18F]MAGL-4-11) also demonstrated excellent in vivo binding specificity toward MAGL in peripheral organs. This work may pave the way for the development of MAGL-targeted positron emission tomography tracers with tunability in reversible and irreversible binding mechanisms.

Muscarinic acetylcholine receptors (mAChRs) have five distinct subunits (M1–M5) and are involved in the action of the neurotransmitter acetylcholine in the central and peripheral nervous system. Attributed to the promising clinical efficacy of xanomeline, an M1/M4‐preferring agonist, in patients of schizophrenia and Alzheimer's disease, M1‐ or M4‐selective mAChR modulators have been developed that target the topographically distinct allosteric sites. Herein we report the synthesis and preliminary evaluation of 11C‐labeled positron emission tomography (PET) ligands based on a validated M4R positive allosteric modulator VU0467485 (AZ13713945) to facilitate drug discovery. [11C]VU0467485 and two other ligands were prepared in high radiochemical yields (>30 %, decay‐corrected) with high radiochemical purity (>99 %) and high molar activity (>74 GBq μmol−1). In vitro autoradiography studies indicated that these three ligands possess moderate‐to‐high in vitro specific binding to M4R. Nevertheless, further physiochemical property optimization is necessary to overcome the challenges associated with limited brain permeability.

Positron Emission Tomography (PET) Imaging of Metabotropic Glutamate Receptor Subtype 2 (mGlu2) Based on a Negative Allosteric Modulator Radioligand

Objectives: We have identified 4-hydroxy-6-(3-methoxyphenethyl)pyridazin-3(2H)-one (1) as a lead molecule for D-amino acid oxidase (DAAO)-specific PET ligand. It has been reported that inhibitory activity of compound 1 for human, mouse and rat DAAOs showed dose-dependent responses with IC50 valves of 8.8, 4.4 and 5.6 nM, respectively. [1] The goal of the project was to synthesize the standard and precursor molecules for this target and perform radiolabeling of compound 1 with carbon-11. Methods: The leading compound 1 was synthesized in the following procedures. The cyclization of 3,4,6-trichloropyridazine 2 and pyrocatechol 3 led to intermediate 4 in 80% yield. Through the Sonagashira coupling reaction of compound 4 and 1-ethynyl-3-methoxybenzene 5, the alkyne 6 was obtained in 40% yield. Exchanged with phenylmethanol under base, compound 6 was converted to benzyl-protected ether 7 in 58% yield. After Pd-mediated hydrogenation of compound 7, the desired product 1 was obtained in 77% yield (Scheme 1A). The synthesis of the labeling precursor for compound 1 was showed in Scheme 1B. Phenylmethanol was used to react with compound 4 to get benzyl-protected ether 8 in 48% yield. After hydrogenation of compound 8 under hydrogen using palladium/carbon catalysis, 6-chloro-4-hydroxypyridazin-3(2H)-one 9 was obtained in 84% yield. Hydroxy groups were then protected by the treatment with chloromethyl methyl ether to give compound 10 in 40% yield, followed by the Sonagashira coupling reaction with alkyne 12 to produce compound 13 in 38% yield. At the end, benzyl protecting group was removed by palladium hydroxide/carbon catalysis under hydrogen to yield the precursor 14 in 71% yield. Preliminary 11C-labeling was carried out using precursor (14; 1.6 mg) with [11C]methyl iodide in the presence of Cs2CO3 in DMF (0.4 mL) at 80°C for 5 min to generate [11C]15 (Scheme 1C). Then, 4 M HCl in dioxane was added to remove the MOM group and the mixture was neutralized with NaHCO3 aqueous solution to get [11C]1. Results: The synthesis of candidate compound 1 was obtained from commercially available 3,4,6-trichloropyridazine 2 and pyrocatechol 3 in four steps with overall yield of 14%. The radioligand [11C]1 was synthesized in 13 ± 3% radiochemical yield based on [11C]CH3I (decay corrected). The tracer was obtained at end-of-synthesis (70 min synthesis time) with high radiochemical purity (>99%) and molar activity (> 1.0 Ci/μmol; 37 GBq/μmol). No radiolysis was observed within 90 min. Conclusions: We have successfully prepared one potent and selective DAAO ligand and performed 11C-labeling with a reasonable radiochemical yield and high molar activity. Preliminary in vivo evaluation by PET are underway. References: [1] T. Hondo, M. Warizaya, T. Niimi, I. Namatame, T. Yamaguchi, K. Nakanishi, T. Hamajima, K. Harada, H. Sakashita, Y. Matsumoto, J. Med. Chem. 2013, 56, 3582., SNMMI 2020 Annual Meeting

Purpose Using our chelate-free, heat-induced radiolabeling (HIR) method, we show that a wide range of metals, including those with radioactive isotopologues used for diagnostic imaging and radionuclide therapy, bind to the Feraheme (FH) nanoparticle (NP), a drug approved for the treatment of iron anemia. Material and methods FH NPs were heated (120°C) with nonradioactive metals, the resulting metal-FH NPs were characterized by inductively coupled plasma mass spectrometry (ICP-MS), dynamic light scattering (DLS), and r1 and r2 relaxivities obtained by nuclear magnetic relaxation spectrometry (NMRS). In addition, the HIR method was performed with [90Y]Y3+, [177Lu]Lu3+, and [64Cu]Cu2+, the latter with an HIR technique optimized for this isotope. Optimization included modifying reaction time, temperature, and vortex technique. Radiochemical yield (RCY) and purity (RCP) were measured using size exclusion chromatography (SEC) and thin-layer chromatography (TLC). Results With ICP-MS, metals incorporated into FH at high efficiency were bismuth, indium, yttrium, lutetium, samarium, terbium and europium (>75% @ 120 oC). Incorporation occurred with a small (less than 20%) but statistically significant increases in size and the r2 relaxivity. An improved HIR technique (faster heating rate and improved vortexing) was developed specifically for copper and used with the HIR technique and [64Cu]Cu2+. Using SEC and TLC analyses with [90Y]Y3+, [177Lu]Lu3+ and [64Cu]Cu2+, RCYs were greater than 85% and RCPs were greater than 95% in all cases. Conclusion The chelate-free HIR technique for binding metals to FH NPs has been extended to a range of metals with radioisotopes used in therapeutic and diagnostic applications. Cations with f-orbital electrons, more empty d-orbitals, larger radii, and higher positive charges achieved higher values of RCY and RCP in the HIR reaction. The ability to use a simple heating step to bind a wide range of metals to the FH NP, a widely available approved drug, may allow this NP to become a platform for obtaining radiolabeled nanoparticles in many settings., Video abstract Point your SmartPhone at the code above. If you have a QR code reader the video abstract will appear. Or use: https://youtu.be/_SYXGLeZuWM

表記のタイトルにてポスター発表を行った。, ISRS2019