Fully Automated Cassette-Based Synthesis of 2-Deoxy-2-[18F]Fluorocellobiose Using Trasis AllInOne Module.

Due to the continuous rise in global incidence and severity of invasive fungal infections (IFIs), particularly among immunocompromised and immunodeficient patients, there is an urgent demand for swift and accurate fungal pathogen diagnosis. Therefore, the need for fungal-specific positron emission tomography (PET) imaging agents that can detect the infection in the early stages is increasing. Cellobiose, a disaccharide, is readily metabolized by fungal pathogens such as Aspergillus species. Recently, our group reported fluorine-18 labeled cellobiose, 2-deoxy-2-[18F]fluorocellobiose ([18F]FCB), for specific imaging of Aspergillus infection. The positive imaging findings with very low background signal on delayed imaging make this ligand a promising fungal-specific imaging ligand. Inspired by this result, the decision was made to automate the radiolabeling procedure for better reproducibility and to facilitate clinical translation. A Trasis AllInOne (Trasis AIO) automated module was used for this purpose. The reagent vials contain commercially available 2-deoxy-2-[18F]fluoroglucose ([18F]FDG), glucose-1-phosphate, and enzyme (cellobiose phosphorylase). A Sep-Pak cartridge was used to purify the tracer. The overall radiochemical yield was 50%-70% (n = 6, decay corrected) in 75-min synthesis time with a radiochemical purity of > 98%. This is a highly reliable protocol to produce current good manufacturing practice (cGMP)-compliant [18F]FCB for clinical PET imaging.

Design and radiosynthesis of class-IIa HDAC inhibitor with high molar activity via repositioning the 18F-radiolabel.

The design and radiosynthesis of [18F]NT376, a high potency inhibitor of class-IIa histone deacetylases (HDAC) is reported. We utilized a three-step radiochemical approach that led to the radiosynthesis of [18F]NT376 in a good radiochemical yield, (17.0 ± 3%, decay corrected), high radiochemical purity (> 97%) and relatively high molar activity of 185.0 GBq/µmol (> 5.0 Ci/µmol). The repositioning of the 18F-radiolabel into a phenyl ring (18F-Fluoro-aryl) of the class-IIa HDAC inhibitor avoided the shortcomings of the direct radiolabeling of the 5-trifluoromethyl-1,2,4-oxadiazole moiety that was reported by us previously and was associated with low molar activity (0.74-1.51 GBq/µmol, 20-41 mCi/µmol). This radiochemical approach could find a wider application for radiolabeling similar molecules with good radiochemical yield and high molar activity.

Radiation Chemistry and Radiation Research: A History from the Beginning to the Platinum Edition.

At the dawn of the 20th Century, the underlying chemistry that produced the observed effects of ionizing radiation, e.g., X rays and Radium salts, on aqueous solutions was either unknown or restricted to products found postirradiation. For example, the Curies noted that sealed aqueous solutions of Radium inexplicably decomposed over time, even when kept in the dark. By 1928 there were numerous papers describing the phenomenological effects of ionizing radiation on a wide variety of materials, including the irradiated hands of early radiologists. One scientist who became intensely interested in these radiation effects was Hugo Fricke (Fricke Dosimetry) who established a laboratory in 1928 dedicated to studies on chemical effects of radiation, the results of which he believed were necessary to understand observed radiobiological effects. In this Platinum Issue of Radiation Research (70 years of continuous publication), we present the early history of the development of radiation chemistry and its contributions to all levels of mechanistic radiobiology. We summarize its development as one of the four disciplinary pillars of the Radiation Research Society and its Journal, Radiation Research, founded during the period 1952-1954. In addition, the work of scientists who contributed substantially to the discipline of Radiation Chemistry and to the birth, life and culture of the Society and its journal is presented. In the years following 1954, the increasing knowledge about the underlying temporal and spatial properties of the species produced by various types of radiation is summarized and related to its radiobiology and to modern technologies (e.g., pulsed radiolysis, electron paramagnetic resonance) which became available as the discipline of radiation chemistry developed. A summary of important results from these studies on Radiation Chemistry/Biochemistry in the 20th and 21st Century up to the present time is presented. Finally, we look into the future to see what possible directions radiation chemistry studies might take, based upon promising current research. We find at least two possible directions that will need radiation chemistry expertise to ensure proper experimental design and interpretation of data. These are FLASH radiotherapy, and mechanisms underlying the effects of low doses of radiation delivered at low dose rates. Examples of how radiation chemists could provide beneficial input to these studies are provided.

Improved Radiosynthesis and Automation of [11C]2-(2,6-Difluoro-4-((2-(N-methylphenylsulfonamido)ethyl)thio)phenoxy)acetamide ([11C]K2) for Positron Emission Tomography of the Glutamate α-Amino-3-hydroxy-5-methyl-4-isoxazole Propionic Acid (AMPA) Receptor.

A new automated radiosynthesis of [11C]2-(2,6-difluoro-4-((2-(N-methylphenylsulfonamido)ethyl)thio)phenoxy)acetamide ([11C]K2), a radiopharmaceutical for the glutamate α-amino-3-hydroxy-5-methyl-4-isoxazole propionic acid (AMPA) receptor, is reported. Although manual syntheses have been described, these are unsuitable for routine production of larger batches of [11C]K2 for (pre)clinical PET imaging applications. To meet demands for the imaging agent from our functional neuroimaging collaborators, herein, we report a current good manufacturing practice (cGMP)-compliant synthesis of [11C]K2 using a commercial synthesis module. The new synthesis is fully automated and has been validated for clinical use. The total synthesis time is 33 min from end of bombardment, and the production method provides 2.66 ± 0.3 GBq (71.9 ± 8.6 mCi) of [11C]K2 in 97.7 ± 0.5% radiochemical purity and 754.1 ± 231.5 TBq/mmol (20,382.7 ± 6256.1 Ci/mmol) molar activity (n = 3). Batches passed all requisite quality control testing confirming suitability for clinical use.

Synthesis of 64Cu-, 55Co-, and 68Ga-Labeled Radiopharmaceuticals Targeting Neurotensin Receptor-1 for Theranostics: Adjusting In Vivo Distribution Using Multiamine Macrocycles.

The development of theranostic radiotracers relies on their binding to specific molecular markers of a particular disease and the use of corresponding radiopharmaceutical pairs thereafter. This study reports the use of multiamine macrocyclic moieties (MAs), as linkers or chelators, in tracers targeting the neurotensin receptor-1 (NTSR-1). The goal is to achieve elevated tumor uptake, minimal background interference, and prolonged tumor retention in NTSR-1-positive tumors. Methods: We synthesized a series of neurotensin antagonists bearing MA linkers and metal chelators. The MA unit is hypothesized to establish a strong interaction with the cell membrane, and the addition of a second chelator may enhance water solubility, consequently reducing liver uptake. Small-animal PET/CT imaging of [64Cu]Cu-DOTA-SR-3MA, [64Cu]Cu-NT-CB-NOTA, [68Ga]Ga-NT-CB-NOTA, [64Cu]Cu-NT-CB-DOTA, and [64Cu]Cu-NT-Sarcage was acquired at 1, 4, 24, and 48 h after injection using H1299 tumor models. [55Co]Co-NT-CB-NOTA was also tested in HT29 (high NTSR-1 expression) and Caco2 (low NTSR-1 expression) colorectal adenocarcinoma tumor models. Saturation binding assay and internalization of [55Co]Co-NT-CB-NOTA were used to test tracer specificity and internalization in HT29 cells. Results: In vivo PET imaging with [64Cu]Cu-NT-CB-NOTA, [68Ga]Ga-NT-CB-NOTA, and [55Co]Co-NT-CB-NOTA revealed high tumor uptake, high tumor-to-background contrast, and sustained tumor retention (≤48 h after injection) in NTSR-1-positive tumors. Tumor uptake of [64Cu]Cu-NT-CB-NOTA remained at 76.9% at 48 h after injection compared with uptake 1 h after injection in H1299 tumor models, and [55Co]Co-NT-CB-NOTA was retained at 60.2% at 24 h compared with uptake 1 h after injection in HT29 tumor models. [64Cu]Cu-NT-Sarcage also showed high tumor uptake with low background and high tumor retention 48 h after injection Conclusion: Tumor uptake and pharmacokinetic properties of NTSR-1-targeting radiopharmaceuticals were greatly improved when attached with different nitrogen-containing macrocyclic moieties. The study results suggest that NT-CB-NOTA labeled with either 64Cu/67Cu, 55Co/58mCo, or 68Ga (effect of 177Lu in tumor to be determined in future studies) and NT-Sarcage labeled with 64Cu/67Cu or 55Co/58mCo may be excellent diagnostic and therapeutic radiopharmaceuticals targeting NTSR-1-positive cancers. Also, the introduction of MA units to other ligands is warranted in future studies to test the generality of this approach.

A third generation PSMA-targeted agent [211At]YF2: Synthesis and in vivo evaluation.

INTRODUCTION: Targeted α-particle therapy agents have shown promising responses in patients who have developed resistance to ß--particle emitting radionuclides, albeit off-target toxicity remains a concern. Astatine-211 emits only one α-particle per decay and may alleviate the toxicity from α-emitting daughter radionuclides. Previously, we developed the low-molecular-weight PSMA-targeted agent [211At]L3-Lu that showed suitable therapeutic efficacy and was well tolerated in mice. Although [211At]L3-Lu had good characteristics, we now have evaluated a closely related analogue, [211At]YF2, to determine the better molecule for clinical translation. METHODS: The tin precursors and unlabeled iodo standards for [211At]YF2 and [211At]L3-Lu each were synthesized and a new one-step labeling method was developed to produce [211At]YF2 and [211At]L3-Lu from the respective tin precursor. RCY and RCP were determined using RP-HPLC. Cell uptake, internalization and in vitro cell-killing (MTT) assays were performed on PSMA+ PC-3 PIP cells in parallel experiments to compare [211At]YF2 and [211At]L3-Lu directly. A paired-label biodistribution study was performed in athymic mice with subcutaneous PSMA-positive PC-3 PIP xenografts as a head-to-head comparison of [131I]YF2 and [125I]L3-Lu. The tissue distribution of [211At]YF2 and [211At]L3-Lu were determined individually in the same animal model. RESULTS: The syntheses of tin precursors and unlabeled iodo standards were accomplished in reasonable yields. A streamlined and scalable radiolabeling method (1 h total synthesis time) was developed for the radiosynthesis of both [211At]YF2 and [211At]L3-Lu with 86 ± 7 % (n = 10) and 87 ± 5 % (n = 7) RCY, respectively, and > 95 % RCP for both. The maximum activity of [211At]YF2 produced to date was 666 MBq. An alternative method that did not involve HPLC purification was developed that provided similar RCY and RCP. Significantly higher cell uptake, internalization and cytotoxicity was seen for [211At]YF2 compared with [211At]L3-Lu. Significantly higher uptake and longer retention in tumor was seen for [131I]YF2 than for co-administered [125I]L3-Lu, while considerably higher renal uptake was seen for [131I]YF2. The biodistribution of [211At]YF2 was consistent with that of [131I]YF2. CONCLUSION: [211At]YF2 exhibited higher cellular uptake, internalization and cytotoxicity than [211At]L3-Lu on PSMA-positive PC3 PIP cells. Likewise, higher uptake and longer retention in tumor was seen for [211At]YF2. Experiments to evaluate the dosimetry and therapeutic efficacy of [211At]YF2 are under way.

Synthesis and evaluation of a 68Ga-labeled spermine derivative for tumor PET imaging.

BACKGROUND: The polyamine transporter system (PTS), which renders it a promising target for tumor therapy and imaging applications, facilitates the transmembrane transport of polyamines. We reported a novel derivative of spermine labeled with gallium-68 ([68Ga]Ga-NOTA-Spermine) for the imaging of the PTS in mouse models of tumor. RESULTS: The radiochemical yield of [68Ga]Ga-NOTA-Spermine was determined to be 64-69 %, demonstrating exceptional stability and radiochemical purity (>98 %). Cellular uptake experiments revealed that A549 cells exhibited peak uptake of [68Ga]Ga-NOTA-Spermine at 90 min (15.4 % ± 0.68 %). Biodistribution analysis demonstrated significant accumulation of [68Ga]Ga-NOTA-Spermine in kidneys and liver, while exhibiting low uptake levels in muscle, brain, and bones. Furthermore, Micro-PET/CT scans conducted on A549 tumor-bearing mouse models indicated substantial uptake of [68Ga]Ga-NOTA-Spermine, with maximum tumor/muscle (T/M) ratios reaching 3.71. CONCLUSION: These results suggest that [68Ga]Ga-NOTA-Spermine holds potential as a PET imaging agent for tumors with high levels of PTS.

Application of copper (I) selective ligands for PET imaging of reactive oxygen species through metabolic trapping.

INTRODUCTION: Reactive oxygen species (ROS) are attractive targets for clinical PET imaging. In this study, we hypothesized that PET imaging of ROS would be possible by using chelating ligands (L) that form stable complexes with copper (I) but not with copper (II), based on metabolic trapping. Namely, when [64Cu][CuI(L)2]+ is oxidized by ROS, the oxidized complex will release [64Cu]Cu2+. Then, the released [64Cu]Cu2+ will be trapped inside the cell, resulting in PET signal depending on the redox potential of ROS. To examine the potential of this novel molecular design for ROS imaging, we synthesized copper (I) complexes with bicinchoninic acid (BCA) disodium salt and bathocuproinedisulfonic acid (BCS) disodium salt and evaluated their reactivity with several kinds of ROS. In addition, the cellular uptake of [64Cu][CuI(BCS)2]3- and the stability of [64Cu][CuI(BCS)2]3- in a biological condition were also evaluated. METHODS: [64Cu]Cu2+ was reduced to [64Cu]Cu+ by ascorbic acid and coordinated with BCA and BCS in the acetate buffer to synthesize [64Cu][CuI(BCA)2]3- and [64Cu][CuI(BCS)2]3-. The radiochemical yields were determined by thin-layer chromatography (TLC). After [64Cu][CuI(BCS)2]3- was incubated with hydroxyl radical, lipid peroxide, superoxide, and hydrogen peroxide, the percentage of released [64Cu]Cu2+ from the parent complex was evaluated by TLC. HT-1080 human fibrosarcoma cells were treated with 0.1 % Dimethyl sulfoxide (control), imidazole ketone erastin (IKE), or IKE + ferrostatin-1 (Fer-1). Then, the uptake of [64Cu][CuI(BCS)2]3- to HT-1080 cells in each group was evaluated as %Dose/mg protein. Lastly, [64Cu][CuI(BCS)2]3- was incubated in human plasma, and its intact ratio was determined by TLC. RESULTS: The radiochemical yield of [64Cu][CuI(BCS)2]3- (86 ± 1 %) was higher than that of [64Cu][CuI(BCA)2]3- (44 ± 3 %). [64Cu][CuI(BCA)2]3- was unstable and partially decomposed on TLC. After [64Cu][CuI(BCS)2]3- was reacted with hydroxyl radical, lipid peroxide, and superoxide, 67 ± 2 %, 44 ± 13 %, and 22 ± 3 % of total radioactivity was detected as [64Cu]Cu2+, respectively. On the other hand, the reaction with hydrogen peroxide did not significantly increase the ratio of [64Cu]Cu2+ (4 ± 1 %). These results suggest that [64Cu][CuI(BCS)2]3- could be used for detecting high-redox-potential ROS such as hydroxyl radical and lipid peroxide with high selectivity. The cellular uptake values of [64Cu][CuI(BCS)2]3- in the control, IKE, and Fer-1 group were 42 ± 2, 54 ± 2, and 47 ± 5 %Dose/mg protein (n = 3), respectively, suggesting the ROS specific uptake of [64Cu][CuI(BCS)2]3-. On the other hand, the intact ratio after the incubation of [64Cu][CuI(BCS)2]3- in human plasma was 9 ± 5 %. CONCLUSION: PET imaging of ROS would be possible by using a copper (I) selective ligand, based on metabolic trapping. Although improvement of the membrane permeability and the stability of copper (I) complexes is required, the present results pave the way for the development of novel 64Cu-labeled complexes for PET imaging of ROS.

Study of activation cross sections of double neutron capture reaction on 193Ir for the reactor production route of radiotherapeutic 195mPt.

The radiotherapeutic 195mPt is among the most effective Auger electron emitters of the currently studied radionuclides that have a potential theranostic application in nuclear medicine. Production of 195mPt through double neuron capture of enriched 193Ir followed by ß--decay to the radioisotope of interest carried out at the research reactor IBR-2 is described. Because of the high radiation background, radiochemical purification procedure of 195mPt from bulk of iridium was needed to be developed and is detailed here as well. For the first time, cross section and resonance integral for the reaction 194Ir(n,γ)195mIr were determined. Resonance neutrons contribution was established to exceed that of thermal neutrons, and resonance integral for the reaction 194Ir(n,γ)195mIr is calculated to be 2900 b. Specific activity of 195mPt was estimated to reach a value of 38.7 GBq/(g Pt) at IBR-2 by the end of bombardment (EOB).

Development of 225Ac/213Bi generator based on α-ZrP-PAN composite for targeted alpha therapy.

BACKGROUND: Radioligand therapy using alpha emitters has gained more and more prominence in the last decade. Despite continued efforts to identify new appropriate radionuclides, the combination of 225Ac/213Bi remains among the most promising. Bismuth-213 has been employed in clinical trials in combination with appropriate vectors to treat patients with various forms of cancer, such as leukaemia, bladder cancer, neuroendocrine tumours, melanomas, gliomas, or lymphomas. However, the half-life of 213Bi (T½ = 46 min) implies that its availability for clinical use is limited to hospitals possessing a 225Ac/213Bi radionuclide generator, which is still predominantly scarce. We investigated a new Ac/Bi generator system based on using the composite sorbent α-ZrP-PAN (zirconium(IV) phosphate as active component and polyacrylonitrile as matrix). The developed 225Ac/213Bi generator was subjected to long-term testing after its development. The elution profile was determined and the elution yield, the contamination of the eluate with the parent 225Ac and the contamination of the eluate with the column material were monitored over time. RESULTS: The high activity (75 MBq of parent 225Ac) generator with a length of 75 mm and a diameter of 4 mm containing the composite sorbent α-ZrP-PAN with a particle size of 0.8 to 1.0 mm as the stationary phase, eluted with a mixture of 10 mM DTPA in 5 mM nitric acid, provided 213Bi with yields ranging from 77 % to 96 % in 2.8 mL of eluate, with parent 225Ac contamination in the order of 10-3 %, up to twenty days of use. CONCLUSION: All the results of the monitored parameters indicate that the composite sorbent α-ZrP-PAN based separation system for the elution of 213Bi is a very promising and functional solution.

Synthesis and preclinical evaluation of novel 18F-vancomycin-based tracers for the detection of bacterial infections using positron emission tomography.

INTRODUCTION: Bacterial infections are a major problem in medicine, and the rapid and accurate detection of such infections is essential for optimal patient outcome. Bacterial infections can be diagnosed by nuclear imaging, but most currently available modalities are unable to discriminate infection from sterile inflammation. Bacteria-targeted positron emission tomography (PET) tracers have the potential to overcome this hurdle. In the present study, we compared three 18F-labelled PET tracers based on the clinically applied antibiotic vancomycin for targeted imaging of Gram-positive bacteria. METHODS: [18F]FB-NHS and [18F]BODIPY-FL-NHS were conjugated to vancomycin. The resulting conjugates, together with our previously developed [18F]PQ-VE1-vancomycin, were tested for stability, lipophilicity, selective binding to Gram-positive bacteria, antimicrobial activity and biodistribution. For the first time, the pharmacokinetic properties of all three tracers were compared in healthy animals to identify potential binding sites. RESULTS: [18F]FB-vancomycin, [18F]BODIPY-FL-vancomycin, and [18F]PQ-VE1-vancomycin were successfully synthesized with radiochemical yields of 11.7%, 2.6%, and 0.8%, respectively. [18F]FB-vancomycin exhibited poor in vitro and in vivo stability and, accordingly, no bacterial binding. In contrast, [18F]BODIPY-FL-vancomycin and [18F]PQ-VE1-vancomycin showed strong and specific binding to Gram-positive bacteria, including methicillin-resistant Staphylococcus aureus (MRSA), which was outcompeted by unlabeled vancomycin only at concentrations exceeding clinically relevant vancomycin blood levels. Biodistribution showed renal clearance of [18F]PQ-VE1-vancomycin and [18F]BODIPY-FL-vancomycin with low non-specific accumulation in muscles, fat and bones. CONCLUSION: Here we present the synthesis and first evaluation of the vancomycin-based PET tracers [18F]BODIPY-FL-vancomycin and [18F]PQ-VE1-vancomycin for image-guided detection of Gram-positive bacteria. Our study paves the way towards real-time bacteria-targeted diagnosis of soft tissue and implant-associated infections that are oftentimes caused by Gram-positive bacteria, even after prophylactic treatment with vancomycin.

Development and first-in-human study of PSMA-targeted PET tracers with improved pharmacokinetic properties.

PURPOSE: A series of new 68Ga-labeled tracers based on [68Ga]Ga-PSMA-617 were developed to augment the tumor-to-kidney ratio and reduce the activity accumulation in bladder, ultimately minimize radiation toxicity to the urinary system. METHODS: We introduced quinoline group, phenylalanine and decanoic acid into different tracers to enhance their lipophilicity, strategically limiting their metabolic pathway through the urinary system. Their binding affinity onto LNCaP cells was determined through in vitro saturation assays and competition binding assays. In vivo metabolic study, PET imaging and biodistribution experiment were performed in LNCaP tumor-bearing B-NSG male mice. The most promising tracer was selected for first-in-human study. RESULTS: Four radiotracers were synthesized with radiochemical purity (RCP) > 95% and molar activity in a range of 20.0-25.5 GBq/µmol. The binding affinities (Ki) of TWS01, TWS02 to PSMA were in the low nanomolar range (< 10 nM), while TWS03 and TWS04 exhibited binding affinities with Ki > 20 nM (59.42 nM for TWS03 and 37.14 nM for TWS04). All radiotracers exhibited high stability in vivo except [68Ga]Ga-TWS03. Micro PET/CT imaging and biodistribution analysis revealed that [68Ga]Ga-TWS02 enabled clear tumor visualization in PET images at 1.5 h post-injection, with higher tumor-to-kidney ratio (T/K, 0.93) and tumor-to-muscle ratio (T/M, 107.62) compared with [68Ga]Ga-PSMA-617 (T/K: 0.39, T/M: 15.01) and [68Ga]Ga-PSMA-11 (T/K: 0.15, T/M: 24.00). In first-in-human study, [68Ga]Ga-TWS02 effectively detected PCa-associated lesions including primary and metastatic lesions, with lower accumulation in urinary system, suggesting that [68Ga]Ga-TWS02 might be applied in the detection of bladder invasion, with minimized radiation toxicity to the urinary system. CONCLUSION: Introduction of quinoline group, phenylalanine and decanoic acid into different tracers can modulate the binding affinity and pharmacokinetics of PSMA in vivo. [68Ga]Ga-TWS02 showed high binding affinity to PSMA, excellent pharmacokinetic properties and clear imaging of PCa-associated lesions, making it a promising radiotracer for the clinical diagnosis of PCa. Moreover, TWS02 with a chelator DOTA could also label 177Lu and 225Ac, which could be used for PCa treatment without significant side effects. TRIAL REGISTRATION: The clinical evaluation of this study was registered On October 30, 2021 at https://www.chictr.org.cn/ (No: ChiCTR2100052545).

An Improved HPLC Separation Method for TSPO Radioligand [11C]ER176 Clinical Production.

Mitochondrial membrane translocator protein 18 kDa (TSPO) expression is increased in activated microglia, established as a plausible target of neuroinflammation imaging. [11C]ER176, specifically binding to TSPO, has been developed as the third generation of radioligand for PET imaging of TSPO, which showed the potential in better quantifying neuroinflammation than its predecessors. In the current study, we developed an automated radiosynthesis with an improved HPLC purification method for [11C]ER176 clinical production. The improved HPLC separation was integrated into the automated production of [11C]ER176 using a reverse phase semi-preparative HPLC column with an isocratic pump and the mixture of methanol and 50 mM ammonium acetate as the mobile phase. The fraction corresponding to [11C]ER176 was collected around 8.5-9.0 min without the risk of getting contaminations from nearby impurities. The automated production process took about 30 min after end of bombardment (EOB) and the quality of the final product [11C]ER176 met all specifications for clinical use based on current US Pharmacopeia and FDA CGMP requirements.

Development of High-Throughput Experimentation Approaches for Rapid Radiochemical Exploration.

Positron emission tomography is a widely used imaging platform for studying physiological processes. Despite the proliferation of modern synthetic methodologies for radiolabeling, the optimization of these reactions still primarily relies on inefficient one-factor-at-a-time approaches. High-throughput experimentation (HTE) has proven to be a powerful approach for optimizing reactions in many areas of chemical synthesis. However, to date, HTE has rarely been applied to radiochemistry. This is largely because of the short lifetime of common radioisotopes, which presents major challenges for efficient parallel reaction setup and analysis using standard equipment and workflows. Herein, we demonstrate an effective HTE workflow and apply it to the optimization of copper-mediated radiofluorination of pharmaceutically relevant boronate ester substrates. The workflow utilizes commercial equipment and allows for rapid analysis of reactions for optimizing reactions, exploring chemical space using pharmaceutically relevant aryl boronates for radiofluorinations, and constructing large radiochemistry data sets.

Synthesis and preclinical evaluation of 11C-labeled 7-Oxo-2,4,5,7-tetrahydro-6H-pyrazolo[3,4-c]pyridine radioligands for RIPK1 positron emission tomography imaging.

Targeting receptor-interacting protein kinase 1 (RIPK1) has emerged as a promising therapeutic strategy for various neurodegenerative disorders. The development of a positron emission tomography (PET) probe for brain RIPK1 imaging could offer a valuable tool to assess therapeutic effectiveness and uncover the neuropathology associated with RIPK1. In this study, we present the development and characterization of two new PET radioligands, [11C]PB218 and [11C]PB220, which have the potential to facilitate brain RIPK1 imaging. [11C]PB218 and [11C]PB220 were successfully synthesized with a high radiochemical yield (34 % - 42 %) and molar activity (293 - 314 GBq/µmol). PET imaging characterization of two radioligands was conducted in rodents, demonstrating that both newly developed tracers have good brain penetration (maximum SUV = 0.9 - 1.0) and appropriate brain clearance kinetic profiles. Notably, [11C]PB218 has a more favorable binding specificity than [11C]PB220. A PET/MR study of [11C]PB218 in a non-human primate exhibited good brain penetration, desirable kinetic properties, and a safe profile, thus supporting the translational applicability of our new probe. These investigations enable further translational exploration of [11C]PB218 for drug discovery and PET probe development targeting RIPK1.

Arginine-Selective Bioconjugation Reagent for Effective 18F-labeling of Native Proteins.

Protein-based 18F-PET tracers offer new possibilities in early disease detection and personalized medicine. Their development relies heavily on the availability and effectiveness of 18F-prosthetic groups. We prepared and evaluated a novel arginine-selective prosthetic group, 4-[18F]fluorophenylglyoxal ([18F]FPG). [18F]FPG was radiosynthesized by a one-pot, two-step procedure with a non-decay-corrected (n.d.c.) isolated radiochemical yield (RCY) of 41 ± 8% (n = 10). [18F]FPG constitutes a generic tool for 18F-labeling of various proteins, including human serum albumin (HSA), ubiquitin, interleukin-2, and interleukin-4 in ∼30-60% n.d.c. isolated RCYs. [18F]FPG conjugation with arginine residues is highly selective, even in the presence of a large excess of lysine, cysteine, and histidine. [18F]FPG protein conjugates are able to preserve the binding affinity of the native proteins while also demonstrating excellent in vivo stability. The [18F]FPG-HSA conjugate has prolonged blood retention, which can be applied as a potential blood pool PET imaging agent. Thus, [18F]FPG is an arginine-selective bioconjugation reagent that can be effectively used for the development of 18F-labeled protein radiopharmaceuticals.

Second generation Al18F-labeled D-amino acid peptide for CXCR4 targeted molecular imaging.

BACKGROUND: The C-X-C chemokine receptor type 4 (CXCR4) is overexpressed in many cancers, e.g. multiple myeloma and acute leukemia, yet solely [68Ga]PentixaFor is used for clinical PET imaging. The aim of this study was to develop and assess a second generation Al18F-labeled D-amino acid peptide based on the viral macrophage inflammatory protein II for CXCR4 targeted molecular imaging. METHODS: We designed a library of monomer and multimer constructs and evaluated their binding affinity for human and mouse CXCR4. Based on these results, we selected the best vector molecule for development of an Al18F-labeled ligand, [18F]AlF-NOTA-2xDV1(c11sc12s), which was further evaluated in a cell-based binding assay to assess its binding properties and specificity for CXCR4. Next, pharmacokinetics and tumor uptake of [18F]AlF-NOTA-2xDV1(c11sc12s) were evaluated in naïve mice and mice with xenografts derived from U87.CXCR4 cells. Finally, we performed an imaging study in a non-human primate to assess the in vivo distribution of this novel radioligand in a species closely related to humans. RESULTS: The lead ligand AlF-NOTA-2xDV1(c11sc12s) showed six-fold higher affinity for human CXCR4 compared to Ga-Pentixafor. The corresponding radiotracer was obtained in a good radiochemical yield of 40.1 ± 13.5 % (n = 4) and apparent molar activity of 20.4 ± 3.3 MBq/nmol (n = 4) after optimization. In U87.CD4.CXCR4 cell binding assays, the total bound fraction of [18F]AlF-NOTA-(2×)DV1(c11sc12s) was 32.4 ± 1.8 %. This fraction could be reduced by 82.5 % in the presence of 75 µM AMD3100. In naïve mice, [18F]AlF-NOTA-2xDV1(c11sc12s) accumulated in organs expressing mouse CXCR4, e.g. the liver (SUVmean (mean standardized uptake value) 75 min p.i. 11.7 ± 0.6), which was blockable by co-injecting AMD3100 (5 mg/kg). In U87.CXCR4 xenografted tumor mice, the tumor uptake of [18F]AlF-NOTA-2xDV1(c11sc12s) remained low (SUVmean 0.5 ± 0.1), but was reduced by co-administration of AMD3100. Surprisingly, [18F]AlF-NOTA-2xDV1(c11sc12s) exhibited a similar biodistribution in a non-human primate as in mice indicating off-target binding of [18F]AlF-NOTA-2xDV1(c11sc12s) in liver tissue. We confirmed that [18F]AlF-NOTA-2xDV1(c11sc12s) is taken up by hepatocytes using in vitro studies and that the uptake can be blocked with AMD3100 and rifampicin, a potent organic anion-transporting-polypeptide (OATP)1B1 and OATP1B3 inhibitor. CONCLUSION: The second generation D-peptide AlF-NOTA-2xDV1(c11sc12s) showed high affinity for human CXCR4 and the corresponding radiotracer was produced in good radiochemical yields. However, [18F]AlF-NOTA-2xDV1(c11sc12s) is not specific for CXCR4 and is also a substrate for OATP1B1 and/or OATP1B3, known to mediate hepatic uptake. Therefore, D-amino acid peptides, based on the viral macrophage inflammatory protein II, are not the prefered vector molecule for the development of CXCR4 targeting molecular imaging tools.

Preclinical validation of a novel brain-penetrant PET ligand for visualization of histone deacetylase 6: a potential imaging target for neurodegenerative diseases.

PURPOSE: Histone deacetylase 6 (HDAC6) has emerged as a therapeutic target for neurodegenerative diseases such as Alzheimer's disease. Noninvasive imaging of HDAC6 in the brain by positron emission tomography (PET) would accelerate research into its roles in these diseases. We recently discovered an 18F-labeled derivative of the selective HDAC6 inhibitor SW-100 ([18F]FSW-100) as a potential candidate for brain HDAC6 radioligand. As a mandatory step prior to clinical translation, we performed preclinical validation of [18F]FSW-100. METHODS: Process validation of [18F]FSW-100 radiosynthesis for clinical use and assessment of preclinical toxicity and radiation dosimetry estimated from mouse distribution data were performed. In vitro selectivity of FSW-100 for 28 common receptors in the brain and HDAC isoforms was characterized. [18F]FSW-100 PET imaging was performed in non-human primates in a conscious state to estimate the feasibility of HDAC6 imaging in humans. RESULTS: Three consecutive validation runs of the automated radiosynthesis gave [18F]FSW-100 injections with radiochemical yields of 12%, and the injections conformed to specified quality control criteria for batch release. No acute toxicity was observed for non-radiolabeled FSW-100 or radioactivity decayed [18F]FSW-100 injection, and the former was negative in the Ames test. The whole-body effective dose estimated from biodistribution in mice was within the range of that of previously reported 18F-radioligands in humans. In vitro selectivity against common receptors and other HDAC isoforms was confirmed. [18F]FSW-100 demonstrated good penetration in monkey brain, and in vivo blocking studies suggested that the uptake was specific. CONCLUSION: These results support the clinical utility of [18F]FSW-100 for in vivo imaging of HDAC6 in the brain.

Selection of radionuclide(s) for targeted alpha therapy based on their nuclear decay properties.

Targeted alpha therapy (TAT) is a promising form of oncology treatment utilising alpha-emitting radionuclides that can specifically accumulate at disease sites. The high energy and high linear energy transfer associated with alpha emissions causes localised damage at target sites whilst minimising that to surrounding healthy tissue. The lack of appropriate radionuclides has inhibited research in TAT. The identification of appropriate radionuclides should be primarily a function of the radionuclide's nuclear decay properties, and not their biochemistry or economic factors since these last two factors can change; however, the nuclear decay properties are fixed to that nuclide. This study has defined and applied a criterion based on nuclear decay properties useful for TAT. This down-selection exercise concluded that the most appropriate radionuclides are: 149 Tb, 211 At/ 211 Po, 212 Pb/ 212 Bi/ 212 Po, 213 Bi/ 213 Po, 224 Ra, 225 Ra/ 225 Ac/ 221 Fr, 226 Ac/ 226 Th, 227 Th/ 223 Ra/ 219 Rn, 229 U, 230 U/ 226 Th, and 253 Fm, the majority of which have previously been considered for TAT. 229 U and 253 Fm have been newly identified and could become new radionuclides of interest for TAT, depending on their decay chain progeny.

Automated radiosynthesis of mGluR5 PET tracer [18F]FPEB from aryl-chloro precursor and validation for clinical application.

The radioligand [18F]FPEB, used for PET imaging of the brain's metabotropic glutamate receptor subtype 5 (mGluR5), undergoes a thorough validation process to ensure its safety, efficacy, and quality for clinical use. The process starts by optimizing the synthesis of [18F]FPEB to achieve high radiochemical yield and purity. This study focuses on optimizing the radiolabeling process using an aryl-chloro precursor and validating the GMP production for clinical applications. Fully automated radiolabeling was achieved via one-step nucleophilic substitution reaction. [18F]FPEB was produced and isolated in high radioactivity and radiochemical purity. Throughout the validation process, thorough quality control measures are implemented. Radiopharmaceutical batch release criteria are established, including testing for physical appearance, filter integrity, pH, radiochemical purity, molar activity, radiochemical identity, chemical impurity, structural identity, stability, residual solvent, sterility, and endotoxin levels. In conclusion, the validation of [18F]FPEB involved a comprehensive process of synthesis optimization, quality control, which ensure the safety, efficacy, and quality of [18F]FPEB, enabling its reliable use in clinical PET. Here, we successfully radiolabeled and validated [18F]FPEB using aryl-chloro precursor according to GMP production for clinical application.