Small animal PET imaging with the 68Ga-labeled pH (low) insertion peptide-like peptide YJL-4 in a triple-negative breast cancer mouse model.

BACKGROUND: The aim of this study was to prepare a novel 68Ga-labeled pH (low) insertion peptide (pHLIP)-like peptide, YJL-4, and determine its value for the early diagnosis of triple-negative breast cancer (TNBC) via in vivo imaging of tumor-bearing nude mice. The novel peptide YJL-4 was designed using a template-assisted method and synthesized by solid-phase peptide synthesis. After modification with the chelator 1,4,7­triazacyclononane-N,N',N″-triacetic acid (NOTA), the peptide was labeled with 68Ga. Then, the biodistribution of 68Ga-YJL-4 in tumor-bearing nude mice was investigated, and the mice were imaged by small animal positron emission tomography (PET). RESULTS: The radiochemical yield and radiochemical purity of 68Ga-YJL-4 were 89.5 ± 0.16% and 97.95 ± 0.06%, respectively. The biodistribution of 68Ga-YJL-4 in tumors (5.94 ± 1.27% ID/g, 6.72 ± 1.69% ID/g and 4.54 ± 0.58% ID/g at 1, 2 and 4 h after injection, respectively) was significantly greater than that of the control peptide in tumors at the corresponding time points (P < 0.01). Of the measured off-target organs, 68Ga-YJL-4 was highly distributed in the liver and blood. The small animal PET imaging results were consistent with the biodistribution results. The tumors were visualized by PET at 2 and 4 h after the injection of 68Ga-YJL-4. No tumors were observed in the control group. CONCLUSIONS: The novel pHLIP family peptide YJL-4 can adopt an α-helical structure for easy insertion into the cell membrane in an acidic environment. 68Ga-YJL-4 was produced in high radiochemical yield with good stability and can target TNBC tissue. Moreover, the strong concentration of radioactive 68Ga-YJL-4 in the abdomen does not hinder the imaging of early TNBC.

Functionalization of 68Ga-Radiolabeled Nanodiamonds with Octreotide Does Not Improve Tumor-Targeting Capabilities.

Nanodiamonds (NDs) are emerging as a novel nanoparticle class with growing interest in medical applications. The surface coating of NDs can be modified by attaching binding ligands or imaging probes, turning them into multi-modal targeting agents. In this investigation, we assessed the targeting efficacy of octreotide-functionalized 68Ga-radiolabelled NDs for cancer imaging and compared it with the tumor uptake using [68Ga]Ga-DOTA-TOC. In vivo studies in mice bearing AR42J tumors demonstrated the highest accumulation of the radiolabeled functionalized NDs in the liver and spleen, with relatively low tumor uptake compared to [68Ga]Ga-DOTA-TOC. Our findings suggest that, within the scope of this study, functionalization did not enhance the tumor-targeting capabilities of NDs.

Development of New CD38 Targeted Peptides for Cancer Imaging.

PURPOSE: Multiple myeloma (MM) affects over 35,000 patients each year in the US. There remains a need for versatile Positron Emission Tomography (PET) tracers for the detection, accurate staging, and monitoring of treatment response of MM that have optimal specificity and translational attributes. CD38 is uniformly overexpressed in MM and thus represents an ideal target to develop CD38-targeted small molecule PET radiopharmaceuticals to address these challenges. PROCEDURES: Using phage display peptide libraries and pioneering algorithms, we identified novel CD38 specific peptides. Imaging bioconjugates were synthesized using solid phase peptide chemistry, and systematically analyzed in vitro and in vivo in relevant MM systems. RESULTS: The CD38-targeted bioconjugates were radiolabeled with copper-64 (64Cu) with100% radiochemical purity and an average specific activity of 3.3 - 6.6 MBq/nmol. The analog NODAGA-PEG4-SL022-GGS (SL022: Thr-His-Tyr-Pro-Ile-Val-Ile) had a Kd of 7.55 ± 0.291 nM and was chosen as the lead candidate. 64Cu-NODAGA-PEG4-SL022-GGS demonstrated high binding affinity to CD38 expressing human myeloma MM.1S-CBR-GFP-WT cells, which was blocked by the non-radiolabeled version of the peptide analog and anti-CD38 clinical antibodies, daratumumab and isatuximab, by 58%, 73%, and 78%, respectively. The CD38 positive MM.1S-CBR-GFP-WT cells had > 68% enhanced cellular binding when compared to MM.1S-CBR-GFP-KO cells devoid of CD38. Furthermore, our new CD38-targeted radiopharmaceutical allowed visualization of tumors located in marrow rich bones, remaining there for up to 4 h. Clearance from non-target organs occurred within 60 min. Quantitative PET data from a murine disseminated tumor model showed significantly higher accumulation in the bones of tumor-bearing animals compared to tumor-naïve animals (SUVmax 2.06 ± 0.4 versus 1.24 ± 0.4, P = 0.02). Independently, tumor uptake of the target compound was significantly higher (P = 0.003) compared to the scrambled peptide, 64Cu-NODAGA-PEG4-SL041-GGS (SL041: Thr-Tyr-His-Ile-Pro-Ile-Val). The subcutaneous MM model demonstrated significantly higher accumulation in tumors compared to muscle at 1 and 4 h after tracer administration (SUVmax 0.8 ± 0.2 and 0.14 ± 0.04, P = 0.04 at 1 h; SUVmax 0.89 ± 0.01 and 0.09 ± 0.01, P = 0.0002 at 4 h). CONCLUSIONS: The novel CD38-targeted, radiolabeled bioconjugates were specific and allowed visualization of MM, providing a starting point for the clinical translation of such tracers for the detection of MM.

Validity and value of metabolic connectivity in mouse models of ß-amyloid and tauopathy.

Among functional imaging methods, metabolic connectivity (MC) is increasingly used for investigation of regional network changes to examine the pathophysiology of neurodegenerative diseases such as Alzheimer's disease (AD) or movement disorders. Hitherto, MC was mostly used in clinical studies, but only a few studies demonstrated the usefulness of MC in the rodent brain. The goal of the current work was to analyze and validate metabolic regional network alterations in three different mouse models of neurodegenerative diseases (ß-amyloid and tau) by use of 2-deoxy-2-[18F]fluoro-d-glucose positron emission tomography (FDG-PET) imaging. We compared the results of FDG-µPET MC with conventional VOI-based analysis and behavioral assessment in the Morris water maze (MWM). The impact of awake versus anesthesia conditions on MC read-outs was studied and the robustness of MC data deriving from different scanners was tested. MC proved to be an accurate and robust indicator of functional connectivity loss when sample sizes ≥12 were considered. MC readouts were robust across scanners and in awake/ anesthesia conditions. MC loss was observed throughout all brain regions in tauopathy mice, whereas ß-amyloid indicated MC loss mainly in spatial learning areas and subcortical networks. This study established a methodological basis for the utilization of MC in different ß-amyloid and tau mouse models. MC has the potential to serve as a read-out of pathological changes within neuronal networks in these models.

Fragmentation of functional resting state brain networks in a transgenic mouse model of tau pathology: A metabolic connectivity study using [18F]FDG-PET.

In a previous study, regional reductions in cerebral glucose metabolism have been demonstrated in the tauopathy mouse model rTg4510 (Endepols et al., 2022). Notably, glucose hypometabolism was present in some brain regions without co-localized synaptic degeneration measured with [18F]UCB-H. We hypothesized that in those regions hypometabolism may reflect reduced functional connectivity rather than synaptic damage. To test this hypothesis, we performed seed-based metabolic connectivity analyses using [18F]FDG-PET data in this mouse model. Eight rTg4510 mice at the age of seven months and 8 non-transgenic littermates were injected intraperitoneally with 11.1 ± 0.8 MBq [18F]FDG and spent a 35-min uptake period awake in single cages. Subsequently, they were anesthetized and measured in a small animal PET scanner for 30 min. Three seed-based connectivity analyses were performed per group. Seeds were selected for apparent mismatch between [18F]FDG and [18F]UCB-H. A seed was placed either in the medial orbitofrontal cortex, dorsal hippocampus or dorsal thalamus, and correlated with all other voxels of the brain across animals. In the control group, the emerging correlative pattern was strongly overlapping for all three seed locations, indicating a uniform fronto-thalamo-hippocampal resting state network. In contrast, rTg4510 mice showed three distinct networks with minimal overlap. Frontal and thalamic networks were greatly diminished. The hippocampus, however, formed a new network with the whole parietal cortex. We conclude that resting-state functional networks are fragmented in the brain of rTg4510 mice. Thus, hypometabolism can be explained by reduced functional connectivity of brain areas devoid of tau-related pathology, such as the thalamus.

An iodine­labelled Antibody-drug conjugate PET probe for noninvasive monitoring of Nectin-4 expression in urothelial carcinoma.

BACKGROUND AND PURPOSE: Some kinds of antibody-drug conjugate (ADC) with high affinity to Nectin-4 have demonstrated breakthrough progress in the third-line setting for bladder cancer. However, many patients are still difficult to benefit from treatment based on the heterogeneity of tumour. As the most advanced auxiliary treatment technology, treatment visualization can most intuitively predict the effectiveness of drug treatment, and timely detect the occurrence of drug resistance. Among them, nuclear medicine molecular probes play an important role in this field. METHODS: 124/125I-EV was prepared by labelling Enfortumad Vedetin (EV), an ADC drugs widely used in clinic targeted Nectin-4, with Na124/125I using N-bromine succinimide as oxidant. The radiochemical purity was analyzed via radio-TLC and bioactivity was measured by enzyme-linked immunosorbent assay. Cell uptake assay and small-animal PET imaging were performed to verified the specificity and targeting. KEY RESULTS: 124/125I-EV was prepared with high labeling yield and radiochemical purity. ELISA assays demonstrated that 124I-EV maintained the same high bioactivity as EV with significantly higher uptake in SW780 cells (Nectin-4 positive, 4.05 ± 0.32 %IA/5 × 105 cells at 8 h) than that in T24 cells (Nectin-4 negative, 1.34 ± 0.18 %IA/5 × 105 cells, p < 0.001). In PET imaging, 124I-EV had a significantly higher accumulation in SW780 tumour than that in T24 tumour and the uptake in SW780 tumour could be specifically blocked when co-injected with cold EV. The signal-to-noise ratio at the tumour site gradually increased with time, and peaked at 72 h. CONCLUSION AND IMPLICATIONS: 124I-EV was successfully prepared with high specificity and binding affinity of Nectin-4. This radioactive probe completely simulates the internal circulation of ADC drugs and tumour uptake and retention, which will greatly improve the clinical application of ADC therapy.

Design study of a novel geometrical arrangement for an in-beam small animal positron emission tomography scanner.

Objective.We designed a geometrical solution for a small animal in-beam positron emission tomography (PET) scanner to be used in the project SIRMIO (Small animal proton irradiator for research in molecular image-guided radiation-oncology). The system is based on 56 scintillator blocks of pixelated LYSO crystals. The crystals are arranged providing a pyramidal-step shape to optimize the geometrical coverage in a spherical configuration.Approach.Different arrangements have been simulated and compared in terms of spatial resolution and sensitivity. The chosen setup enables us to reach a good trade-off between a solid angle coverage and sufficient available space for the integration of additional components of the first design prototype of the SIRMIO platform. The possibility of moving the mouse holder inside the PET scanner furthermore allows for achieving the optimum placement of the irradiation area for all the possible tumor positions in the body of the mouse. The work also includes a study of the scintillator material where LYSO and GAGG are compared with a focus on the random coincidence noise due to the natural radioactivity of Lutetium in LYSO, justifying the choice of LYSO for the development of the final system.Main results.The best imaging performance can be achieved with a sub-millimeter spatial resolution and sensitivity of 10% in the center of the scanner, as verified in thorough simulations of point sources. The simulation of realistic irradiation scenarios of proton beams in PMMA targets with/without air gaps indicates the ability of the proposed PET system to detect range shifts down to 0.2 mm.Significance.The presented results support the choice of the identified optimal design for a novel spherical in-beam PET scanner which is currently under commissioning for application to small animal proton and light ion irradiation, and which might find also application, e.g. for biological image-guidance in x-ray irradiation.

Effect of depth of interaction resolution on the spatial resolution of SIAT aPET.

Objective.Spatial resolution is a crucial parameter for a positron emission tomography (PET) scanner. The spatial resolution of a high-resolution small animal PET scanner is significantly influenced by the effect of depth of interaction (DOI) uncertainty. The aim of this work is to investigate the impact of DOI resolution on the spatial resolution of a small animal PET scanner called SIAT aPET and determine the required DOI resolution to achieve nearly uniform spatial resolution within the field of view (FOV).Approach. The SIAT aPET detectors utilize 1.0 × 1.0 × 20 mm3crystals, with an average DOI resolution of ∼2 mm. A default number of 16 DOI bins are used during data acquisition. First, a Na-22 point source was scanned in the center of the axial FOV with different radial offsets. Then, a Derenzo phantom was scanned at radial offsets of 0 and 15 mm in the center axial FOV. The measured DOI information was rebinned to 1, 2, 4 and 8 DOI bins to mimic different DOI resolutions of the detectors during image reconstruction.Main results. Significant artifacts were observed in images obtained from both the point source and Derenzo phantom when using only one DOI bin. When accurate measurement of DOI is not achieved, degradation in spatial resolution is more pronounced in the radial direction compared to tangential and axial directions for large radial offsets. The radial spatial resolutions at a 30 mm radial offset are 5.05, 2.62, 1.24, 0.86 and 0.78 mm when using 1, 2, 4, 8, or 16 DOI bins, respectively. The axial spatial resolution improved from ∼1.3 to 0.7 mm as the number of DOI bins increased from 1 to 16 at radial offsets from 0 to 25 mm. Two DOI bins are required to obtain images without significant artifacts. The required DOI resolution is about three times the crystal width of SIAT aPET to achieve a uniform submillimeter spatial resolution within the central 60 mm FOV and resolve the 1 mm rods of the Derenzo phantom at both positions.

Integrated Small Animal PET/CT/RT with Onboard PET/CT Image Guidance for Preclinical Radiation Oncology Research.

We have integrated a compact and lightweight PET with an existing CT image-guided small animal irradiator to enable practical onboard PET/CT image-guided preclinical radiation therapy (RT) research. The PET with a stationary and full-ring detectors has ~1.1 mm uniform spatial resolution over its imaging field-of-view of 8.0 cm diameter and 3.5 cm axial length and was mechanically installed inside the irradiator in a tandem configuration with CT and radiation unit. A common animal bed was used for acquiring sequential dual functional and anatomical images with independent PET and CT control and acquisition systems. The reconstructed dual images were co-registered based on standard multi-modality image calibration and registration processes. Phantom studies were conducted to evaluate the integrated system and dual imaging performance. The measured mean PET/CT image registration error was ~0.3 mm. With one-bed and three-bed acquisitions, initial tumor focused and whole-body [18F]FDG animal images were acquired to test the capability of onboard PET/CT image guidance for preclinical RT research. Overall, the results have shown that integrated PET/CT/RT can provide advantageous and practical onboard PET/CT image to significantly enhance the accuracy of tumor delineation and radiation targeting that should enhance the existing and enable new and potentially breakthrough preclinical RT research and applications.

Rest/stress myocardial perfusion imaging by positron emission tomography with 18F-Flurpiridaz: A feasibility study in mice.

BACKGROUND: Myocardial perfusion imaging by positron emission tomography (PET-MPI) is the current gold standard for quantification of myocardial blood flow. 18F-flurpiridaz was recently introduced as a valid alternative to currently used PET-MPI probes. Nonetheless, optimum scan duration and time interval for image analysis are currently unknown. Further, it is unclear whether rest/stress PET-MPI with 18F-flurpiridaz is feasible in mice. METHODS: Rest/stress PET-MPI was performed with 18F-flurpiridaz (0.6-3.0 MBq) in 27 mice aged 7-8 months. Regadenoson (0.1 µg/g) was used for induction of vasodilator stress. Kinetic modeling was performed using a metabolite-corrected arterial input function. Image-derived myocardial 18F-flurpiridaz uptake was assessed for different time intervals by placing a volume of interest in the left ventricular myocardium. RESULTS: Tracer kinetics were best described by a two-tissue compartment model. K1 ranged from 6.7 to 20.0 mL·cm-3·min-1, while myocardial volumes of distribution (VT) were between 34.6 and 83.6 mL·cm-3. Of note, myocardial 18F-flurpiridaz uptake (%ID/g) was significantly correlated with K1 at rest and following pharmacological vasodilation for all time intervals assessed. However, while Spearman's coefficients (rs) ranged between 0.478 and 0.681, R2 values were generally low. In contrast, an excellent correlation of myocardial 18F-flurpiridaz uptake with VT was obtained, particularly when employing the averaged myocardial uptake from 20 to 40 min post tracer injection (R2 ≥ 0.98). Notably, K1 and VT were similarly sensitive to pharmacological vasodilation induction. Further, mean stress-to-rest ratios of K1, VT, and %ID/g 18F-flurpiridaz were virtually identical, suggesting that %ID/g 18F-flurpiridaz can be used to estimate coronary flow reserve (CFR) in mice. CONCLUSION: Our findings suggest that a simplified assessment of relative myocardial perfusion and CFR, based on image-derived tracer uptake, is feasible with 18F-flurpiridaz in mice, enabling high-throughput mechanistic CFR studies in rodents.

Synthesis, radiolabeling, and preclinical in vivo evaluation of 68Ga-radiolabelled nanodiamonds.

PURPOSE: Nanodiamonds (NDs) represent a new class of nanoparticles and have gained increasing interest in medical applications. Modifying the surface coating by attaching binding ligands or imaging probes can transform NDs into multi-modal targeting probes. This study evaluated the biokinetics and biodistribution of 68Ga-radiolabelled NDs in a xenograft model. PROCEDURES: NDs were coated with an albumin-derived copolymer modified with desferrioxamine to provide a chelator for radiolabeling. In vivo studies were conducted in AR42J tumor-bearing CD1 mice to evaluate biodistribution and tumor accumulation of the NDs. RESULTS: Coated NDs were successfully radiolabeled using 68Ga at room temperature with radiolabeling efficiencies up to 91.8 ± 3.2 % as assessed by radio-TLC. In vivo studies revealed the highest accumulation in the liver and spleen, whereas tumor radioactivity concentration was low. CONCLUSIONS: Radiolabeling of coated NDs could be achieved. However, the obtained results indicate these coated NDs' limitations in their biodistribution within the conducted studies.

Intramuscular [18F]F-FDG Administration for Successful PET Imaging of Golden Hamsters in a Maximum Containment Laboratory Setting.

Positron emission tomography (PET) is becoming an important tool for the investigation of emerging infectious diseases in animal models. Usually, PET imaging is performed after intravenous (IV) radiotracer administration. However, IV injections are difficult to perform in some small animals, such as golden hamsters. This challenge is particularly evident in longitudinal imaging studies, and even more so in maximum containment settings used to study high-consequence pathogens. We propose the use of intramuscular (IM) administration of 2-deoxy-2[18F]fluoro-D-glucose ([18F]F-FDG) for PET imaging of hamsters in a biosafety level 4 (BSL-4) laboratory setting. After [18F]F-FDG administration via IM or IV (through surgically implanted vascular access ports), eight hamsters underwent static or dynamic PET scans. Time-activity curves (TACs) and standardized uptake values (SUVs) in major regions of interest (ROIs) were used to compare the two injection routes. Immediately after injection, TACs differed between the two routes. At 60 min post-injection, [18F]F-FDG activity for both routes reached a plateau in most ROIs except the brain, with higher accumulation in the liver, lungs, brain, and nasal cavities observed in the IM group. IM delivery of [18F]F-FDG is an easy, safe, and reliable alternative for longitudinal PET imaging of hamsters in a BSL-4 laboratory setting.

Radiosynthesis and Preclinical Evaluation of an 18F-Labeled Triazolopyridopyrazine-Based Inhibitor for Neuroimaging of the Phosphodiesterase 2A (PDE2A).

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.

A guideline proposal for mice preparation and care in 18F-FDG PET imaging.

The experimental outcomes of small-animal positron emission tomography (PET) imaging with 18F-labelled fluorodeoxyglucose (18F-FDG) can be particularly compromised by animal preparation and care. Several works intend to improve research reporting and amplify the quality and reliability of published research. Though these works provide valuable information to plan and conduct animal studies, manuscripts describe different methodologies-standardization does not exist. Consequently, the variation in details reported can explain the difference in the experimental results found in the literature. Additionally, the resources and guidelines defining protocols for small-animal imaging are scarce, making it difficult for researchers to obtain and compare accurate and reproducible data. Considering the selection of suitable procedures key to ensure animal welfare and research improvement, this paper aims to prepare the way for a future guideline on mice preparation and care for PET imaging with 18F-FDG. For this purpose, a global standard protocol was created based on recommendations and good practices described in relevant literature.

A quantitative in vivo imaging platform for tracking pathological tau depositions and resultant neuronal death in a mouse model.

PURPOSE: Depositions of tau fibrils are implicated in diverse neurodegenerative disorders, including Alzheimer's disease, and precise assessments of tau pathologies and their impacts on neuronal survival are crucial for pursuing the neurodegenerative tau pathogenesis with and without potential therapies. We aimed to establish an in vivo imaging system to quantify tau accumulations with positron emission tomography (PET) and brain atrophy with volumetric MRI in rTg4510 transgenic mice modeling neurodegenerative tauopathies. METHODS: A total of 91 rTg4510 and non-transgenic control mice underwent PET with a tau radiotracer, 18F-PM-PBB3, and MRI at various ages (1.8-12.3 months). Using the cerebellum as reference, the radiotracer binding in target regions was estimated as standardized uptake value ratio (SUVR) and distribution volume ratio (DVR). Histopathological staining of brain sections derived from scanned animals was also conducted to investigate the imaging-neuropathology correlations. RESULTS: 18F-PM-PBB3 SUVR at 40-60 min in the neocortex, hippocampus, and striatum of rTg4510 mice agreed with DVR, became significantly different from control values around 4-5 months of age, and progressively and negatively correlated with age and local volumes, respectively. Neocortical SUVR also correlated with the abundance of tau inclusions labeled with PM-PBB3 fluorescence, Gallyas-Braak silver impregnation, and anti-phospho-tau antibodies in postmortem assays. The in vivo and ex vivo 18F-PM-PBB3 binding was blocked by non-radioactive PM-PBB3. 18F-PM-PBB3 yielded a 1.6-fold greater dynamic range for tau imaging than its ancestor, 11C-PBB3. CONCLUSION: Our imaging platform has enabled the quantification of tau depositions and consequent neuronal loss and is potentially applicable to the evaluation of candidate anti-tau and neuroprotective drugs.

Assessment of the In Vivo Relationship Between Cerebral Hypometabolism, Tau Deposition, TSPO Expression, and Synaptic Density in a Tauopathy Mouse Model: a Multi-tracer PET Study.

Cerebral glucose hypometabolism is a typical hallmark of Alzheimer's disease (AD), usually associated with ongoing neurodegeneration and neuronal dysfunction. However, underlying pathological processes are not fully understood and reproducibility in animal models is not well established. The aim of the present study was to investigate the regional interrelation of glucose hypometabolism measured by [18F]FDG positron emission tomography (PET) with various molecular targets of AD pathophysiology using the PET tracers [18F]PI-2620 for tau deposition, [18F]DPA-714 for TSPO expression associated with neuroinflammation, and [18F]UCB-H for synaptic density in a transgenic tauopathy mouse model. Seven-month-old rTg4510 mice (n = 8) and non-transgenic littermates (n = 8) were examined in a small animal PET scanner with the tracers listed above. Hypometabolism was observed throughout the forebrain of rTg4510 mice. Tau pathology, increased TSPO expression, and synaptic loss were co-localized in the cortex and hippocampus and correlated with hypometabolism. In the thalamus, however, hypometabolism occurred in the absence of tau-related pathology. Thus, cerebral hypometabolism was associated with two regionally distinct forms of molecular pathology: (1) characteristic neuropathology of the Alzheimer-type including synaptic degeneration and neuroinflammation co-localized with tau deposition in the cerebral cortex, and (2) pathological changes in the thalamus in the absence of other markers of AD pathophysiology, possibly reflecting downstream or remote adaptive processes which may affect functional connectivity. Our study demonstrates the feasibility of a multitracer approach to explore complex interactions of distinct AD-pathomechanisms in vivo in a small animal model. The observations demonstrate that multiple, spatially heterogeneous pathomechanisms can contribute to hypometabolism observed in AD mouse models and they motivate future longitudinal studies as well as the investigation of possibly comparable pathomechanisms in human patients.

Ultra-high-resolution depth-encoding small animal PET detectors: Using GAGG and LYSO crystal arrays.

PURPOSE: Small animal positron emission tomography (PET) scanners are widely used in current biomedical research. The study aimed to develop high-efficiency and ultra-high-resolution detectors that could be used to develop a small animal PET scanner with high sensitivity and spatial resolution approaching to its physical limit. METHODS: Four crystal arrays were fabricated and measured in this study. Crystal arrays 1 and 2 consisted of 38 × 38 gadolinium aluminum gallium garnet (GAGG) and lutetium-yttrium oxyorthosilicate (LYSO) crystals of 0.4 × 0.4 × 20 mm3 size. Crystal array 3 consisted of 16 × 16 GAGG crystals of 0.3 × 0.3 × 20 mm3 size, and crystal array 4 consisted of 24 × 24 LYSO crystals 0.3 × 0.3 × 20 mm3 in size. The crystal arrays were dual-ended readouts using 8 × 8 silicon photomultiplier (SiPM) arrays of 2 × 2 mm2 pixel area. The SiPM array was readout using a signal multiplexing circuit to convert the 64 output signals into four position-encoding signals. The performances of the four detectors in terms of flood histogram, energy resolution, depth of interaction (DOI) resolution, and timing resolution were measured. RESULTS: The GAGG detectors provided better flood histograms, ∼30% higher photopeak amplitude, ∼20% higher energy resolution, ∼12% worse DOI resolution, and ∼15% worse timing resolution compared with LYSO detectors of the same crystal size. These four detectors provided DOI resolutions of <2 mm, energy resolutions of <22%, and timing resolutions of <1.6 ns. All crystals of 0.4 × 0.4 × 20 mm3 and 0.3 × 0.3 × 20 mm3 could be clearly resolved if the crystal array was 1 mm smaller in the four sides than that in the SiPM array. CONCLUSIONS: High DOI resolution PET detectors were developed using both GAGG and LYSO arrays with crystal sizes of 0.3 and 0.4 mm, respectively, and a length of 20 mm. The detectors can be used in the future to develop small animal PET scanners, especially dedicated mouse imaging PET scanners, which can simultaneously achieve high sensitivity and ultra-high spatial resolution.

Discovery, synthesis and evaluation of novel reversible monoacylglycerol lipase radioligands bearing a morpholine-3-one scaffold.

Monoacylglycerol lipase (MAGL) is a serine hydrolase that plays an important role in the endocannabinoid degradation in the brain. It has recently emerged as a promising therapeutic target in the treatment of neuroinflammatory and neurodegenerative diseases, such as multiple sclerosis, Alzheimer's disease and Parkinson's disease. Development of MAGL-specific radioligands for non-invasive imaging by positron-emission tomography (PET) would deepen our knowledge on the relevant pathological changes in diseased states and accelerate drug discovery. In this study, we report the selection and synthesis of two morpholine-3-one derivatives as potential reversible MAGL PET tracer candidates based on their multiparameter optimization scores. Both compounds ([11C]1, [11C]2) were radiolabeled by direct [11C]CO2 fixation and the in vitro autoradiographic studies demonstrated their specificity and selectivity towards MAGL. Dynamic PET imaging using MAGL knockout and wild-type mice confirmed the in vivo specificity of [11C]2. Our preliminary results indicate that morpholine-3-one derivative [11C]2 ([11C]RO7279991) binds to MAGL in vivo, and this molecular scaffold could serve as an alternative lead structure to image MAGL in the central nervous system.

The ultra high sensitivity blood counter: a compact, MRI-compatible, radioactivity counter for pharmacokinetic studies inµl volumes.

Quantification of physiological parameters in preclinical pharmacokinetic studies based on nuclear imaging requires the monitoring of arterial radioactivity over time, known as the arterial input function (AIF). Continuous derivation of the AIF in rodent models is very challenging because of the limited blood volume available for sampling. To address this challenge, an Ultra High Sensitivity Blood Counter (UHS-BC) was developed. The device detects beta particles in real-time using silicon photodiodes, custom low-noise electronics, and 3D-printed plastic cartridges to hold standard catheters. Two prototypes were built and characterized in two facilities. Sensitivities up to 39% for18F and 58% for11C-based positron emission tomography (PET) tracers were demonstrated.99mTc and125I based Single Photon Emission Computed Tomography (SPECT) tracers were detected with greater than 3% and 10% sensitivity, respectively, opening new applications in nuclear imaging and fundamental biology research. Measured energy spectra show all relevant peaks down to a minimum detectable energy of 20 keV. The UHS-BC was shown to be highly reliable, robust towards parasitic background radiation and electromagnetic interference in the PET or MRI environment. The UHS-BC provides reproducible results under various experimental conditions and was demonstrated to be stable over days of continuous operation. Animal experiments showed that the UHS-BC performs accurate AIF measurements using low detection volumes suitable for small animal models in PET, SPECT and PET/MRI investigations. This tool will help to reduce the time and number of animals required for pharmacokinetic studies, thus increasing the throughput of new drug development.

Administration Routes for SSTR-/PSMA- and FAP-Directed Theranostic Radioligands in Mice.

The NETTER-1, VISION, and TheraP trials proved the efficacy of repeat intravenous application of small radioligands. Application by subcutaneous, intraperitoneal, or oral routes is an important alternative and may yield comparable or favorable organ and tumor radioligand uptake. Here, we assessed organ and tumor biodistribution for various radioligand application routes in healthy mice and models of cancer expressing somatostatin receptor (SSTR), prostate-specific membrane antigen (PSMA), and fibroblast activation protein (FAP). Methods: Healthy and tumor-bearing male C57BL/6 or NOD SCID γ-mice, respectively, were administered a mean of 6.0 ± 0.5 MBq of 68Ga-DOTATOC (RM1-SSTR allograft), 5.3 ± 0.3 MBq of 68Ga-PSMA11 (RM1-PSMA allograft), or 4.8 ± 0.2 MBq of 68Ga-FAPI46 (HT1080-FAP xenograft) by intravenous, intraperitoneal, subcutaneous, or oral routes. In vivo PET images and ex vivo biodistribution in tumor, organs, and the injection site were assessed up to 5 h after injection. Healthy mice were monitored for up to 7 d after the last scan for signs of stress or adverse reactions. Results: After intravenous, intraperitoneal, and subcutaneous radioligand administration, average residual activity at the injection site was less than 17 percentage injected activity per gram (%IA/g) at 1 h after injection, less than 10 %IA/g at 2 h after injection, and no more than 4 %IA/g at 4 h after injection for all radioligands. After oral administration, at least 50 %IA/g remained within the intestines until 4 h after injection. Biodistribution in organs of healthy mice was nearly equivalent after intravenous, intraperitoneal, and subcutaneous application at 1 h after injection and all subsequent time points (≤1 %IA/g for liver, blood, and bone marrow; 11.2 ± 1.4 %IA/g for kidneys). In models for SSTR-, PSMA- and FAP-expressing cancer, tumor uptake was increased or equivalent for intraperitoneal/subcutaneous versus intravenous injection at 5 h after injection (ex vivo): SSTR, 7.2 ± 1.0 %IA/g (P = 0.0197)/6.5 ± 1.3 %IA/g (P = 0.0827) versus 2.9 ± 0.3 %IA/g, respectively; PSMA, 3.4 ± 0.8 %IA/g (P = 0.9954)/3.9 ± 0.8 %IA/g (P = 0.8343) versus 3.3 ± 0.7% IA/g, respectively; FAP, 1.1 ± 0.1 %IA/g (P = 0.9805)/1.1 ± 0.1 %IA/g (P = 0.7446) versus 1.0 ± 0.2 %IA/g, respectively. Conclusion: In healthy mice, biodistribution of small theranostic ligands after intraperitoneal/subcutaneous application is nearly equivalent to that after intravenous injection. Subcutaneous administration resulted in the highest absolute SSTR tumor and tumor-to-organ uptake as compared with the intravenous route, warranting further clinical assessment.