Efficient Synthesis and HPLC-Based Characterization for Developing Vanadium-48-Labeled Vanadyl Acetylacetonate as a Novel Cancer Radiotracer for PET Imaging.

Bis(acetylacetonato)oxidovanadium(IV) [(VO(acac)2], generally known as vanadyl acetylacetonate, has been shown to be preferentially sequestered in malignant tissue. Vanadium-48 (48V) generated with a compact medical cyclotron has been used to label VO(acac)2 as a potential radiotracer in positron emission tomography (PET) imaging for the detection of cancer, but requires lengthy synthesis. Current literature protocols for the characterization of VO(acac)2 require macroscale quantities of reactants and solvents to identify products by color and to enable crystallization that are not readily adaptable to the needs of radiotracer synthesis. We present an improved method to produce vanadium-48-labeled VO(acac)2, [48V]VO(acac)2, and characterize it using high-performance liquid chromatography (HPLC) with radiation detection in combination with UV detection. The approach is suitable for radiotracer-level quantities of material. These methods are readily applicable for production of [48V]VO(acac)2. Preliminary results of preclinical, small-animal PET studies are presented.

Evaluation of an Image-Derived Input Function for Kinetic Modeling of Nicotinic Acetylcholine Receptor-Binding PET Ligands in Mice.

Positron emission tomography (PET) radioligands that bind with high-affinity to α4ß2-type nicotinic receptors (α4ß2Rs) allow for in vivo investigations of the mechanisms underlying nicotine addiction and smoking cessation. Here, we investigate the use of an image-derived arterial input function and the cerebellum for kinetic analysis of radioligand binding in mice. Two radioligands were explored: 2-[18F]FA85380 (2-FA), displaying similar pKa and binding affinity to the smoking cessation drug varenicline (Chantix), and [18F]Nifene, displaying similar pKa and binding affinity to nicotine. Time-activity curves of the left ventricle of the heart displayed similar distribution across wild type mice, mice lacking the ß2-subunit for ligand binding, and acute nicotine-treated mice, whereas reference tissue binding displayed high variation between groups. Binding potential estimated from a two-tissue compartment model fit of the data with the image-derived input function were higher than estimates from reference tissue-based estimations. Rate constants of radioligand dissociation were very slow for 2-FA and very fast for Nifene. We conclude that using an image-derived input function for kinetic modeling of nicotinic PET ligands provides suitable results compared to reference tissue-based methods and that the chemical properties of 2-FA and Nifene are suitable to study receptor response to nicotine addiction and smoking cessation therapies.

Accelerator-Based Production of Scandium Radioisotopes for Applications in Prostate Cancer: Toward Building a Pipeline for Rapid Development of Novel Theranostics.

In the field of nuclear medicine, the ß+ -emitting 43Sc and ß- -emitting 47Sc are promising candidates in cancer diagnosis and targeted radionuclide therapy (TRT) due to their favorable decay schema and shared pharmacokinetics as a true theranostic pair. Additionally, scandium is a group-3 transition metal (like 177Lu) and exhibits affinity for DOTA-based chelators, which have been studied in depth, making the barrier to implementation lower for 43/47Sc than for other proposed true theranostics. Before 43/47Sc can see widespread pre-clinical evaluation, however, an accessible production methodology must be established and each isotope's radiolabeling and animal imaging capabilities studied with a widely utilized tracer. As such, a simple means of converting an 18 MeV biomedical cyclotron to support solid targets and produce 43Sc via the 42Ca(d,n)43Sc reaction has been devised, exhibiting reasonable yields. The NatTi(γ,p)47Sc reaction is also investigated along with the successful implementation of chemical separation and purification methods for 43/47Sc. The conjugation of 43/47Sc with PSMA-617 at specific activities of up to 8.94 MBq/nmol and the subsequent imaging of LNCaP-ENZaR tumor xenografts in mouse models with both 43/47Sc-PSMA-617 are also presented.

Trapping of Nicotinic Acetylcholine Receptor Ligands Assayed by In Vitro Cellular Studies and In Vivo PET Imaging.

A question relevant to nicotine addiction is how nicotine and other nicotinic receptor membrane-permeant ligands, such as the anti-smoking drug varenicline (Chantix), distribute in brain. Ligands, like varenicline, with high pKa and high affinity for α4ß2-type nicotinic receptors (α4ß2Rs) are trapped in intracellular acidic vesicles containing α4ß2Rs in vitro Nicotine, with lower pKa and α4ß2R affinity, is not trapped. Here, we extend our results by imaging nicotinic PET ligands in vivo in male and female mouse brain and identifying the trapping brain organelle in vitro as Golgi satellites (GSats). Two PET 18F-labeled imaging ligands were chosen: [18F]2-FA85380 (2-FA) with varenicline-like pKa and affinity and [18F]Nifene with nicotine-like pKa and affinity. [18F]2-FA PET-imaging kinetics were very slow consistent with 2-FA trapping in α4ß2R-containing GSats. In contrast, [18F]Nifene kinetics were rapid, consistent with its binding to α4ß2Rs but no trapping. Specific [18F]2-FA and [18F]Nifene signals were eliminated in ß2 subunit knock-out (KO) mice or by acute nicotine (AN) injections demonstrating binding to sites on ß2-containing receptors. Chloroquine (CQ), which dissipates GSat pH gradients, reduced [18F]2-FA distributions while having little effect on [18F]Nifene distributions in vivo consistent with only [18F]2-FA trapping in GSats. These results are further supported by in vitro findings where dissipation of GSat pH gradients blocks 2-FA trapping in GSats without affecting Nifene. By combining in vitro and in vivo imaging, we mapped both the brain-wide and subcellular distributions of weak-base nicotinic receptor ligands. We conclude that ligands, such as varenicline, are trapped in neurons in α4ß2R-containing GSats, which results in very slow release long after nicotine is gone after smoking.SIGNIFICANCE STATEMENT Mechanisms of nicotine addiction remain poorly understood. An earlier study using in vitro methods found that the anti-smoking nicotinic ligand, varenicline (Chantix) was trapped in α4ß2R-containing acidic vesicles. Using a fluorescent-labeled high-affinity nicotinic ligand, this study provided evidence that these intracellular acidic vesicles were α4ß2R-containing Golgi satellites (GSats). In vivo PET imaging with F-18-labeled nicotinic ligands provided additional evidence that differences in PET ligand trapping in acidic vesicles were the cause of differences in PET ligand kinetics and subcellular distributions. These findings combining in vitro and in vivo imaging revealed new mechanistic insights into the kinetics of weak base PET imaging ligands and the subcellular mechanisms underlying nicotine addiction.

Modelling cyclotron-based production of radioisotopes via TOPAS.

Objective. In this work, the irradiation of natural titanium foils in the beam-stop of a compact medical cyclotron, an IBA CYCLONE 18/9, is simulated to assess the efficacy of using a beam-stop as a target holder, and using two different target geometries, in the production of vanadium-48, a positron-emitting radioisotope with potential utility as a cancer imaging agent in positron emission tomography.Approach. TOPAS, the TOol for PArticle Simulation, a Geant4-based Monte Carlo program, was used to model the cyclotron beam parameters, choose an appropriate physics list, and simulate the irradiation of targets made from foils of 12 or 12.5µm thickness. These simulation yields were compared to theoretical yields calculated using cross section data from the literature, as well as assayed yields from experimental irradiations.Main results.We found that most physics lists in TOPAS overestimate the cross section in the desired energy range (16-20 MeV) by at least 136%, with the exception of those using the Bertini Cascade Model. Compared to assayed yields, TOPAS provided a minimum of 0.4% error for cup-shaped targets and at least a 12% overestimation for sphere-shaped targets.Significance.These simulations provide a tool to help explain irregularities in radioisotope production yield and motivate modifications to increase target yield.

The optimal 18F-fluoromisonidazole PET threshold to define tumor hypoxia in preclinical squamous cell carcinomas using pO2 electron paramagnetic resonance imaging as reference truth.

PURPOSE: To identify the optimal threshold in 18F-fluoromisonidazole (FMISO) PET images to accurately locate tumor hypoxia by using electron paramagnetic resonance imaging (pO2 EPRI) as ground truth for hypoxia, defined by pO2 [Formula: see text] 10 mmHg. METHODS: Tumor hypoxia images in mouse models of SCCVII squamous cell carcinoma (n = 16) were acquired in a hybrid PET/EPRI imaging system 2 h post-injection of FMISO. T2-weighted MRI was used to delineate tumor and muscle tissue. Dynamic contrast enhanced (DCE) MRI parametric images of Ktrans and ve were generated to model tumor vascular properties. Images from PET/EPR/MRI were co-registered and resampled to isotropic 0.5 mm voxel resolution for analysis. PET images were converted to standardized uptake value (SUV) and tumor-to-muscle ratio (TMR) units. FMISO uptake thresholds were evaluated using receiver operating characteristic (ROC) curve analysis to find the optimal FMISO threshold and unit with maximum overall hypoxia similarity (OHS) with pO2 EPRI, where OHS = 1 shows perfect overlap and OHS = 0 shows no overlap. The means of dice similarity coefficient, normalized Hausdorff distance, and accuracy were used to define the OHS. Monotonic relationships between EPRI/PET/DCE-MRI were evaluated with the Spearman correlation coefficient ([Formula: see text]) to quantify association of vasculature on hypoxia imaged with both FMISO PET and pO2 EPRI. RESULTS: FMISO PET thresholds to define hypoxia with maximum OHS (both OHS = 0.728 [Formula: see text] 0.2) were SUV [Formula: see text] 1.4 [Formula: see text] SUVmean and SUV [Formula: see text] 0.6 [Formula: see text] SUVmax. Weak-to-moderate correlations (|[Formula: see text]|< 0.70) were observed between PET/EPRI hypoxia images with vascular permeability (Ktrans) or fractional extracellular-extravascular space (ve) from DCE-MRI. CONCLUSION: This is the first in vivo comparison of FMISO uptake with pO2 EPRI to identify the optimal FMISO threshold to define tumor hypoxia, which may successfully direct hypoxic tumor boosts in patients, thereby enhancing tumor control.

Preliminary investigation of 48V-labeled VO(acac)2 for cancer imaging: An initial proof-of-concept study.

In this preliminary study, a procedure for synthesizing novel PET radiotracer vanadium-48-labeled-vanadyl acetylacetonate was developed, including radioisotope production via cyclotron, separation of 48V, chelation as 48VO(acac)2, and assessment through in vitro cellular studies. We employed the beam-stop setup in a cyclotron as the target holder to irradiate titanium foils in the reaction of natTi (p,n)48V. The radioisotope production rate was 4.84 ± 0.67 µCi/µA-h. Overall radiochemical yield was 12.86 ± 0.51% with gamma-ray spectroscopy showing no detectable contaminant peaks. HPLC of 48VO(acac)2 showed a retention time (1:48) corresponding closely to that (1:50) of commercial VO(acac)2, verifying the successful synthesis of 48VO(acac)2. In vitro cellular studies demonstrated radiotracer uptake and saturation around 0.48 nM. These studies pave the way for improving methodologies and in vivo experiments, including imaging studies, in future investigations.

Improving Tumor Hypoxia Location in 18F-Misonidazole PET with Dynamic Contrast-enhanced MRI Using Quantitative Electron Paramagnetic Resonance Partial Oxygen Pressure Images.

Purpose: To enhance the spatial accuracy of fluorine 18 (18F) misonidazole (MISO) PET imaging of hypoxia by using dynamic contrast-enhanced (DCE) MR images as a basis for modifying PET images and by using electron paramagnetic resonance (EPR) partial oxygen pressure (pO2) as the reference standard. Materials and Methods: Mice (n = 10) with leg-borne MCa4 mammary carcinomas underwent EPR imaging, T2-weighted and DCE MRI, and 18F-MISO PET/CT. Images were registered to the same space for analysis. The thresholds of hypoxia for PET and EPR images were tumor-to-muscle ratios greater than or equal to 2.2 mm Hg and less than or equal to 14 mm Hg, respectively. The Dice similarity coefficient (DSC) and Hausdorff distance (d H ) were used to quantify the three-dimensional overlap of hypoxia between pO2 EPR and 18F-MISO PET images. A training subset (n = 6) was used to calculate optimal DCE MRI weighting coefficients to relate EPR to the PET signal; the group average weights were then applied to all tumors (from six training mice and four test mice). The DSC and d H were calculated before and after DCE MRI-corrected PET images were obtained to quantify the improvement in overlap with EPR pO2 images for measuring tumor hypoxia. Results: The means and standard deviations of the DSC and d H between hypoxic regions in original PET and EPR images were 0.35 mm ± 0.23 and 5.70 mm ± 1.7, respectively, for images of all 10 mice. After implementing a preliminary DCE MRI correction to PET data, the DSC increased to 0.86 mm ± 0.18 and the d H decreased to 2.29 mm ± 0.70, showing significant improvement (P < .001) for images of all 10 mice. Specifically, for images of the four independent test mice, the DSC improved with correction from 0.19 ± 0.28 to 0.80 ± 0.29 (P = .02), and the d H improved from 6.40 mm ± 2.5 to 1.95 mm ± 0.63 (P = .01). Conclusion: Using EPR information as a reference standard, DCE MRI information can be used to correct 18F-MISO PET information to more accurately reflect areas of hypoxia.Keywords: Animal Studies, Molecular Imaging, Molecular Imaging-Cancer, PET/CT, MR-Dynamic Contrast Enhanced, MR-Imaging, PET/MR, Breast, Oncology, Tumor Mircoenvironment, Electron Paramagnetic ResonanceSupplemental material is available for this article.© RSNA, 2021.

In†Vivo Imaging of the Tumor-Associated Enzyme NCEH1 with a Covalent PET Probe.

Herein, we report the development of an 18 F-labeled, activity-based small-molecule probe targeting the cancer-associated serine hydrolase NCEH1. We undertook a focused medicinal chemistry campaign to simultaneously preserve potent and specific NCEH1 labeling in live cells and animals, while permitting facile 18 F radionuclide incorporation required for PET imaging. The resulting molecule, [18 F]JW199, labels active NCEH1 in live cells at nanomolar concentrations and greater than 1000-fold selectivity relative to other serine hydrolases. [18 F]JW199 displays rapid, NCEH1-dependent accumulation in mouse tissues. Finally, we demonstrate that [18 F]JW199 labels aggressive cancer tumor cells in vivo, which uncovered localized NCEH1 activity at the leading edge of triple-negative breast cancer tumors, suggesting roles for NCEH1 in tumor aggressiveness and metastasis.

A novel approach to measuring macrophage-specific reverse cholesterol transport in vivo in humans.

Reverse cholesterol transport (RCT) is thought to be an atheroprotective function of HDL, and macrophage-specific RCT in mice is inversely associated with atherosclerosis. We developed a novel method using 3H-cholesterol nanoparticles to selectively trace macrophage-specific RCT in vivo in humans. Use of 3H-cholesterol nanoparticles was initially tested in mice to assess the distribution of tracer and response to interventions known to increase RCT. Thirty healthy subjects received 3H-cholesterol nanoparticles intravenously, followed by blood and stool sample collection. Tracer counts were assessed in plasma, nonHDL, HDL, and fecal fractions. Data were analyzed by using multicompartmental modeling. Administration of 3H-cholesterol nanoparticles preferentially labeled macrophages of the reticuloendothelial system in mice, and counts were increased in mice treated with a liver X receptor agonist or reconstituted HDL, as compared with controls. In humans, tracer disappeared from plasma rapidly after injection of nanoparticles, followed by reappearance in HDL and nonHDL fractions. Counts present as free cholesterol increased rapidly and linearly in the first 240 min after nadir; counts in cholesteryl ester increased steadily over time. Estimates of fractional transfer rates of key RCT steps were obtained. These results support the use of 3H-cholesterol nanoparticles as a feasible approach for the measurement of macrophage RCT in vivo in humans.

A Dedicated Breast Positron Emission Tomography Scanner: Proof of Concept.

PURPOSE: This study developed and tested a novel scanner constructed for dedicated positron emission tomography (PET) of the breast. The breast PET (B-PET) scanner is designed with two opposing detectors using curve plate NaI(Tl) detectors to achieve a combination of high spatial resolution and energy resolution. METHODS: Phantom and clinical studies (n = 20) with 18F-fluorodeoxyglucose were carried out on the whole-body Philips Allegro scanner and the B-PET scanner. Images were subjectively assessed by an expert panel. RESULTS: Phantom studies indicated improved contrast for B-PET over conventional PET. Of the 20 clinical studies with breast cancer demonstrated on whole-body fluorodeoxyglucose PET, 10 B-PET scans showed agreement. Of the remaining 10 studies, three had breasts that were too small to be imaged, four had lesions that were too deep to be captured in the field of view, and three were excluded due to technical errors. CONCLUSIONS: Compared with conventional PET, B-PET images provided greater detail in breast lesions suggesting that the low-cost and relatively simple design of B-PET may potentially be an important adjunct to traditional mammography in helping determine the nature of a lesion.

Biodistribution and dosimetry of (18)F-EF5 in cancer patients with preliminary comparison of (18)F-EF5 uptake versus EF5 binding in human glioblastoma.

PURPOSE: The primary purpose of this study was to assess the biodistribution and radiation dose resulting from administration of (18)F-EF5, a lipophilic 2-nitroimidazole hypoxia marker in ten cancer patients. For three of these patients (with glioblastoma) unlabeled EF5 was additionally administered to allow the comparative assessment of (18)F-EF5 tumor uptake with EF5 binding, the latter measured in tumor biopsies by fluorescent anti-EF5 monoclonal antibodies. METHODS: (18)F-EF5 was synthesized by electrophilic addition of (18)F(2) gas, made by deuteron bombardment of a neon/fluorine mixture in a high-pressure gas target, to an allyl precursor in trifluoroacetic acid at 0° then purified and administered by intravenous bolus. Three whole-body images were collected for each of ten patients using an Allegro (Philips) scanner. Gamma counts were determined in blood, drawn during each image, and urine, pooled as a single sample. PET images were analyzed to determine radiotracer uptake in several tissues and the resulting radiation dose calculated using OLINDA software and standard phantom. For three patients, 21 mg/kg unlabeled EF5 was administered after the PET scans, and tissue samples obtained the next day at surgery to determine EF5 binding using immunohistochemistry techniques (IHC). RESULTS: EF5 distributes evenly throughout soft tissue within minutes of injection. Its concentration in blood over the typical time frame of the study (∼3.5 h) was nearly constant, consistent with a previously determined EF5 plasma half-life of ∼13 h. Elimination was primarily via urine and bile. Radiation exposure from labeled EF5 is similar to other (18)F-labeled imaging agents (e.g., FDG and FMISO). In a de novo glioblastoma multiforme patient, focal uptake of (18)F-EF5 was confirmed by IHC. CONCLUSION: These results confirm predictions of biodistribution and safety based on EF5's characteristics (high biological stability, high lipophilicity). EF5 is a novel hypoxia marker with unique pharmacological characteristics allowing both noninvasive and invasive measurements.

The radiation response of cells from 9L gliosarcoma tumours is correlated with [F18]-EF5 uptake.

PURPOSE: Tumour hypoxia affects cancer biology and therapy-resistance in both animals and humans. The purpose of this study was to determine whether EF5 ([2-(2-nitro-1-H-imidazol-1-yl)-N-(2,2,3,3,3-pentafluoropropyl)-acetamide]) binding and/or radioactive drug uptake correlated with single-dose radiation response in 9L gliosarcoma tumours. MATERIALS AND METHODS: Twenty-two 9L tumours were grown in male Fischer rats. Rats were administered low specific activity (18)F-EF5 and their tumours irradiated and assessed for cell survival and hypoxia. Hypoxia assays included EF5 binding measured by antibodies against bound-drug adducts and gamma counts of (18)F-EF5 tumour uptake compared with uptake by normal muscle and blood. These assays were compared with cellular radiation response (in vivo to in vitro assay). In six cases, uptake of tumour versus muscle was also assayed using images from a PET (positron emission tomography) camera (PENN G-PET). RESULTS: The intertumoural variation in radiation response of 9L tumour-cells was significantly correlated with uptake of (18)F-labelled EF5 (i.e., including both bound and non-bound drug) using either tumour to muscle or tumour to blood gamma count ratios. In the tumours where imaging was performed, there was a significant correlation between the image analysis and gamma count analysis. Intertumoural variation in cellular radiation response of the same 22 tumours was also correlated with mean flow cytometry signal due to EF5 binding. CONCLUSION: To our knowledge, this is the first animal model/drug combination demonstrating a correlation of radioresponse for tumour-cells from individual tumours with drug metabolism using either immunohistochemical or non-invasive techniques.

Comparison of diffuse optical tomography of human breast with whole-body and breast-only positron emission tomography.

We acquire and compare three-dimensional tomographic breast images of three females with suspicious masses using diffuse optical tomography (DOT) and positron emission tomography (PET). Co-registration of DOT and PET images was facilitated by a mutual information maximization algorithm. We also compared DOT and whole-body PET images of 14 patients with breast abnormalities. Positive correlations were found between total hemoglobin concentration and tissue scattering measured by DOT, and fluorodeoxyglucose (18F-FDG) uptake. In light of these observations, we suggest potential benefits of combining both PET and DOT for characterization of breast lesions.

Accuracy of [18F]fluorodopa positron emission tomography for diagnosing and localizing focal congenital hyperinsulinism.

OBJECTIVES: Focal lesions in infants with congenital hyperinsulinism (HI) represent areas of adenomatosis that express a paternally derived ATP-sensitive potassium channel mutation due to embryonic loss of heterozygosity for the maternal 11p region. This study evaluated the accuracy of 18F-fluoro-l-dihydroxyphenylalanine ([18F]DOPA) positron emission tomography (PET) scans in diagnosing focal vs. diffuse disease and identifying the location of focal lesions. DESIGN: A total of 50 infants with HI unresponsive to medical therapy were studied. Patients were injected iv with [18F]DOPA, and PET scans were obtained for 50-60 min. Images were coregistered with abdominal computed tomography scans. PET scan interpretations were compared with histological diagnoses. RESULTS: The diagnosis of focal or diffuse HI was correct in 44 of the 50 cases (88%). [18F]DOPA PET identified focal areas of high uptake of radiopharmaceutical in 18 of 24 patients with focal disease. The locations of these lesions matched the areas of increased [18F]DOPA uptake on the PET scans in all of the cases. PET scan correctly located five lesions that could not be visualized at surgery. The positive predictive value of [18F]DOPA in diagnosing focal adenomatosis was 100%, and the negative predictive value was 81%. CONCLUSIONS: [18F]DOPA PET scans correctly diagnosed 75% of focal cases and were 100% accurate in identifying the location of the lesion. These results suggest that [18F]DOPA PET imaging provides a useful guide to surgical resection of focal adenomatosis and should be considered as a guide to surgery in all infants with congenital HI who have medically uncontrollable disease.

Diagnosis and localization of focal congenital hyperinsulinism by 18F-fluorodopa PET scan.

OBJECTIVES: To assess the accuracy of 18F-fluoro-L-dihydroxyphenylalanine ([18F]-DOPA) PET scans to diagnose focal versus diffuse disease and to localize focal lesions in infants with congenital hyperinsulinism. STUDY DESIGN: Twenty-four infants with hyperinsulinism unresponsive to medical therapy were studied. Patients were injected intravenously with [18F]-DOPA, and PET scans were obtained for 1 hour. Images were coregistered with abdominal CT scans. RESULTS: The diagnosis of focal or diffuse hyperinsulinism was correct in 23 of the 24 cases (96%) and equivocal in 1 case. [18F]-DOPA PET identified focal areas of high uptake of radiopharmaceutical in 11 patients. Pathology results confirmed that all 11 had focal adenomatosis, and the locations of these lesions matched the areas of increased [18F]-DOPA uptake on the PET scans in all of the cases. CONCLUSIONS: [18F]-DOPA PET scans were 96% accurate in diagnosing focal or diffuse disease and 100% accurate in localizing the focal lesion. These results suggest that [18F]-DOPA PET imaging should be considered in all infants with congenital hyperinsulinism who need to have pancreatectomy.