Preparation and labelling of red blood cells with [68Ga]Ga-oxine for PET/CT imaging of the human spleen.

INTRODUCTION: With the introduction of automated synthetization methods, the in-house production of several 68Ga-based tracers became feasible in hospital laboratories. We describe a possible standard operating procedure (SOP) for [68Ga]Ga-oxine-labeled heat-denaturated erythrocytes, which can be used for selective imaging in patients with splenic disorders. METHODS: Heat-denaturated erythrocytes were labeled with [68Ga]Ga-oxine, which was produced from 68Ga and 8-hydroxyquinoline on an automated synthesizer. The workflow was validated in a good manufacturing/good radiopharmaceutical practice (GMP/GRP) certified laboratory. A patient underwent [68Ga]Ga-oxine-erythrocyte PET/CT for differentiation of an intrapancreatic mass. RESULTS: [68Ga]Ga-oxine and [68Ga]Ga-oxine-labeled erythrocytes could be synthesized reproducibly and reliably. The products met GMP quality standards. The tracer showed high accumulation in the intrapancreatic mass, consistent with an accessory spleen. CONCLUSIONS: PET/CT imaging with [68Ga]Ga-oxine-labeled, heat-denaturated erythrocytes can be a backup method for the differentiation of functioning splenic tissue from tumors. An SOP for the production of the tracer in a clinical setting could be established.

Liver PET Reloaded: Automated Synthesis of [68Ga]Ga-BP-IDA for Positron Imaging of the Hepatobiliary Function and First Clinical Experience.

Hepatobiliary scintigraphy is a well-established nuclear imaging method for evaluating liver function and displaying the biliary system, but the spatial and temporal resolution is limited, and, there is still no established PET equivalent. Adapted from the work of Schuhmacher et al. in 1983, the production of a 68Gallium-labeled substitute, tetrabromophthalein ([68Ga]Ga-BP-IDA), was undertaken according to current Good Manufacturing Practice (GMP) standards and proved feasible and reproducible. PET/CT with the radiotracer was performed in two complex patients with hepatocellular carcinoma in preparation for transarterial radioembolization. Due to its high spatial and temporal resolution, localization of areas with impaired liver function and visualization of the biliary system were possible. We could demonstrate that this 68Gallium-labeled, IDA-based PET tracer is feasible and could advance hepatic and biliary function PET imaging.

PET/CT of the Spleen with Gallium-Oxine-Labeled, Heat-Damaged Red Blood Cells: Clinical Experience and Technical Aspects.

Several scintigraphic techniques have been supplemented or replaced by PET/CT methods because of their superior sensitivity, high resolution, and absolute activity quantification capability. The purpose of this project was the development of a PET tracer for splenic imaging, its radiopharmaceutical validation, and its application in selected patients in whom unclear constellations of findings could not be resolved with established imaging methods. Heat-damaged red blood cells (RBCs) were labeled with [68Ga]gallium-oxine, which was produced from [68Ga]gallium and 8-Hydroxyquinoline (oxine) on an automated synthesizer. Ten patients underwent [68Ga]gallium-oxine-RBC-PET/CT for the classification of eleven unclear lesions (3 intra-, 8 extrapancreatic). [68Ga]gallium-oxine and [68Ga]gallium-oxine-labeled RBCs could be synthesized reproducibly and reliably. The products met GMP quality standards. The tracer showed high accumulation in splenic tissue. Of the 11 lesions evaluated by PET/CT, 3 were correctly classified as non-splenic, 6 as splenic, 1 as equivocal, and 1 lesion as a splenic hypoplasia. All lesions classified as non-splenic were malignant, and all lesions classified as splenic did not show malignant features during follow-up. PET/CT imaging of the spleen with [68Ga]gallium-oxine-labeled, heat-damaged RBCs is feasible and allowed differentiation of splenic from non-splenic tissues, and the diagnosis of splenic anomalies.

A Novel Breast Cancer Xenograft Model Using the Ostrich Chorioallantoic Membrane-A Proof of Concept.

The avian chorioallantoic membrane (CAM) assay has attracted scientific attention in cancer research as an alternative or complementary method for in vivo animal models. Here, we present a xenograft model based on the ostrich (struthio camelus) CAM assay for the first time. The engraftment of 2 × 106 breast cancer carcinoma MDA-MB-231 cells successfully lead to tumor formation. Tumor growth monitoring was evaluated in eight fertilized eggs after xenotransplantation. Cancer cells were injected directly onto the CAM surface, close to a well-vascularized area. Histological analysis confirmed the epithelial origin of tumors. The CAM of ostrich embryos provides a large experimental surface for the xenograft, while the comparably long developmental period allows for a long experimental window for tumor growth and treatment. These advantages could make the ostrich CAM assay an attractive alternative to the well-established chick embryo model. Additionally, the large size of ostrich embryos compared to mice and rats could help overcome the limitations of small animal models. The suggested ostrich model is promising for future applications, for example, in radiopharmaceutical research, the size of the embryonal organs may compensate for the loss in image resolution caused by physical limitations in small animal positron emission tomography (PET) imaging.

67 Ga-PSMA I&T for Radioguided Surgery of Lymph Node Metastases in Patients With Biochemical Recurrence of Prostate Cancer.

PURPOSE: Radioguided lymph node dissection in patients with prostate cancer, and suffering from biochemical recurrence has been described thoroughly during the past few years. Several prostate-specific membrane antigen (PSMA)-directed ligands labeled with 111 In, 99m Tc, and 68 Ga have been published; however, limitations regarding availability, short half-life, high costs, and unfavorable high energy might restrict frequent use. This study aims at introducing 67 Ga as a promising radionuclide for radioguided surgery. METHODS: Retrospective analysis was performed on 6 patients with 7 PSMA-positive lymph node metastases. 67 Ga-PSMA I&T (imaging and therapy) was synthesized in-house and intravenously applied according to §13 2b of the German Medicinal Products Act. Radioguided surgery was performed 24 hours after injection of 67 Ga-PSMA I&T using a gamma probe. Patient urine samples were collected. Occupational and waste dosimetry was performed to describe hazards arising from radiation. RESULTS: 67 Ga-PSMA application was tolerated without adverse effects. Five of 7 lymph nodes were detected on 22-hour SPECT/CT in 4 of 6 patients. During surgery, all 7 lymph node metastases were identified by positive gamma probe signal. Relevant accumulation of 67 Ga was observed in lymph node metastases (32.1 ± 15.1 kBq). Histology analysis of near-field lymph node dissection revealed more lymph node metastases than PET/CT (and gamma probe measurements) identified. Waste produced during inpatient stay required decay time of up to 11 days before reaching exemption limits according to German regulations. CONCLUSIONS: Radioguided surgery using 67 Ga-PSMA I&T is a safe and feasible option for patients suffering from biochemical recurrence of prostate cancer. 67 Ga-PSMA I&T was successfully synthesized according to Good Manufacturing Practice guidelines. Radioguided surgery with 67 Ga-PSMA I&T does not lead to relevant radiation burden to urology surgeons and represents a novel interdisciplinary approach in nuclear medicine and urology.

Radionuclide Cisternography with [64Cu]Cu-DOTA.

Radionuclide cisternography (RNC) is a method for conducting imaging of the cerebrospinal system and can be used to identify cerebrospinal fluid leaks. So far, RNC has commonly employed radiopharmaceutical agents suitable only for single-photon emission tomography techniques, which are thus lacking in terms of image resolution and can potentially lead to false-negative results. Therefore, [64Cu]Cu-DOTA was investigated as an alternative radiopharmaceutical for RNC, employing positron emission tomography (PET) instead of single-photon emission tomography. A formulation of [64Cu]Cu-DOTA was produced according to the guidelines for good manufacturing practice. The product met the requirements of agents suitable for intrathecal application. [64Cu]Cu-DOTA was administered to a patient and compared to the approved scintigraphic RNC agent, [111In]In-DTPA. While no cerebrospinal fluid leak was detected with [111In]In-DTPA, [64Cu]Cu-DOTA RNC exhibited a posterolateral leak between the vertebral bodies C1 and C2. Thus, in this patient, PET RNC with [64Cu]Cu-DOTA was superior to RNC with [111In]In-DTPA. Since radiopharmaceuticals have a very good safety profile regarding the occurrence of adverse events, PET RNC with [64Cu]Cu-DOTA may become an attractive alternative to scintigraphic methods, and also to computed tomography or magnetic resonance imaging, which often require contrast agents, causing adverse events to occur much more frequently.

Hepatobiliary Excretion PET/CT With 68Ga-TAoS-DAZA to Evaluate Bile Duct Patency.

ABSTRACT: Tri-alkoxysalicyl-1,4-diazepan-6-amine (TAoS-DAZA) ligands, radiolabelled with 68Ga, have been proposed as PET/CT agents for depiction and quantification of hepatobiliary function and evaluation of bile excretion. In the presented case, a patient with hepatocellulary carcinoma underwent PET/CT with the TAoS-derivate 68Ga-tri-methoxysalicyl-(TMoS)-DAZA to determine the patency of intrahepatic and extrahepatic bile ducts, in particular of a stent in the common bile duct. The PET/CT was performed without complications. Evaluation of bile excretion over time was possible. 68Ga-TAoS-DAZA PET/CT may be an option for dynamic imaging of the excretory hepatic function to visualize the biliary tree and to rule out cholestasis.

Ectopic Retrolaryngeal Parathyroid Adenoma Detected by 18F-Ethylcholine PET/US Fusion Imaging.

ABSTRACT: A 68-year-old woman was referred with the diagnosis of symptomatic pHPT (primary hyperparathyroidism). Ultrasonography (US) and 99mTc-MIBI scintigraphy could not clearly identify a parathyroid adenoma. A primary unilateral surgical exploration remained unsuccessful to remove adenomatous parathyroid tissue and the pHPT persisted. 18F-ethylcholine PET/CT showed a left-sided retropharyngeal lesion with intense tracer uptake that could not be detected on US images at first sight. Therefore, additional 18F-ethylcholine PET/US fusion imaging was performed and revealed a poorly definable nodular structure on US that could be unambiguously correlated to the PET finding. Surgical excision confirmed an oncocytic variant of parathyroid adenoma, and parathormone and calcium levels normalized immediately.

In-ovo imaging using ostrich eggs: Biomagnetism for detection of cardiac signals and embryonal motion.

In-ovo imaging using ostrich eggs has been described as a potential alternative to common animal testing. The main advantage is its independence from small animal imaging devices as ostrich eggs provide good image quality on regular CT, MRI, or PET used in examinations of humans. However, embryonal motion during dynamic imaging studies produce artifacts. The aims of this study were (1) to explore the feasibility of biomagnetism to detect cardiac signals and embryonal motion and to use these findings (2) to investigate the effect of isoflurane anesthesia on ostrich embryos. A standard magnetoencephalography developed for brain studies was used to detect embryonal signals of ostrich eggs on developmental day 34. Signals were instantly shown on a screen and data were also postprocessed. For assessing the effects of anesthesia, nine ostrich eggs were investigated using isoflurane 6% for 90 min. Biomagnetic signals were recorded simultaneously. A control group consisting of eight different ostrich eggs was also investigated. Cardiac signals similar to electrocardiography were observed in all eggs. Postprocessing revealed frequent motion of embryos without anesthesia. The exposure to isoflurane led to a significant decrease in motion signals in 9/9 ostrich embryos after 8 min. Motion was significantly reduced in the isoflurane group versus control group. There were no isoflurane-related deaths. This study shows that biomagnetism is feasible to detect cardiac signals and motion of ostrich embryos in-ovo. Application of isoflurane is safe and leads to a rapid decrease in embryonal motion, which is an important prerequisite for the implementation of in-ovo imaging using ostrich eggs.

Automated GMP Production and Preclinical Evaluation of [68Ga]Ga-TEoS-DAZA and [68Ga]Ga-TMoS-DAZA.

[68Ga]Ga-TEoS-DAZA and [68Ga]Ga-TMoS-DAZA are two novel radiotracers suitable for functional PET liver imaging. Due to their specific liver uptake and biliary excretion, the tracers may be applied for segmental liver function quantification, gall tree imaging and the differential diagnosis of liver nodules. The purpose of this study was to investigate problems that occurred initially during the development of the GMP compliant synthesis procedure and to evaluate the tracers in a preclinical model. After low radiolabeling yields were attributed to precursor instability at high temperatures, an optimized radiolabeling procedure was established. Quality controls were in accordance with Ph. Eur. requirements and gave compliant results, although the method for the determination of the 68Ga colloid is partially inhibited due to the presence of a radioactive by-product. The determination of logP revealed [68Ga]Ga-TEoS-DAZA (ethoxy bearing) to be more lipophilic than [68Ga]Ga-TMoS-DAZA (methoxy bearing). Accordingly, biodistribution studies in an in ovo model showed a higher liver uptake for [68Ga]Ga-TEoS-DAZA. In dynamic in ovo PET imaging, rapid tracer accumulation in the liver was observed. Similarly, the activity in the intestines rose steadily within the first hour p.i., indicating biliary excretion. As [68Ga]Ga-TEoS-DAZA and [68Ga]Ga-TMoS-DAZA can be prepared according to GMP guidelines, transition into the early clinical phase is now possible.

Differences in Distribution and Detection Rate of the [68Ga]Ga-PSMA Ligands PSMA-617, -I&T and -11-Inter-Individual Comparison in Patients with Biochemical Relapse of Prostate Cancer.

The biochemical relapse of prostate cancer is diagnostically challenging but of high clinical impact for subsequent patient treatment. PET/CT with radiolabeled PSMA ligands outperforms conventional diagnostic methods in the detection of tumor recurrence. Several radiopharmaceuticals were and are available for use. The aim of this study was to investigate whether the routinely applied [68Ga]Ga-PSMA ligands PSMA-617, -I&T and -11 (HBED-CC) differ in physiological and pathological distribution, or in tumor detection rate. A retrospective evaluation of 190 patients (39 patients received PSMA-617, 68 patients PSMA-I&T and 83 patients PSMA-11) showed significant differences in tracer accumulation within all organs examined. The low retention within the compartments blood pool, bone and muscle tissue is a theoretical advantage of PSMA-11. Evaluation of tumor lesion uptake and detection rate did not reveal superiority of one of the three radiopharmaceuticals, neither in the whole population, nor in particularly challenging subgroups like patients with very low PSA levels. We conclude that all three [68Ga]Ga-PSMA ligands are equally feasible in this clinically important scenario, and may replace each other in case of unavailability or production restrictions.

Dynamic PET/CT with the Hepatobiliary Tracer [68Ga]Ga-Tmos-DAZA for Characterization of a Hepatic Tumor.

Established imaging modalities for the characterization of liver tumors are computed tomography (CT), magnetical resonance (MR) imaging, sonography, and hepatobiliary scintigraphy. In some cases, their results may be inconclusive or certain examinations not possible due to contraindications. Positron emission tomography (PET)/CT has the capability of dynamic imaging with high temporal resolution. With radiolabeled tri-alkoxysalicyl-1,4-diazepan-6-amine (TAoS-DAZA) tracers, imaging of liver perfusion and hepatobiliary function is possible in a single examination. In the presented case, the PET/CT was performed in a patient with suspected hepatocellular carcinoma and atypical CT findings. PET imaging characteristics were consistent with a hepatocellular carcinoma (HCC). PET with DAZA ligands may be a supplemental method for liver tumor characterization in difficult cases.

Design and practical evaluation of a shielded application system for intravenously administered radionuclide therapies.

AIM: Intravenous radionuclide therapies are gaining importance. With the increased frequency of these therapies, safe application procedures in combination with effective radiation protection are needed. We present a shielded system which can be universally used for the application of liquid radiopharmaceuticals. MATERIALS AND METHODS: The application system consists of a cuboidal box made of lead with stainless steel coating, an inner layer of polymethyl methacrylate (PMMA), a disposable line/bottle system, and a medical infusion pump. No proprietary disposable parts are used. The system was tested and validated ex-vivo, and evaluated for Lu-177 peptide receptor radionuclide and for radioligand therapies (PRRT, RLT). RESULTS: Construction of the application system was performed in accordance with the physical characteristics of the used isotopes. 10 validation procedures with 1 GBq Tc-99m-pertechnetate confirmed functionality. 38 PRRT and 13 RLT procedures were performed successfully and completely. At least 98 % of the prescribed activities were infused. No leakage of radioactivity occurred. Dose rate measurements showed that the radiation in the box is shielded completely, and that exposure arises only from the infusion line outside the box and from the patient. CONCLUSION: The presented application system for intravenous radionuclide therapies is feasible, safe, and cost-efficient. It has been shown that Lu-177-PRRT and -RLT can be performed without complications while ensuring nearly complete infusion of the prescribed radiopharmaceutical dose. Radiation exposure of the applying physician and other staff is low. The construction of the shielding box ensures complete shielding of all radionuclides currently used for radiomolecular therapies.

Metal-Based Complexes as Pharmaceuticals for Molecular Imaging of the Liver.

This article reviews the use of metal complexes as contrast agents (CA) and radiopharmaceuticals for the anatomical and functional imaging of the liver. The main focus was on two established imaging modalities: magnetic resonance imaging (MRI) and nuclear medicine, the latter including scintigraphy and positron emission tomography (PET). The review provides an overview on approved pharmaceuticals like Gd-based CA and 99mTc-based radiometal complexes, and also on novel agents such as 68Ga-based PET tracers. Metal complexes are presented by their imaging modality, with subsections focusing on their structure and mode of action. Uptake mechanisms, metabolism, and specificity are presented, in context with advantages and limitations of the diagnostic application and taking into account the respective imaging technique.

N,1,4-Tri(4-alkoxy-2-hydroxybenzyl)-DAZA: efficient one-pot synthesis and labelling with 68Ga for PET liver imaging in ovo.

We report the isolation of a new type of 1,4-diazepan-6-amine (DAZA)-based ligand. Condensation of aldehydes with DAZA gives a novel class of 1,5-diazabicyclo[3.2.1]octanes in nearly quantitative yields. Subsequent reductive cleavage of these bicyclic aminal species with sodium borohydride selectively leads to N,1,4-tri(4-alkoxy-2-hydroxybenzyl)-1,4-diazepan-6-amines (alkoxy = Me: TMeOHB-DAZA; alkoxy = Et: TEtOHB-DAZA) via a unique reductive alkylation reaction in which a substituent is added to the DAZA moiety without the presence of an alkylating agent. Mass spectrometry studies of the intermediates suggest that the mechanism involves insertion of in situ released carbonyl species into an aminal bond to form hemiaminal intermediates, and subsequent reduction. TMeOHB-DAZA and TEtOHB-DAZA are hexadentate ligands suitable for effectively coordinating Ga(iii) ions. Chelation of the radionuclide 68Ga was achieved within 5 min at 100 °C. In vitro stability studies in PBS and human serum confirmed the kinetic inertness of the tracers as no 68Ga demetallation was observed over a period of 4 h. Positron emission tomography (PET)/computed tomography (CT) imaging after in ovo administration to incubated ostrich eggs showed a high uptake in the liver, namely 27% (60 min post injection), and subsequent biliary excretion. These results suggest that [68Ga]Ga-TMeOHB-DAZA and [68Ga]Ga-TEtOHB-DAZA have excellent potential as hepatobiliary PET/CT imaging agents.

Investigations on the Ga(III) Complex of EOB-DTPA and Its 68Ga Radiolabeled Analogue.

We demonstrate a method for the isolation of EOB-DTPA (3,6,9-triaza-3,6,9-tris(carboxymethyl)-4-(ethoxybenzyl)-undecanedioic acid) from its Gd(III) complex and protocols for the preparation of its novel non-radioactive, i.e., natural Ga(III) as well as radioactive (68)Ga complex. The ligand as well as the Ga(III) complex were characterized by nuclear magnetic resonance (NMR) spectroscopy, mass spectrometry and elemental analysis. (68)Ga was obtained by a standard elution method from a (68)Ge/(68)Ga generator. Experiments to evaluate the (68)Ga-labeling efficiency of EOB-DTPA at pH 3.8-4.0 were performed. Established analysis techniques radio TLC (thin layer chromatography) and radio HPLC (high performance liquid chromatography) were used to determine the radiochemical purity of the tracer. As a first investigation of the (68)Ga tracers' lipophilicity the n-octanol/water distribution coefficient of (68)Ga species present in a pH 7.4 solution was determined by an extraction method. In vitro stability measurements of the tracer in various media at physiological pH were performed, revealing different rates of decomposition.