Clostridioides difficile infection (CDI) is a major healthcare challenge that is associated with high morbidity and mortality. Current diagnostic methods are limited in terms of specificity and invasiveness, necessitating novel, non-invasive imaging techniques. In this study, we develop and evaluate an immunoPET radiotracer targeting C. difficile toxin B for in vivo detection of CDI in a murine model.
The monoclonal antibody bezlotoxumab, was radiolabeled with [125I]I for in vitro characterization and with [89Zr]Zr for in vivo PET imaging, resulting in high radiochemical yields (75.36 ± 4.11% for [125I]I and 71.58 ± 8.19% for [89Zr]Zr) and purities (> 99.99% in both cases), with stable binding properties. PET/CT imaging 48 h post-infection in an animal model of CDI (C57BL/6 mice and ribotype 027 strain) demonstrated specific accumulation of [89Zr]Zr-DFO-Beztxab in the colon and cecum of infected mice, thus making it possible to distinguish CDI from dysbiosis without specific C. difficile infection and healthy controls. Findings were confirmed by PET-based quantification and ex vivo biodistribution.
We successfully developed an immunoPET radiotracer targeting toxin B for detection of CDI. Its application in an animal CDI model proved its capacity to detect the source of infection with high specificity, while avoiding confusion with non-specific inflammation.
Despite recent therapeutic advances, managing liver cancer remains a significant medical and economic priority for many countries. Patients often present at an advanced stage, preventing them from benefiting from salvage surgery. Consequently, palliative treatments play a crucial role in managing these cancers. Interventional radiology techniques are well-known for providing significant benefits to patients. [90Y]Y-L4-Lipo/Lipiodol® Ultra-Fluid is a novel transarterial radioembolization formulation designed for selective arterial injection to deliver localized radiation treatment of advanced unresectable liver tumors. This study aimed to confirm the stability of [90Y]Y-L4-Lipo/Lipiodol® Ultra-Fluid, investigate its biodistribution in an animal model, and conduct an initial dosimetry evaluation.
Less than 3% of 90Y was released from the [90Y]Y-L4-Lipo/Lipiodol® Ultra-Fluid formulation in human serum. The tumor-to-liver activity ratio (T/NT) expressed in %ID/g tissue at 1 h, 24 h, 3 days, and 6 days (5.50 ± 2.42, 3.28 ± 1.94, 4.89 ± 3.41 and 4.77 ± 1.59, respectively) suggests effective targeting of tumor tissue compared to healthy liver tissue. The extrapolated absorbed doses in Gy in humans to the tumor, normal liver, lung, and red marrow per GBq of [90Y]Y-L4-Lipo/Lipiodol® Ultra-Fluid administered were 54.3, 16.2, 0.69 and 0.89, for males, and 54.3, 22.3, 0.84 and 0.89, respectively, for females.
With good in vitro stability at low activity lasting at least 3 half-lives and a T/NT ratio of 4 in vivo, [90Y]Y-L4-Lipo/Lipiodol® Ultra-Fluid is confirmed as a valid candidate treatment for transarterial radioembolization of hepatocellular carcinoma. High activity stability in both in vitro and in vivo studies is needed to complete the formulation’s safety profile.
Epithelial cell adhesion molecule (EpCAM) is a transmembrane glycoprotein, which is overexpressed in several types of malignancies. Designed ankyrin repeat protein (DARPin) Ec1 is a 19 kDa engineered scaffold protein that binds with high affinity to EpCAM. Radiolabelled Ec1 might be used as a companion diagnostic for the selection of patients for personalized therapy. This study aimed to investigate the influence of different radiometal-chelator complexes on the biodistribution and imaging contrast of 68Ga-labelled Ec1. To investigate this, two macrocyclic chelators, 1,4,7-triazacyclononane-N,N,N-triacetic acid (NOTA) and 1-(1,3-carboxypropyl)-1,4,7-triazacyclononane-4,7-diacetic acid (NODAGA) were conjugated to the C-terminus of the Ec1. The previously developed DARPin Ec1 conjugated to 1,4,7,10-tetraazacylododecane-1,4,7,10-tetraacetic acid (DOTA) was used as a comparator.
All Ec1 variants were successfully labelled with 68Ga. The use of NOTA and NODAGA provided twice higher radiochemical yield and improved label stability compared to DOTA. All labelled Ec1 variants bound to the EpCAM-expressing cells with nanomolar affinity and preserved targeting specificity in vitro and in vivo. Biodistribution studies in mice bearing EpCAM-expressing SKOV-3 xenografts showed that [68Ga]Ga-Ec1-NOTA had lower uptake in most normal organs while maintaining tumor uptake. Among all variants, [68Ga]Ga-Ec1-NOTA showed the lowest liver uptake, with no significant differences in tumor uptake. Additionally, [68Ga]Ga-Ec1-NOTA provided the highest tumor-to-blood ratio compared to [68Ga]Ga-Ec1-DOTA and [68Ga]Ga-Ec1-NODAGA.
[68Ga]Ga-Ec1-NOTA is the preferred radioconjugate for PET imaging of EpCAM expression.
Radiolabeling is a technique that involves attaching radioactive isotopes to molecules, allowing for their tracking and analysis in biological systems. Radiolabeled d-glucose and its derivatives have a very prominent role in exploring metabolic pathways, the enzymatic system, and measuring the flow of the metabolites through biochemical reactions, as accumulation or deficiency of metabolites occurs along with metabolic disorders. Glucose as the main source of energy in the body is involved in different metabolic pathways like glycolysis, pentose phosphate pathway, and tricarboxylic acid cycle. Various derivatives of glucose are labeled at different positions by 14C and 3H. The aim of this review is to summarize some of the most significant aspects of the use of different radiolabeled d-glucose, 2-deoxy-d-glucose, and methyl-α-d-glucopyranoside.
This review focuses on the application of radiolabeled glucose derivatives in studying glucose transport systems, metabolic pathways, enzyme activity, and glucose utilization across various tissues. It highlights their role in understanding disease mechanisms in diabetes, cancer, heart failure, and metabolic disorders, and the impact of pharmacological agents and environmental pollutants.
In conclusion, radiolabeled glucose derivatives are invaluable tools for studying glucose metabolism across various tissues and organs. They provide critical insights into metabolic dysfunctions, disease mechanisms, and therapeutic interventions, aiding in the development of targeted treatments for conditions like diabetes, cancer, and cardiovascular diseases.
Simultaneous targeting of RGD and FA receptors on breast carcinoma could improve the diagnostic outcome of breast cancer patients. In this study, we have designed and synthesized an FA-RGD heteromeric targeting vector, with both RGD and FA motifs, in one single molecule for positron emission tomography (PET) diagnostic imaging of breast carcinoma.
Aoa-FA-RGD peptide conjugate was radiolabeled efficiently with [18F]FDG, resulting in high labeling efficiency (≥ 85%). The in vitro stability of the radiotracer in human plasma was found to be high. The Aoa-FA-RGD peptide conjugate showed the nanomolar affinity (≤ 51 nM) to the TNBC MDA-MB-231 cell line. In the MDA-MB-231 xenografts model, [18F]FDG-Aoa-FA-RGD peptide conjugate exhibited efficient clearance from the blood and excretion predominantly by the renal pathway (~ 56% ID), possibly due to its hydrophilic nature. A rapid accumulation of 3.30% ID/g in the TNBC MDA-MB-231 tumors was observed at 45 min p.i. Whereas a low accumulation of radioactivity was seen in the normal organs, including the heart, lungs, liver, stomach, spleen, intestines, and kidneys (< 4% ID/g). The receptor specificity of the radiotracer was confirmed by the receptor-blocking assay. A rapid and efficient tumor targeting, together with the favorable pharmacokinetics, highlights the tumor-targeting potential of the radiofluroconjugate. Furthermore, PET imaging provided sufficient visualization of MDA-MB-231 tumors in mice.
Our findings suggest that the [18F]FDG-labeled FA-RGD peptide conjugate can be a useful agent for the efficient targeting of TNBC cells. This study suggests the potential of this innovative heteromeric targeting agent for rapid and efficient targeting of tumors and merits further advancement.
N,1,4-Tri(4-ethoxy-2-hydroxybenzyl)-1,4-diazepan-6-amine (TEoS-DAZA), a novel radiopharmaceutical precursor for a liver-specific 68Ga-based diagnostic radiopharmaceutical, was tested for toxicity in rats to ensure its safe applicability and to fulfil the preclinical requirements in preparation of a clinical study. The study was performed according to EMA draft Guideline on the non-clinical requirements for radiopharmaceuticals, as well as to the so-called microdosing approach of the ICH guideline M3 (R2).
This randomized study was conducted using Wistar rats. The test item was administered intravenously at three different dose levels, the vehicle solution was administered to a separate group as control. Toxicity assessment included a 24 h observation period in three dose groups, and a 14-day recovery period in the high dose group. Animals were monitored regarding clinical behaviour, bodyweight, food and water consumption, additionally undergoing modified IRWIN, grip-strength and beam-walking tests. Following euthanisation, extensive haematological and clinical biochemical parameters were analysed. Necropsy and histopathology were performed. There was no evidence to any test-item related adversities at any dose level. No delayed effects were identified in any animal at the end of the recovery phase. Some small, albeit significant changes in haematology and clinical biochemistry could not be related to the test item administration. The NOAEL of TEoS-DAZA was determined at 1.4 mg/kg bodyweight.
Administration of a thousandfold clinical dose of TEoS-DAZA in rats did not cause any observable adverse events. An injectable solution of [68Ga]Ga-TEoS-DAZA containing 100 µg of the precursor is safe for clinical application to humans from the pharmacological point of view. Subsequent dosimetry studies need to be undertaken to reveal any radiation related toxicity.
The increasing use of [⁶⁸Ga]Ga-based radiopharmaceuticals in PET imaging, requires efficient quality control procedures. The standard r-TLC method for verifying [⁶⁸Ga]Ga-EDOTREOTIDE (Somakit-TOC®) radiochemical purity (RCP) is time-consuming, creating workflow challenges in radiopharmacies. This study evaluates an optimized r-TLC method with a reduced migration distance (4 cm vs. 9 cm) to improve efficiency while maintaining analytical reliability. Tests for specificity, accuracy and robustness were performed using ITLC-SG – Acetate and ITLC-SG – Citrate systems. Additionally, migration time was analyzed to evaluate whether the alternative method could offer added benefits.
The mean Rs for ITLC-SG – Acetate at 4 cm was 2.43 ± 0.28, while for ITLC-SG – Citrate with added [⁶⁸Ga]GaCl₃ was 5.58 ± 0.23, both exceeding the threshold of 1.5. The mean RCP at 4 cm was 98.90% ± 0.25%, and 99.21% ± 0.19% at 9 cm, with [⁶⁸Ga]Ga-uncomplexed remaining within acceptable limits. No [⁶⁸Ga]GaCl₃ was detected. The coefficient of variation (CV) for RCP between methods was < 2% (0.22%). Operator-based analysis yielded a mean Rs of 3.95 ± 0.06 (CV = 1.52%) and a mean [⁶⁸Ga]Ga-EDOTREOTIDE percentage of 99.60% ± 0.03% (CV = 0.03%). Migration times were significantly reduced with the alternative method (85% reduction).
Shortening the migration distance in r-TLC did not compromise specificity, accuracy or robustness while significantly reducing analysis time. The proposed method enhances PET radiopharmaceutical workflows, allowing faster patient dose preparation without quality loss. This approach could be investigated to other [68Ga]Ga-labeled compounds, supporting improved clinical and research applications in nuclear medicine.
Colony-stimulating factor 1 receptor (CSF1R) is a promising imaging biomarker for neuroinflammation and tumor-associated macrophages. However, existing positron emission tomography (PET) tracers for CSF1R imaging often suffer from limited specificity or sensitivity.
We have performed 11C-labeled radiosynthesis of compound FJRD (3-((2-amino-5-(1-methyl-1H-pyrazol-4-yl)pyridin-3-yl)ethynyl)-N-(4-methoxyphenyl)-4-methylbenzamide), which exhibits excellent affinity for CSF1R, and evaluated its in vivo and in vitro binding properties. PET images of [11C]FJRD show low brain uptake and specific binding in the living organs, except the kidneys in both normal mice and rats. In vitro autoradiographs demonstrate high levels of specific binding in all investigated organs, including the brain, spleen, liver, kidneys and lungs, when self-blocking was used. The addition of CPPC partially blocked in vitro [11C]FJRD binding in these organs, with blocking effects ranging from 9 to 67%. In contrast, the other two CSF1R inhibitors, GW2580 and BLZ945, showed minimal blocking effects, suggesting unignorable off-target binding in these organs. Furthermore, specific binding of [11C]CPPC and [11C]GW2580 was faint in the mouse organs, with [11C]CPPC demonstrating detectable binding only in the spleen.
These results suggest that [11C]FJRD is a potential CSF1R-PET tracer for more sensitive detection of CSF1R, compared to [11C]CPPC and [11C]GW2580. However, the high level off-target binding necessitates further improvements in specificity for CSF1R imaging.
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