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Advances in chelator design are the cornerstone for the development of metals like copper and gallium based biomedical agents and radiopharmaceuticals. To develop optimal chelating ligands, we explored the synthesis and chelating properties of azaheterocycle pendant armed 1,4,7-triazacyclononane (TACN) dimethylcarboxylate derivatives and dimethylphosphonate derivatives. In the complexation kinetics test, dicarboxylate pendant armed TACN derivatives 2,2'-(7-((1H-imidazol-2-yl)methyl)-1,4,7-triazonane-1,4-diyl)diacetic acid (NODA-Im), 2,2'-(7-((1-methyl-1H-imidazol-2-yl)methyl)-1,4,7-triazonane-1,4-diyl)diacetic acid (NODA-MeIm), and 2,2'-(7-(thiazol-2-ylmethyl)-1,4,7-triazonane-1,4-diyl)diacetic acid (NODA-Thia) exhibited fast complexation kinetics towards Cu (II) cations, which were comparable to the frequently explored ligand 1,4,7-triazacyclononane-1,4,7-triacetic acid (NOTA). And the diphosphonate pendant armed TACN derivative ((7-(thiazol-2-ylmethyl)-1,4,7-triazonane-1,4-diyl)bis(methylene))bis(phosphonic acid) (NODP-Thia) bound with Ga (III) cations at a much faster rate than NOTA. Density functional theory studies confirmed that the better complexation kinetics and metal chelating efficiency of NODA-Im, NODA-MeIm, NODA-Thia, and NODP-Thia could be ascribed to the lower Gibbs energies of corresponding chelator-metal complexes than NOTA-metal complexes. The kinetic inertness of the Cu (II) complex with NODA-Im, NODA-MeIm, and NODA-Thia was also demonstrated by cyclic voltammetry studies. Subsequently radiolabeling experiment demonstrated that these metal chelators could efficiently labeled with

Targeting metastatic esophageal squamous cell carcinoma (ESCC) has been a challenge in clinical practice. Emerging evidence demonstrates that C-X-C chemokine receptor 4 (CXCR4) highly expresses in ESCC and plays a pivotal role in the process of tumor metastasis. We developed a copper-64 (

The aim of this work is to estimate the biological effect of targeted radionuclide therapy using Cu-64, which is a well-known Auger electron emitter. To do so, we evaluate the absorbed dose of emitted particles from Cu-64 using the Geant4-DNA Monte Carlo simulation toolkit. The contribution of beta particles to the absorbed dose is higher than that of Auger electrons. The simulation result agrees with experimental ones evaluated using coumarin-3-carboxylic acid chemical dosimeter. The simulation result is also in good agreement with previous ones obtained using fluorescent nuclear track detector. From the results of present simulation (i.e., absorbed dose estimation) and previous biological experiments using two cell lines (i.e., evaluation of survival curves), we have estimated the relative biological effectiveness (RBE) of Cu-64 emitted particles on CHO wild-type cells and xrs5 cells. The RBE of xrs5 cells exposed to Cu-64 is almost equivalent to that with gamma rays and protons and C ions. This result indicates that the radiosensitivity of xrs5 cells is independent of LET. In comparison to this, the RBE on CHO wild-type cells exposed to Cu-64 is significantly higher than gamma rays and almost equivalent to that irradiated with C ions with a linear energy transfer of 70 keV/μm.

Herein, we report a new series of bifunctional chelators (BFCs) with a high affinity for amyloid aggregates, a strong binding affinity toward Cu(II), and favorable lipophilicity for potential blood-brain barrier penetration. The alkyl carboxylate ester pendant arms offer up to 3 orders of magnitude higher binding affinity toward Cu(II) and enable the BFCs to form stable

Purpose Many radiotracers are currently available for the detection of recurrent prostate cancer (rPC), yet many have not been compared head-to-head in comparative imaging studies. There is therefore an unmet need for evidence synthesis to guide evidence-based decisions in the selection of radiotracers. The objective of this study was therefore to assess the detection rate of various radiotracers for the rPC. Methods The PUBMED, EMBASE, and the EU and NIH trials databases were searched without date or language restriction for comparative imaging tracers for 13 radiotracers of principal interest. Key search terms included 18F-PSMA-1007, 18F-DCPFyl, 68Ga-PSMA-11, 18F-PSMA-11, 68Ga-PSMA-I&T, 68Ga-THP-PSMA, 64Cu-PSMA-617, 18F-JK-PSMA-7, 18F-Fluciclovine, 18F-FABC, 18F-Choline, 11C-Choline, and 68Ga-RM2. Studies reporting comparative imaging data in humans in rPC were selected. Single armed studies and matched pair analyses were excluded. Twelve studies with eight radiotracers were eligible for inclusion. Two independent reviewers screened all studies (using the PRISMA-NMA statement) for inclusion criteria, extracted data, and assessed risk of bias (using the QUADAS-2 tool). A network meta-analysis was performed using Markov-Chain Monte Carlo Bayesian analysis to obtain estimated detection rate odds ratios for each tracer combination. Results A majority of studies were judged to be at risk of publication bias. With the exception of 18F-PSMA-1007, little difference in terms of detection rate was revealed between the three most commonly used PSMA-radiotracers (68Ga-PSMA-11, 18F-PSMA-1007, 18F-DCFPyl), which in turn showed clear superiority to choline and fluciclovine using the derived network. Conclusion Differences in patient-level detection rates were observed between PSMA- and choline-radiotracers. However, there is currently insufficient evidence to favour one of the four routinely used PSMA-radioligands (PSMA-11, PSMA-1007, PSMA-I&T, and DCFPyl) over another owing to the limited evidence base and risk of publication bias revealed by our systematic review. A further limitation was lack of reporting on diagnostic accuracy, which might favour radiotracers with low specificity in an analysis restricted only to detection rate. The NMA derived can be used to inform the design of future clinical trials and highlight areas where current evidence is weak.

PURPOSE: Despite unprecedented responses to immune checkpoint inhibitors and targeted therapy in melanoma, a major subset of patients progresses and have few effective salvage options. We have previously demonstrated robust, selective uptake of the peptidomimetic LLP2A labeled with Cu-64 ([(64)Cu]-LLP2A) for positron emission tomography (PET) imaging in subcutaneous and metastatic models of B16F10 murine melanoma. LLP2A binds with high affinity to very late antigen-4 (VLA-4, integrin α(4)β(1)), a transmembrane protein overexpressed in melanoma and other cancers that facilitates tumor growth and metastasis. Yet B16F10 fails to faithfully reflect human melanoma biology, as it lacks certain oncogenic driver mutations, including BRAF mutations found in ≥50% of clinical specimens. Here, we evaluated the PET tracer [(64)Cu]-CB-TE1A1P-PEG(4)-LLP2A ([(64)Cu]-LLP2A) in novel, translational BRAF(V600E) mutant melanoma models differing in VLA-4 expression - BPR (VLA-4(−)) and BPRα (VLA-4(+). PROCEDURES: BPR cells were transduced with α(4) (CD49d) to overexpress intact cell surface VLA-4 (BPRα). The binding affinity of [(64)Cu]-LLP2A to BPR and BPRα cells was determined by saturation binding assays. [(64)Cu]-LLP2A internalization into B16F10, BPR, and BPRα cells was quantified via a plate-based assay. Tracer biodistribution and PET/CT imaging were evaluated in mice bearing subcutaneous BPR and BPRα tumors. RESULTS: [(64)Cu]-LLP2A demonstrated high binding affinity to BPRα (K(d) = 1.4 nM) but indeterminate binding to BPR cells. VLA-4(+) BPRα and B16F10 displayed comparable time-dependent [(64)Cu]-LLP2A internalization, whereas BPR internalization was undetectable. PET/CT showed increased tracer uptake in BPRα tumors vs. BPR tumors in vivo, which was validated by significantly greater (p

Compared with photon-induced binary cancer therapy, such as photothermal therapy (PTT) and photodynamic therapy (PDT), boron neutron capture therapy (BNCT) emerges as an alternative noninvasive treatment strategy that could overcome the shallow penetration of light. One key factor in performing successful BNCT is to accumulate a sufficient amount of B-10 (>20 ppm) within tumor cells, which has been a long-standing challenge for small-molecule-based boron drugs. Boron nitride nanoparticles (BNNPs) are promising boron carriers due to their high boron content and good biocompatibility, as certain types of BNNPs can undergo rapid degradation under physiological conditions. To design an on-demand degradable boron carrier, BNNPs were coated by a phase-transitioned lysozyme (PTL) that protects BNNPs from hydrolysis during blood circulation and can be readily removed by vitamin C after neutron capture therapy. According to PET imaging, the coated BNNPs exhibited high tumor boron accumulation while maintaining a good tumor to nontumor ratio. Tail-vein injections of vitamin C were followed by neutron irradiation, and BNNPs were found to be rapidly cleared from major organs according to ex vivo ICP-OES analysis. Compared with the control group, animals treated with BNCT showed suppression of tumor growth, while almost negligible side effect was observed. This strategy not only utilized the high boron content of BNNPs but also successfully performed an on-demand degradation of BNNPs to avoid the potential toxicity caused by the long-term accumulation of nanoparticles.

Background:68Ga-PSMA is a widely useful PET/CT tracer for prostate cancer imaging. Being a transmembrane protein acting as a glutamate carboxypeptidase enzyme, PSMA is highly expressed in prostate cancer cells. PSMA can also be labeled with 64Cu, offering a longer half-life and different resolution imaging. Several studies documented bio-distribution and pitfalls of 68Ga-PSMA as well as of 64Cu- PSMA. No data are reported on differences between these two variants of PSMA. Our aim was to evaluate physiological distribution of these two tracers and to analyze false positive cases.Methods:We examined tracer bio-distribution in prostate cancer patients with negative 68Ga-PSMA PET/CT (n=20) and negative 64Ga-PSMA PET/CT (n=10). A diagnostic pitfall for each tracer was documented.Results:Bio-distribution of both tracers was similar, with some differences due to renal excretion of 68Ga- PSMA and biliary excretion of 64Cu-PSMA. 68Ga-PSMA uptake was observed in sarcoidosis while 64Cu- PSMA uptake was recorded in pneumonitis.Discussion:Both tracers may present similar bio-distribution in the human body, with similar uptake in exocrine glands and high intestinal uptake. Similarly to other tracers, false positive cases cannot be excluded in clinical practice.Conclusion:The knowledge of difference in bio-distribution between two tracers may help in interpretation of PET data. Diagnostic pitfalls can be documented, due to the possibility of PSMA uptake in inflammation. Our results are preliminary to future studies comparing diagnostic accuracies of 68Ga-PSMA and 64Cu-PSMA.

Backgrounds Overexpression of epidermal growth factor receptor (EGFR) has been established as a valid therapeutic target of non-small cell lung cancer (NSCLC). However, the clinical benefit of cetuximab as an EGFR-targeting drug is still controversial, partially due to the lack of effective means to identify suitable patients. This study aimed to investigate the potential of radiolabeled cetuximab as a non-invasive tool to predict cetuximab accumulation in NSCLC tumor xenografts with varying EGFR expression levels. Methods The NSCLC tumors in model mice were subjected to in vivo biodistribution study and positron emission tomography (PET) imaging 48 h after injection of either 111In- or 64Cu-labeled cetuximab. The EGFR expression levels of NSCLC tumors were determined by ex vivo immunoblotting. Results We found that tumors with high EGFR expression had significantly higher [111In]In-DOTA-cetuximab accumulation than tumors with moderate to low EGFR expression (P 111In]In-DOTA-cetuximab tumor uptake and EGFR expression level (r = 0.893), and between [64Cu]Cu-DOTA-cetuximab tumor uptake with EGFR expression level (r = 0.915). PET imaging with [64Cu]Cu-DOTA-cetuximab allowed clear visualization of tumors. Conclusion Our findings suggest that this immuno-PET imaging can be clinically translated as a tool to predict cetuximab accumulation in NSCLC cancer patients prior to cetuximab therapy.

Exosomes have attracted tremendous attention due to their important role in physiology, pathology, and oncology, as well as promising potential in biomedical applications. Although great efforts have been dedicated to investigating their biological properties and applications as natural cancer drug-delivery systems, the systemic biodistribution of exosomes remains underexplored. In addition, exosome-based drug delivery is inevitably hindered by the robust liver clearance, leading to suboptimal tumor retention and therapeutic efficiency. In this study, we report one of the first examples using in vivo positron emission tomography (PET) for noninvasive monitoring of copper-64 (64Cu)-radiolabeled polyethylene glycol (PEG)-modified exosomes, achieving excellent imaging quality and quantitative measurement of blood residence and tumor retention. PEGylation not only endowed exosomes with a superior pharmacokinetic profile and great accumulation in the tumor versus traditionally reported native exosomes but also reduced premature hepatic sequestration and clearance of exosomes, findings that promise enhanced therapeutic delivery efficacy and safety in future studies. More importantly, this study provides important guidelines about surface engineering, radiochemistry, and molecular imaging in obtaining accurate and quantitative biodistribution information on exosomes, which may benefit future exploration in the realm of exosomes.

Molecules containing lysine-ureido-glutamate functional groups bind to the active site of prostate specific membrane antigen, which is overexpressed in prostate cancer. To prepare copper radiopharmaceuticals for the diagnosis and therapy of prostate cancer, macrobicyclic sarcophagine ligands tethered to either one or two lysine-ureido-glutamate functional groups through an appropriate linker have been prepared. Sarcophagine ligands can be readily radiolabeled with positron-emitting copper-64 at room temperature. The bivalent agent, in which two targeting groups are tethered to a single copper complex, dramatically outperforms the monomeric agent with respect to tumor uptake and retention. The high tumor uptake, low background, and prolonged tumor retention, even at 24 hours post injection, suggest the bivalent agent is a promising diagnostic for prostate cancer and could be used for prospective dosimetry for therapy with a copper-67 variant.

Scintigraphic imaging of malignant glioblastoma (MG) continues to be challenging. We hypothesized that VPAC1 cell surface receptors can be targeted for positron emission tomography (PET) imaging of orthotopically implanted MG in a mouse model, using a VPAC1-specific peptide [64Cu]TP3805. The expression of VPAC1 in mouse GL261 and human U87 glioma cell lines was determined by western blot. The ability of [64Cu]TP3805 to bind to GL261 and U87 cells was studied by cell-binding. Receptor-blocking studies were performed to validate receptor specificity. GL261 tumors were implanted orthotopically in syngeneic T-bet knockout C57BL/6 mouse brain (N = 15) and allowed to grow for 2–3 weeks. Mice were injected i.v., first with ~ 150 μCi of 2-deoxy-2-[18F]fluoro-D-glucose ([18F]FDG) then 24 h later with ~ 200 μCi of [64Cu]TP3805. In another set of tumor-bearing mice, (N = 5), ionic [64Cu]Cl2 was injected as a control. Mice were imaged at a 2-h post-injection using an Inveon micro-PET/CT, sacrificed and % ID/g of [64Cu]TP3805 and [64Cu]Cl2 were calculated in a tumor, normal brain, and other tissues. For histologic tissue examination, 3-μm thick sections of the tumors and normal brain were prepared, digital autoradiography (DAR) was performed, and then the sections were H&E stained for histologic examination. Western blots showed a strong signal for VPAC1 on both cell lines. [64Cu]TP3805 cell-binding was 87 ± 1.5 %. Receptor-blocking reduced cell-binding to 24.3 ± 1.5 % (P

Nanoparticles have been assessed in preclinical models of atherosclerosis for detection of plaque complexity and treatment. However, their successful clinical translation has been hampered by less than satisfactory plaque detection, and lack of a general strategy for assessing the translational potential of nanoparticles. Herein, nanoparticles based on comb-copolymer assemblies were synthesized through a modular construction approach with precise control over the conjugation of multiple functional building blocks for in vivo evaluation. This high level of design control also allows physicochemical properties to be varied in a controllable fashion. Through conjugation of c-atrial natriuretic factor (CANF) peptide and radiolabeling with (64)Cu, the (64)Cu-CANF-Comb nanoparticle was assessed for plaque imaging by targeting natriuretic peptide clearance receptor (NPRC) in a double-injury atherosclerosis model in rabbits. The extended blood circulation and improved binding capacity of (64)Cu-CANF-Comb nanoparticles afforded sensitive and specific detection of NPRC upregulated in atherosclerotic lesions by positron emission tomography (PET) at intervals during the progression of disease. Ex vivo tissue validation using autoradiography and immunostaining on human carotid endarterectomy specimens demonstrated specific binding of (64)Cu-CANF-Comb to human NPRC receptors. Taken together, this study not only shows the potential of NPRC-targeted (64)Cu-CANF-Comb nanoparticles for increased sensitivity to an epitope that increases during atherosclerosis plaque development, but also provides a useful strategy for the general design and assessment of translational potential of nanoparticles in cardiovascular imaging.

Objective: To investigate Porphysome fluorescence image-guided resection (PYRO-FGR) for detection of uterine tumour, metastatic lymph nodes and abdominal metastases in a model of endometrial cancer. Methods: White New Zealand rabbits were inoculated with VX2 cells via intra-myometrial injection. At 30 days, Porphysomes were administered intravenously. At 24 h the abdomen was imaged and fluorescent tissue identified (PYRO-FGR). After complete resection of fluorescent tissue, fluorescence-negative lymph nodes and peritoneal biopsies were removed. Histopathology including ultra-staging and analysis by a pathologist was used to detect tumour. Fluorescence signal to background ratio (SBR) was calculated and VX2 (+) tissue compared to VX2 (-) tissue. Biodistribution was calculated and Porphysome accumulation in fluorescent VX2 (+) tissue compared to fluorescent VX2 (-) and non-fluorescent VX2 (-) tissue. Results: Of 17 VX2 models, 10 received 4 mg/kg of Porphysomes and 7 received 1 mg/kg. Seventeen tumours (UT), 81 lymph nodes (LN) and 54 abdominal metastases (AM) were fluorescence-positive and resected. Of these, 17 UT, 60 LN and 45 AM were VX2 (+), while 16 LN and 5 AM were VX2 (-). Nine specimens were excluded from analysis. Thirty-one LN and 53 peritoneal biopsies were fluorescence-negative and resected. Of these, all LN and 51/53 biopsies were VX2 (-) with only 2 false-negative biopsies. Sensitivity and specificity of PYRO-FGR for VX2 (+) tissue was 98.4% / 80.0% overall, 100% / 100% for UT, 100% / 66.0 % for LN and 95.7% / 91.4% for AM. Increased SBR and biodistribution was observed in VX2 (+) tissue vs. VX2 (-) tissue. Conclusions: Porphysomes are a highly sensitive imaging agent for intra-operative detection and resection of uterine tumour, metastatic lymph nodes and abdominal metastases.

Purpose: To design and evaluate a small engineered protein binder targeting human programmed death-1 ligand (hPD-L1) in vivo for PET imaging in four mouse tumor models, and in situ in human cancer specimens. Experimental Design: The hPD-L1 protein binder, FN3hPD-L1, was engineered using a 12-kDa human fibronectin type-3 domain (FN3) scaffold. The binder's affinity was assayed in CT26 mouse colon carcinoma cells stably expressing hPD-L1 (CT26/hPD-L1). 64Cu-FN3hPD-L1 was assayed for purity, specific activity, and immunoreactivity. Four groups of NSG mice (n = 3–5/group) were imaged with 64Cu-FN3hPD-L1 PET imaging (1–24 hours postinjection of 3.7 MBq/7 μg of Do-FN3 in 200 μL PBS): Nod SCID Gamma (NSG) mice bearing (i) syngeneic CT26/hPD-L1tumors, (ii) CT26/hPD-L1 tumors blocked (blk) by preinjected nonradioactive FN3hPD-L1 binder, (iii) hPD-L1-negative Raji xenografts, and (iv) MDA-MB-231 xenografts. The FN3hPD-L1 binder staining was evaluated against validated hPD-L1 antibodies by immunostaining in human cancer specimens. Results: FN3hPD-L1 bound hPD-L1 with 1.4 ± 0.3 nmol/L affinity in CT26/hPD-L1 cells. 64Cu-FN3hPD-L1 radiotracer showed >70% yield and >95% purity. 64Cu-FN3hPD-L1 PET imaging of mice bearing CT26/hPD-L1 tumors showed tumor-to-muscle ratios of 5.6 ± 0.9 and 13.1 ± 2.3 at 1 and 4 hours postinjection, respectively. The FN3hPD-L1 binder detected hPD-L1 expression in human tissues with known hPD-L1 expression status based on two validated antibodies. Conclusions: The 64Cu-FN3hPD-L1 radiotracer represents a novel, small, and high-affinity binder for imaging hPD-L1 in tumors. Our data support further exploration and clinical translation of this binder for noninvasive identification of cancer patients who may respond to immune checkpoint blockade therapies.

Amyloid-β plaques, consisting of aggregated amyloid-β peptides, are one of the pathological hallmarks of Alzheimer's disease. Copper complexes formed using positron-emitting copper radionuclides that cross the blood-brain barrier and bind to specific molecular targets offer the possibility of noninvasive diagnostic imaging using positron emission tomography. New thiosemicarbazone-pyridylhydrazone based ligands that incorporate pyridyl-benzofuran functional groups designed to bind amyloid-β plaques have been synthesized. The ligands form stable complexes with copper(II) ( Kd = 10-18 M) and can be radiolabeled with copper-64 at room temperature. Subtle changes to the periphery of the ligand backbone alter the metabolic stability of the complexes in mouse and human liver microsomes, and influenced the ability of the complexes to cross the blood-brain barrier in mice. A lead complex was selected based on possessing the best metabolic stability and brain uptake in mice. Synthesis of this lead complex with isotopically enriched copper-65 allowed us to show that the complex bound to amyloid-β plaques present in post-mortem human brain tissue using laser ablation-inductively coupled plasma-mass spectrometry. This work provides insight into strategies to target metal complexes to amyloid-β plaques, and how small modifications to ligands can dramatically alter the metabolic stability of metal complexes as well as their ability to cross the blood-brain barrier.

International audience; We present here the synthesis of two new bifunctionalized azachelators, no2th-EtBzNCS and Hno2th1tha, as bioconjugable analogues of two previously described di- and trimethylthiazolyl 1,4,7-triazacyclononane (tacn) ligands, no2th and no3th, for potential uses in copper-64 (64Cu) positron emission tomography imaging. The first one bears an isothiocyanate group on the remaining free nitrogen atom of the tacn framework, while the second one presents an additional carboxylic function on one of the three heterocyclic pendants. Their syntheses required regiospecific N-functionalization of the macrocycles. In order to investigate their suitability for in vivo applications, a complete study of their copper(II) chelation was performed. The acid–base properties of the ligands and their thermodynamic stability constants with copper(II) and zinc(II) cations were determined using potentiometric techniques. Structural studies were conducted in both solution and the solid state, consolidated by theoretical calculations. The kinetic inertness in an acidic medium of both copper(II) complexes was determined by spectrophotometry, while cyclic voltammetry experiments were performed to evaluate the stability at the copper(I) redox state. UV–vis, NMR (of the zinc complexes), electron paramagnetic resonance spectroscopy, and density functional theory studies showed excellent agreement between the solution structures of the complexes and their crystallographic data. These investigations unambiguously prove that these bifunctional derivatives display similar coordination properties as their no2th and no3th counterparts, opening the door to targeted bioapplications. The no2th-EtBzNCS and Hno2th1tha ligands were then conjugated to a bombesin antagonist peptide for targeting the gastrin-releasing peptide receptor (GRPr). To highlight the potential of the two chelators for radiopharmaceutical development, the 64Cu-radiolabeling properties, in vitro stability, and binding affinity to GRPr of the corresponding bioconjugates were determined. Altogether, the results of this work warrant the further development of 64Cu-based radiopharmaceuticals comprising our novel bifunctional chelators.

Previous studies has demonstrated the utility of human epidermal growth factor receptor type 2 (HER2) as an attractive target for cancer molecular imaging and therapy. An affibody protein with strong binding affinity for HER2, ZHER2:342, has been reported. Various methods of chelator conjugation for radiolabeling HER2 affibody molecules have been described in the literature including N-terminal conjugation, C-terminal conjugation, and other methods. Cu-64 has recently been extensively evaluated due to its half-life, decay properties, and availability. Our goal was to optimize the radiolabeling method of this affibody molecule with Cu-64, and translate a positron emission tomography (PET) probe with the best in vivo performance to clinical PET imaging of HER2-positive cancers. In our study, three anti-HER2 affibody proteins-based PET probes were prepared, and their in vivo performance was evaluated in mice bearing HER2-positive subcutaneous SKOV3 tumors. The affibody analogues, Ac-Cys-ZHER2:342, Ac-ZHER2:342(Cys39), and Ac-ZHER2:342-Cys, were synthesized using the solid phase peptide synthesis method. The purified small proteins were site-specifically conjugated with the maleimide-functionalized chelator, 1,4,7,10-tetraazacyclododecane-1,4,7-tris- aceticacid-10-maleimidethylacetamide (maleimido-mono-amide-DOTA). The resulting DOTA-affibody conjugates were then radiolabeled with Cu-64. Cell uptake assay of the resulting PET probes, [64Cu]DOTA-Cys-ZHER2:342, [64Cu]DOTA-ZHER2:342(Cys39), and [64Cu]DOTA-ZHER2:342-Cys, was performed in HER2-positive human ovarian SKOV3 carcinoma cells at 4 and 37 °C. The binding affinities of the radiolabeled peptides were tested by cell saturation assay using SKOV3 cells. PET imaging, biodistribution, and metabolic stability studies were performed in mice bearing SKOV3 tumors. Cell uptake assays showed high and specific uptake by incubation of Cu-64-labeled affibodies with SKOV3 cells. The affinities (KD) of the PET radio probes as tested by cell saturation analysis were in the low nanomolar range with the ranking of [64Cu]DOTA-Cys-ZHER2:342 (25.2 ± 9.2 nM) ≈ [64Cu]DOTA-ZHER2:342-Cys (32.6 ± 14.7 nM) > [64Cu]DOTA-ZHER2:342(Cys39) (77.6 ± 22.2 nM). In vitro stability and in vivo metabolite analysis study revealed that all three probes were stable enough for in vivo imaging applications, while [64Cu]DOTA-Cys-ZHER2:342 showed the highest stability. In vivo small-animal PET further demonstrated fast tumor targeting, good tumor accumulation, and good tumor to normal tissue contrast of all three probes. For [64Cu]DOTA-Cys-ZHER2:342, [64Cu]DOTA-ZHER2:342(Cys39), and [64Cu]DOTA-ZHER2:342-Cys, tumor uptake at 24 h are 4.0 ± 1.0 % ID/g, 4.0 ± 0.8 %ID/g, and 4.3 ± 0.7 %ID/g, respectively (mean ± SD, n = 4). Co-injection of the probes with non-labeled anti-HER2 affibody proteins confirmed in vivo specificities of the compounds by tumor uptake reduction. The three Cu-64-labeled ZHER2:342 analogues all display excellent HER2 targeting ability and tumor PET imaging quality. Although varied in the position of the radiometal labeling of these three Cu-64-labeled ZHER2:342 analogues, there is no significant difference in tumor and normal tissue uptakes among the three probes. [64Cu]DOTA-Cys-ZHER2:342 stands out as the most superior PET probe because of its highest affinities and in vivo stability.

Copper-64-Diacetyl-bis(N4-methylthiosemicarbazone) [64Cu][Cu(ATSM)] is a hypoxia-targeting PET tracer with applications in oncology and cardiology. Upon entering a hypoxic cell, [64Cu][Cu(II)(ATSM)] is reduced to a putative [64Cu][Cu(I)(ATSM)]− species which dissociates to deposit radiocopper, thereby providing hypoxic contrast. This process may be dependent upon protonation arising from intracellular acidosis. Since acidosis is a hallmark of ischemic tissue and tumors, the hypoxia specificity of [64Cu][Cu(ATSM)] may be confounded by changes in intracellular pH. We have therefore determined the influence of intracellular pH on [64Cu][Cu(ATSM)] pharmacokinetics. Using isolated perfused rat hearts, acidosis was induced using an ammonium pre-pulse method, with and without hypoxic buffer perfusion. Cardiac [64Cu][Cu(ATSM)] pharmacokinetics were determined using NaI detectors, with intracellular pH and cardiac energetics monitored in parallel by 31P NMR. To distinguish direct acidotic effects on tracer pharmacokinetics from acidosis-induced hypocontractility, parallel studies used lidocaine perfusion to abolish cardiac contraction. Hypoxic myocardium trapped [64Cu][Cu(ATSM)] despite no evidence of it being acidotic when characterised by 31P NMR. Independent induction of tissue acidosis had no direct effect on [64Cu][Cu(ATSM)] pharmacokinetics in either normoxic or hypoxic hearts, beyond decreasing cardiac oxygen consumption to alleviate hypoxia and decrease tracer retention, leading us to conclude that tissue acidosis does not mediate the hypoxia selectivity of [64Cu][Cu(ATSM)].

Most current nanoparticle-based PET tracers are radiolabeled through metal chelators conjugated on the nanoparticle surface. Metal chelation usually requires sophisticated optimization and may impact the physical or chemical properties of nanoparticles, which leads to the changes in their distribution and pharmacokinetics in vivo. A chelator-free radiolabeling approach is thus highly desirable. Here, we report that zinc sulfide (ZnS) quantum dots (QDs) can be rapidly radiolabeled with 68Ga or 64Cu through cation exchange without chelators. The radiolabeling was accomplished in times as short as 5 min at 37 °C in aqueous solution, yielding a high labeling efficiency and radiochemical purity for both isotopes. Surface functionalization with targeting peptides was also readily achieved to enable or enhance the cellular uptake of QDs. In vivo PET imaging showed that 64Cu-labeled QDs had a much higher tumor uptake (7.3% ID g-1) than 64Cu-DOTA in a murine cancer model. Overall, this study presents a QD-based platform to achieve convenient and chelator-free radiolabeling, and improve PET imaging of solid tumors.

A drug delivery system (DDS) for combined therapy, based on a short oxidized multiwalled carbon nanotube, is reported. It was prepared by exploiting a synthetic approach which allowed loading of two drugs, doxorubicin and metformin, the targeting agent biotin and a radiolabeling tag, to enable labeling with Ga-68 or Cu-64 in order to perform an extensive biodistribution study by PET/CT. The DDS biodistribution profile changes with different administration methods. Once administered at therapeutic doses, the DDS showed a marginal beneficial effect on 4T1 tumor bearing mice, a syngeneic and orthotopic model of triple negative breast cancer, with survival extended by 1 week and 2 days in 20% of the mice. This is encouraging given the aggressiveness of the 4T1 tumor. Furthermore, our DDS was well tolerated, ruling out concerns regarding the toxicity of carbon nanotubes.

Albumin is one of the most attractive nanoplatforms for targeted imaging and drug delivery due to its biocompatibility and long circulation half-life. However, previously reported albumin-based nanoplatforms have shown inconsistent blood circulation half-life according to the modified methods, and the affecting factors were not well evaluated, which could hamper the clinical translation of albumin-based nanoplatforms. Herein, we developed a finely tuned click-chemistry based albumin nanoplatform (CAN) with a longer circulation half-life and an efficient tumor targeting ability. Methods: CAN was synthesized in two steps. First, albumin was conjugated with ADIBO-NHS (albumin-ADIBO) by reacting albumin with various molar ratios of ADIBO. The number of attached ADIBO moieties was determined using matrix-assisted laser desorption ionization time of flight (MALDI-TOF). Second, the desired modalities including azide-functionalized chelator, a fluorescence dye, and folate were incorporated into albumin-ADIBO using strain-promoted alkyne-azide cycloaddition reaction (SPAAC reaction). The biodistribution and targeting efficiency of functionalized CANs were demonstrated in mice. Results: The degree of functionalization (DOF) and resulting in vivo biodistribution was controlled precisely using the click chemistry approach. Specifically, the numbers of attached azadibenzocyclooctyne (ADIBO) moieties on albumin, the DOF, were optimized by reacting albumin with varying molar ratios of ADIBO with a high reproducibility. Furthermore, we developed a simple and efficient method to estimate the DOF using UV-visible spectrophotometry (UV-vis), which was further validated by matrix-assisted laser desorption ionization time of flight (MALDI-TOF). The biodistribution of CAN could be controlled by DOF, and CAN with an optimized DOF showed a long circulation half-life (> 18 h). CAN was further functionalized using a simple click chemistry reaction with an azide functionalized chelator, a fluorescence dye, and folate. 64Cu- and folate-labeled CAN (64Cu-CAN-FA) showed effective and specific folate receptor targeting in vivo, with an over two-fold higher uptake than the liver at 24 h post-injection. Conclusions: Our development from the precisely controlled DOF demonstrates that an optimized CAN can be used as a multifunctional nanoplatform to obtain a longer half-life with radioisotopes and ligands, and provides an effective method for the development of albumin-based tumor theranostic agents.

The homeostasis of copper (Cu) in the central nervous system is regulated by the blood-brain barrier and blood-cerebrospinal (CSF) barrier (BCB) in the choroid plexus. While proteins responsible for Cu uptake, release, storage and intracellular trafficking exist in the choroid plexus, the influence of age on Cu clearance from the CSF via the choroid plexus and how Cu transporting proteins contribute to the process are unelucidated. This study was designed to test the hypothesis that the aging process diminishes Cu clearance from the CSF of rats by disrupting Cu transporting proteins in the choroid plexus. Data from ventriculo-cisternal perfusion experiments demonstrated greater 64Cu radioactivity in the CSF effluents of older rats (18 months) compared to younger (1 month) and adult (2 months) rats, suggesting much slower removal of Cu by the choroid plexus in old animals. Studies utilizing qPCR and immunofluorescence revealed an age-specific expression pattern of Cu transporting proteins in the choroid plexus. Moreover, proteomic analyses unraveled age-specific proteomes in the choroid plexus with distinct pathway differences, particularly associated with extracellular matrix and neurodevelopment between young and old animals. Taken together, these findings support an age-dependent deterioration in CSF Cu clearance, which appears to be associated with altered subcellular distribution of Cu transporting proteins and proteomes in the choroid plexus.

Plastics are used commonly in the world because of their convenience and cost effectiveness. Microplastics, an environmental threat and human health risk, are widely detected in food and consequently ingested. However, degraded plastics are found everywhere, creating an environmental threat and human health risk. Therefore, real-time monitoring of orally administered microplastics to trace them in the body is tremendously important.

CD20-overexpressed non-Hodgkin lymphoma typically indicates progressive malignancy. Obinutuzumab is a next-generation Food and Drug Administration–approved humanized monoclonal antibody that targets CD20. Previous studies with (89)Zr-labeled obinutuzumab have successfully imaged CD20 in vivo. However, delayed tumor uptake and increased radioactive exposure caused by long blood circulation limit its clinical translation. This study aimed to develop (64)Cu-labeled F(ab′)(2) fragments of obinutuzumab for imaging CD20 in lymphoma xenograft tumor models. Methods: F(ab′)(2) fragments were produced from obinutuzumab using an IgG-degrading enzyme of Streptococcus pyogenes (IdeS) enzyme and purified with protein A beads. Sodium dodecyl sulfate polyacrylamide gel electrophoresis and high-performance liquid chromatography were performed to evaluate the products and their stability. F(ab′)(2) products were conjugated with p-SCN-Bn-NOTA (NOTA) for (64)Cu radiolabeling. Western blotting was performed to screen the CD20 expression levels of lymphoma cells. Enzyme-linked immunosorbent assay, flow cytometry, and confocal imaging were used to test the binding affinity in vitro. Serial PET imaging and biodistribution studies in subcutaneous lymphoma–bearing mice were performed using (64)Cu-NOTA-F(ab′)(2)-obinutuzumab or (64)Cu-NOTA-F(ab′)(2)-IgG. Results: F(ab′)(2)-obinutuzumab and F(ab′)(2)-IgG produced by the IdeS digestion system were confirmed with sodium dodecyl sulfate polyacrylamide gel electrophoresis and high-performance liquid chromatography. The radiochemical purity of (64)Cu-labeled F(ab′)(2) fragments was no less than 98%, and the specific activity was 56.3 ± 7.9 MBq/mg (n = 6). Among the 5 lymphoma cell lines, Ramos showed the strongest expression of CD20, and CLL-155 showed the lowest, as confirmed by enzyme-linked immunosorbent assay, flow cytometry, and confocal imaging. PET imaging revealed rapid and sustained tumor uptake of (64)Cu-NOTA-F(ab′)(2)-obinutuzumab in Ramos tumor–bearing mice. The peak tumor uptake (9.08 ± 1.67 percentage injected dose per gram of tissue [%ID/g]) in the Ramos model was significantly higher than that in the CCL-155 model (2.78 ± 0.62 %ID/g) or the (64)Cu-NOTA-F(ab′)(2)-IgG control (1.93 ± 0.26 %ID/g, n = 4, P < 0.001). The tumor-to-blood and tumor-to-muscle ratios were 7.3 ± 1.6 and 21.9 ± 9.0, respectively, at 48 h after injection in the (64)Cu-NOTA-F(ab′)(2)-obinutuzumab group. Of the measured off-target organs, the kidneys showed the highest uptake. Ex vivo immunofluorescent staining verified the differential CD20 expression in the Ramos and CCL-155 tumor models. Conclusion: This study demonstrated that (64)Cu-NOTA-F(ab′)(2)-obinutuzumab had a rapid and sustained tumor uptake in CD20-positive lymphoma with high contrast, which could enable noninvasive evaluation of CD20 levels in the clinic.

Background: A research priority in finding a cure for HIV is to establish methods to accurately locate and quantify where and how HIV persists in people living with HIV (PLWH) receiving suppressive antiretroviral therapy (ART). Infusing copper-64 (64Cu) radiolabelled broadly neutralising antibodies targeting HIV envelope (Env) with CT scan and positron emission tomography (PET) identified HIV Env in tissues in SIV infected non-human primates . We aimed to determine if a similar approach was effective in people living with HIV (PLWH). Methods: Unmodified 3BNC117 was compared with 3BNC117 bound to the chelator MeCOSar and 64Cu (64Cu-3BNC117) in vitro to assess binding and neutralization. In a clinical trial 64Cu-3BNC117 was infused into HIV uninfected (Group 1), HIV infected and viremic (viral load, VL >1000 c/mL; Group 2) and HIV infected aviremic (VL

The validation of metal-phenolic nanoparticles (MPNs) in preclinical imaging studies represents a growing field of interest due to their versatility in forming predesigned structures with unique properties. Before MPNs can be used in medicine, their pharmacokinetics must be optimized so that accumulation in nontargeted organs is prevented and toxicity is minimized. Here, we report the fabrication of MPNs made of a coordination polymer core that combines In(III), Cu(II), and a mixture of the imidazole 1,4-bis(imidazole-1-ylmethyl)-benzene and the catechol 3,4-dihydroxycinnamic acid ligands. Furthermore, a phenolic-based coating was used as an anchoring platform to attach poly(ethylene glycol) (PEG). The resulting MPNs, with effective hydrodynamic diameters of around 120 nm, could be further derivatized with surface-embedded molecules, such as folic acid, to facilitate in vivo targeting and multifunctionality. The prepared MPNs were evaluated for in vitro plasma stability, cytotoxicity, and cell internalization and found to be biocompatible under physiological conditions. First, biomedical evaluations were then performed by intrinsically incorporating trace amounts of the radioactive metals 111In or 64Cu during the MPN synthesis directly into their polymeric matrix. The resulting particles, which had identical physicochemical properties to their nonradioactive counterparts, were used to perform in vivo single-photon emission computed tomography (SPECT) and positron emission tomography (PET) in tumor-bearing mice. The ability to incorporate multiple metals and radiometals into MPNs illustrates the diverse range of functional nanoparticles that can be prepared with this approach and broadens the scope of these nanoconstructs as multimodal preclinical imaging agents., This work was supported by grant RTI2018-098027-B-C21 from the Spanish Government funds and by the European Regional Development Fund (ERDF). The ICN2 is funded by the CERCA programme/Generalitat de Catalunya. The ICN2 is supported by the Severo Ochoa Centres of Excellence programme, funded by the Spanish Research Agency (AEI, grant no. SEV-2017-0706). Financial support from the University of Copenhagen, the Phospholipid Research Center (Heidelberg, Germany), and the Lundbeck Foundation (Copenhagen, Denmark) is kindly acknowledged. TRIUMF receives funding via a contribution agreement with the National Research Council of Canada. T.V.F.E. received financial support from a Killam Doctoral Fellowship and a Four-Year Doctoral Fellowship (4YF) from the University of British Columbia (UBC). K.S. acknowledges the generous support of BWXT Isotope Technology Group for the supply of the radioisotope 111In. The authors also thank the technicians and veterinarians at UBC Centre for Comparative Medicine (CCM, Vancouver, Canada), especially Dr. Laura Mowbray, Jana Hodasova, and Ava McHugh for their expert assistance in the imaging lab, and the Canada Foundation for Innovation (project no. 25413) for its support of the imaging facility (http://invivoimaging.ca/). S.S.-G. acknowledges the support from MINECO BES-2015-071492 grant. S.S.-G. and C.R.-R. acknowledge Dr. Pedro L. Esquinas for fruitful discussions about the quantitative measurements of 111In and 64Cu. Special thanks to Dr. Javier Saiz-Poseu from 3D Science Visuals (Instagram: 3dscivisuals) for contributing with graphic design and scientific illustrations. The authors acknowledge the support from the Cost ENBA CA15216.

Background A research priority in finding a cure for HIV is to establish methods to accurately locate and quantify where and how HIV persists in people living with HIV (PLWH) receiving suppressive antiretroviral therapy (ART). Infusing copper-64 (64Cu) radiolabelled broadly neutralising antibodies targeting HIV envelope (Env) with CT scan and positron emission tomography (PET) identified HIV Env in tissues in SIV infected non-human primates . We aimed to determine if a similar approach was effective in people living with HIV (PLWH). Methods Unmodified 3BNC117 was compared with 3BNC117 bound to the chelator MeCOSar and 64Cu (64Cu-3BNC117) in vitro to assess binding and neutralization. In a clinical trial 64Cu-3BNC117 was infused into HIV uninfected (Group 1), HIV infected and viremic (viral load, VL >1000 c/mL; Group 2) and HIV infected aviremic (VL

Herein we present the preparation of two novel cyclam-based macrocycles (

In recent years the use of beta-emitting radiopharmaceuticals for cancer therapy has expanded rapidly following development of therapeutics for neuroendocrine tumors, prostate cancer, and other oncologic malignancies. One emerging beta-emitting radioisotope of interest for therapy is

PURPOSE: Glioblastoma (GBM) is the most common malignant brain tumor in adults. Various immunotherapeutic approaches to improve patient survival are being developed, but the molecular mechanisms of immunotherapy resistance are currently unknown. Here, we explored the ability of a humanized radiolabeled CD8-targeted minibody to non-invasively quantify tumor-infiltrating CD8(+) T cells using positron emission tomography (PET). EXPERIMENTAL DESIGN: We generated a peripheral blood mononuclear cell (PBMC) humanized immune system (HIS) mouse model and quantified the absolute number of CD8(+) T cells by flow cytometry relative to the [(64)Cu]Cu-NOTA-anti-CD8 PET signal. To evaluate a patient derived orthotopic GBM HIS model, we intracranially injected cells into NOG mice, humanized cohorts with multiple HLA-matched PBMC donors and quantified CD8(+) tumor-infiltrating lymphocytes by immunohistochemistry (IHC). To determine whether [(64)Cu]Cu-NOTA-anti-CD8 images brain parenchymal T cell infiltrate in GBM tumors, we performed PET and autoradiography and subsequently stained serial sections of brain tumor tissue by IHC for CD8(+) T cells. RESULTS: Non-tumor-bearing NOG mice injected with human PBMCs showed prominent [(64)Cu]Cu-NOTA-anti-CD8 uptake in the spleen and minimal radiotracer localization to the normal brain. NOG mice harboring intracranial human GBMs yielded high-resolution PET images of tumor-infiltrating CD8(+) T cells. Radiotracer retention correlated with CD8(+) T cell numbers in spleen and tumor tissue. Our study demonstrates the ability of [(64)Cu]Cu-NOTA-anti-CD8 PET to quantify peripheral and tumor-infiltrating CD8(+) T cells in brain tumors. CONCLUSION: Human CD8(+) T cells infiltrate an orthotopic GBM in a donor-dependent manner. Further, [(64)Cu]Cu-NOTA-anti-CD8 quantitatively images both peripheral and brain parenchymal human CD8(+) T cells.

We report herein the preclinical evaluation of new [

Invasive pulmonary aspergillosis (IPA) is a life-threatening lung disease of immunocompromised humans, caused by the opportunistic fungal pathogen Aspergillus fumigatus. Inadequacies in current diagnostic procedures mean that early diagnosis of the disease, critical to patient survival, remains a major clinical challenge, and is leading to the empiric use of antifungal drugs and emergence of azole resistance. A non-invasive procedure that allows both unambiguous detection of IPA and its response to azole treatment is therefore needed. Here, we show that a humanised Aspergillus-specific monoclonal antibody, dual labelled with a radionuclide and fluorophore, can be used in immunoPET/MRI in vivo in a neutropenic mouse model and 3D light sheet fluorescence microscopy ex vivo in the infected mouse lungs to quantify early A. fumigatus lung infections and to monitor the efficacy of azole therapy. Our antibody-guided approach reveals that early drug intervention is critical to prevent complete invasion of the lungs by the fungus, and demonstrates the power of molecular imaging as a non-invasive procedure for tracking IPA in vivo., Invasive pulmonary aspergillosis is a life-threatening fungal lung disease devoid of specific rapid diagnosis and with limited therapeutic options. Here, the authors show how state-of-the-art imaging approaches can enable specific diagnosis and therapy monitoring of this infection.

Imaging of CD8 receptors on T-cells by positron emission tomography (PET) has been considered a promising strategy for monitoring the treatment response to immunotherapy. In this study, a trial of imaging CD8 with our newly developed sequential multiple-agent receptor targeting (SMART) technology was conducted. Mice bearing a subcutaneous colorectal CT26 tumor received three times different immunotherapy treatments (PD1 or CTLA4 or combined). On either day 7 or day 14 after the first time treatment, the PET imaging study was performed with sequentially administered TCO-modified anti-CD8 antibody and 64Cu-labeled MeTz-NOTA-RGD. However, no positive response was detected, probably due to (1) inappropriate selection of biomarkers for the SMART strategy, (2) limited TCO modification on the anti-CD8 antibody, and (3) inadequate response of the CT26 tumor to the selected immunotherapies. Therefore, the potential of applying SMART in imaging CD8 was not demonstrated in this study, and further optimization will be necessary before it can be applied in imaging CD8.

Copper, a cofactor for many enzymes, is a bioelement that is involved in many main biochemical processes; although high levels of copper promote the proliferation of cancer cells. Further development of radiopharmaceuticals based on copper radioisotopes depend on understanding and taking advantage of its biochemical pathways in oncogenesis. As with other radiometals used in molecular imaging and/or targeted therapy, biological vectors are employed to transport copper radioisotopes to a target, aiming for high specific uptake at tumor sites and precise delivery of ionizing radiation. Evidence of the clinical utility of copper radioisotopes in the ionic form CuCl2 were also proven in an in vivo study of the copper metabolism, guiding personalized copper-chelating treatment in cancer patients and in imaging pathological sites associated with copper imbalance. Five of the copper radioisotopes have gained interest for nuclear medicine applications, based on their emissions, energies, and half-lives, as they can be produced with pharmaceutical-grade quality. The uptake mechanism, kinetics, and metabolic parameters are important findings in molecular imaging, which are decisive when designing individualized targeted radiotherapy for dose calculations of high linear energy transfer Auger electrons and β− emissions of 64Cu and 67Cu. As radiation deposits a high amount of energy within the intra-cellular space, the biochemical involvement of copper determines targets in drug design and validation. The biochemical pathways depict copper metabolism in normal cells and highlight its increased activity in tumor progression and angiogenesis. The avid uptake of copper into inter- and intra-mitochondrial spaces, as constituents of cytochrome C oxidase, substantiate the selection of 64/67CuCl2 as theranostic agents.

Purpose The co-stimulatory molecules CD80 and CD86 are upregulated on activated antigen-presenting cells (APC). We investigated whether local APC activation, induced by subcutaneous (s.c.) inoculation of lipopolysaccharides (LPS), can be imaged by positron emission tomography (PET) with CD80/CD86-targeting 64Cu-labelled abatacept. Procedures Mice were inoculated s.c. with extracellular-matrix gel containing either LPS or vehicle (PBS). Immune cell populations were analysed by flow cytometry and marker expression by RT-qPCR. 64Cu-NODAGA-abatacept distribution was analysed using PET/CT and ex vivo biodistribution. Results The number of CD80+ and CD86+ immune cells at the LPS inoculation site significantly increased a few days after inoculation. CD68 and CD86 expression were higher at the LPS than the PBS inoculation site, and CD80 was only increased at the LPS inoculation site. CTLA-4 was highest 10 days after LPS inoculation, when CD80/CD86 decreased again. A few days after inoculation, 64Cu-NODAGA-abatacept distribution to the inoculation site was significantly higher for LPS than PBS (4.2-fold). Co-administration of unlabelled abatacept or human immunoglobulin reduced tracer uptake. The latter reduced the number of CD86+ immune cells at the LPS inoculation site. Conclusions CD80 and CD86 are upregulated in an LPS-induced local inflammation, indicating invasion of activated APCs. 64Cu-NODAGA-abatacept PET allowed following APC activation over time., Molecular Imaging and Biology, 23, ISSN:1860-2002, ISSN:1536-1632

Gastrin-releasing peptide receptor (GRPR) is overexpressed in the majority of prostate cancers. This study aimed to investigate the potential of 64Cu (radionuclide for late time-point PET-imaging) for imaging of GRPR expression using NOTA-PEG2-RM26 and NODAGA-PEG2-RM26. Methods: NOTA/NODAGA-PEG2-RM26 were labeled with 64Cu and evaluated in GRPR-expressing PC-3 cells. Biodistribution of [64Cu]Cu-NOTA/NODAGA-PEG2-RM26 was studied in PC-3 xenografted mice and compared to the biodistribution of [57Co]Co-NOTA/NODAGA-PEG2-RM26 at 3 and 24 h p.i. Preclinical PET/CT imaging was performed in tumor-bearing mice. NOTA/NODAGA-PEG2-RM26 were stably labeled with 64Cu with quantitative yields. In vitro, binding of [64Cu]Cu-NOTA/NODAGA-PEG2-RM26 was rapid and GRPR-specific with slow internalization. In vivo, [64Cu]Cu-NOTA/NODAGA-PEG2-RM26 bound specifically to GRPR-expressing tumors with fast clearance from blood and normal organs and displayed generally comparable biodistribution profiles to [57Co]Co-NOTA/NODAGA-PEG2-RM26, tumor uptake exceeded normal tissue uptake 3 h p.i.. Tumor-to-organ ratios did not increase significantly with time. [64Cu]Cu-NOTA-PEG2-RM26 had a significantly higher liver and pancreas uptake compared to other agents. 57Co-labeled radioconjugates showed overall higher tumor-to-non-tumor ratios, compared to the 64Cu-labeled counterparts. [64Cu]Cu-NOTA/NODAGA-PEG2-RM26 was able to visualize GRPR-expression in a murine PC model using PET. However, [55/57Co]Co-NOTA/NODAGA-PEG2-RM26 provided better in vivo stability and overall higher tumor-to-non-tumor ratios compared with the 64Cu-labeled conjugates.

Boron neutron capture therapy (BNCT) is an atomic targeted radiotherapy that shows fantastic suppression impact on locally intrusive threatening tumors. One key factor for effective BNCT is to aggregate an adequate concentration (>20 ppm) of 10B in the cytoplasm of the tumor. Carborane-loaded polymer nanoparticles are promising because of their outstanding biocompatibility and plasma steadiness. In this study, a new class of carborane-loaded nanoscale covalent organic polymers (BCOPs) was prepared by a Schiff base condensation reaction, and their solubility was greatly improved in common solvents via alkyl chain engineering and size tailoring. The obtained BCOP-5T was further functionalized by biocompatible 1,2-distearoyl-sn-glycero-3-phosphoethanolamine-N-[amino(polyethylene-glycol)-2000] (DSPE-PEG, molecular weight 2000) to form stable aqueous-phase nanoparticles with a hydrodynamic diameter of around 100 nm. After chelating with radioactive copper-64, DSPE-BCOP-5T was tracked by positron emission tomography (PET) imaging and showed significant accumulation in the tumor. DSPE-BCOP-5T + neutron radiation showed remarkable tumor suppression in 4T1 tumor-bearing mice (murine breast cancer). No obvious physical tissue damage and abnormal behavior were observed, demonstrating that the boron delivery was successful and tumor-selective. To conclude, this study presents a theranostic COP-based platform with a well-defined composition, good biocompatibility, and satisfactory tumor accumulation, which is promising for PET imaging, drug delivery, and BNCT.

Nephron number varies widely in humans. A low nephron endowment at birth or a loss of functioning nephrons is strongly linked to increased susceptibility to chronic kidney disease. In this work, we developed a contrast agent, radiolabeled cationic ferritin (RadioCF), to map functioning glomeruli in vivo in the kidney using positron emission tomography (PET). PET radiotracers can be detected in trace doses (/+), a model of congenital hypoplasia and low nephron mass. The average standardized uptake value (SUV) measured by PET 90 min after injection was 21% higher in WT mice than in Os/+ mice, consistent with the higher glomerular density in WT mice. The difference in peak SUV from SUV at 90 min correlated with glomerular density in male mice from both WT and Os/+ cohorts ( R2 = 0.98). Finally, we used RadioCF-PET to map functioning glomeruli in a donated human kidney. SUV within the kidney correlated with glomerular number ( R2= 0.78) measured by CF-enhanced magnetic resonance imaging in the same locations. This work suggests that RadioCF-PET appears to accurately detect nephron mass and has the potential for clinical translation.

As an indicative biomarker for immunotherapy, PD-L1 plays an important role in the clinical decision-making of the immune checkpoint blockade therapy. PET imaging through radiotracer can real-timely, quantitatively, and non-invasively assess the expression of PD-L1 in tumors. Here, we reported a copper-64 radiolabeled NOTA-WL12, 64Cu-NOTA-WL12, and preliminarily evaluated its application in non-invasively detecting the PD-L1 expression.64Cu-NOTA-WL12 was produced with high radiochemical yield (>90%), radiochemical purity (>98%), and specific activity (20 MBq/nmol). 64Cu-NOTA-WL12 showed high in vitro stability and high binding affinity to the PD-L1 (KD ≈ 3.012 nM). The micro-positron emission tomography/computerized tomography (micro-PET/CT) imaging indicated that 64Cu-NOTA-WL12 was specifically accumulated in the tumor with PD-L1 expression. All results demonstrated that 64Cu-NOTA-WL12 holds great potential for noninvasive evaluation of PD-L1 expression levels.

INTRODUCTION Nuclear medicine plays a crucial role for personalized therapy, mainly in oncology. Chemotherapy and radiotherapy present some disadvantages and research is shifting toward nanotechnology with significant improvements in therapy and diagnosis of several cancers. Indeed, nanoparticles can be tagged with different radioisotopes for single photon emission computed tomography (SPECT) and positron emission tomography (PET) imaging and for therapy. This review describes the current state of the art of 64Copper-labeled nanoparticles for PET imaging of cancer. EVIDENCE ACQUISITION We performed a systematic analysis of literature using the terms "64CuCl 2 ," "64Cu," "Copper" AND "nanoparticle" AND "PET" in online databases: i.e. PubMed/MEDLINE and Scopus. The search was limited to English papers and original articles. We excluded articles not in English language, abstracts, case reports, review articles and meeting presentations. EVIDENCE SYNTHESIS Amongst the 116 articles retrieved, 88 were excluded because reviews, or not in English, or only in-vitro studies or meeting presentations. We considered only 28 original papers. The most used nanoparticles are liposomes and they are mainly used in breast cancer although other animal models of cancer have been also investigated. CONCLUSIONS The results showed that nanoparticles can be considered a promising radiopharmaceutical for PET imaging of different type of cancer.

Along with other novel metallic radionuclides, copper-64 (64Cu) is currently being investigated as an alternative option to the gallium-68 (68Ga) and lutetium-177 (177Lu) radiopharmaceuticals widely used for targeting somatostatin receptors, expressed by neuroendocrine tumors (NETs), and recently prostate specific membrane antigen (PSMA), expressed by prostate cancer cells. This interest is mostly driven by the peculiar nuclear properties of 64Cu that make it an almost ideal example of theranostic radionuclide. In fact, 64Cu emits both low-energy positrons, β- particles and a swarm of Auger electrons. This combination of different emissions may allow to collect high-resolution PET images, but also to use the same radiopharmaceutical for eliciting a therapeutic effect. Another unique behavior of 64Cu originates from the fundamental biological role played in organisms by the ionic forms of the copper element, which is naturally involved in a multitude of cellular processes including cell replication. These intrinsic biological characteristics has led to the discovery that 64Cu, under its simplest dicationic form Cu2+, is able to specifically target a variety of cancerous cells and to detect the onset of a metastatic process in its initial stage. This short review reports an outline of the status of 64Cu radiopharmaceuticals and of the most relevant results that are constantly disclosed by preclinical and investigational clinical studies.