ABSTRACT
BACKGROUND: Macrophages, innate immune cells that reside in all organs, defend the host against infection and injury. In the heart and vasculature, inflammatory macrophages also enhance tissue damage and propel cardiovascular diseases. METHODS: We here use in vivo positron emission tomography (PET) imaging, flow cytometry, and confocal microscopy to evaluate quantitative noninvasive assessment of cardiac, arterial, and pulmonary macrophages using the nanotracer 64Cu-Macrin-a 20-nm spherical dextran nanoparticle assembled from nontoxic polyglucose. RESULTS: PET imaging using 64Cu-Macrin faithfully reported accumulation of macrophages in the heart and lung of mice with myocardial infarction, sepsis, or pneumonia. Flow cytometry and confocal microscopy detected the near-infrared fluorescent version of the nanoparticle (VT680Macrin) primarily in tissue macrophages. In 5-day-old mice, 64Cu-Macrin PET imaging quantified physiologically more numerous cardiac macrophages. Upon intravenous administration of 64Cu-Macrin in rabbits and pigs, we detected heightened macrophage numbers in the infarcted myocardium, inflamed lung regions, and atherosclerotic plaques using a clinical PET/magnetic resonance imaging scanner. Toxicity studies in rats and human dosimetry estimates suggest that 64Cu-Macrin is safe for use in humans. CONCLUSIONS: Taken together, these results indicate 64Cu-Macrin could serve as a facile PET nanotracer to survey spatiotemporal macrophage dynamics during various physiological and pathological conditions. 64Cu-Macrin PET imaging could stage inflammatory cardiovascular disease activity, assist disease management, and serve as an imaging biomarker for emerging macrophage-targeted therapeutics.
Subject(s)
Copper Radioisotopes , Dextrans , Heart/diagnostic imaging , Lung/diagnostic imaging , Macrophages/pathology , Molecular Imaging , Positron Emission Tomography Computed Tomography , Radiopharmaceuticals , Animals , Atherosclerosis/diagnostic imaging , Atherosclerosis/pathology , Copper Radioisotopes/administration & dosage , Copper Radioisotopes/pharmacokinetics , Dextrans/administration & dosage , Dextrans/pharmacokinetics , Disease Models, Animal , Injections, Intravenous , Lung/pathology , Macrophages, Alveolar/pathology , Mice , Myocardial Infarction/diagnostic imaging , Myocardial Infarction/pathology , Nanoparticles , Pneumonia/diagnostic imaging , Pneumonia/pathology , Predictive Value of Tests , Rabbits , Radiopharmaceuticals/administration & dosage , Radiopharmaceuticals/pharmacokinetics , Swine , Swine, Miniature , Time FactorsABSTRACT
BACKGROUND: Molecular imaging is of great benefit to early disease diagnosis and timely treatment. One of the most striking innovations is the development of multimodal molecular imaging technology, which integrates two or more imaging modalities, largely in view of making the best of the advantages of each modality while overcoming their respective shortcomings. Hence, engineering a versatile and easily prepared nanomaterial with integrating multimodal molecular imaging function holds great promise, but is still a great challenge. MATERIALS AND METHODS: We firstly designed and synthesized a BDT-DPP conjugated polymer and then noncovalent self-assembly with phospholipid-polyethylene glycol endowed BDT-DPP with water solubility and biocompatibility. Followed by [Cu] labeling, the acquired multifunctional nanoparticles (NPs) were studied in detail for the photophysical property. The cytotoxicity and biocompatibility of DPP-BDT NPs were examined through MTT assay and H&E stained analysis. In addition, we investigated the accumulation of the NPs in HepG2 tumor models by positron emission tomography (PET) and photoacoustic (PA) dual-mode imaging. RESULTS AND DISCUSSION: The DPP-BDT NPs exhibited excellent optical stability, strong near-infrared (NIR) light absorption as well as fine biocompatibility. After tail vein injection into the living mice, the PA signals in the neoplastic tissues were gradually increased and reached to the maximum at the 4-h post-injection, which was consistent with the PET analysis. Such strong PA and PET signals were attributed to the efficient NPs accumulation resulting from the enhanced permeability and retention (EPR) effect. CONCLUSION: The biocompatible DPP-BDT NPs demonstrated to be strong NIR absorption property and PAI sensitivity. Besides, these novel DPP-BDT NPs can act not only as a PA imaging contrast agent but also as an imaging agent for PET.
Subject(s)
Ketones/chemistry , Nanoparticles/chemistry , Neoplasms, Experimental/diagnostic imaging , Photoacoustic Techniques/methods , Positron-Emission Tomography/methods , Pyrroles/chemistry , Animals , Contrast Media/chemistry , Copper Radioisotopes/administration & dosage , Copper Radioisotopes/pharmacokinetics , Female , Hep G2 Cells , Humans , Injections , Mice , Mice, Nude , Multimodal Imaging/methods , NIH 3T3 Cells , Nanoparticles/administration & dosage , Phosphatidylethanolamines/chemistry , Polyethylene Glycols/chemistry , Polymers/chemistry , Radiopharmaceuticals/administration & dosage , Radiopharmaceuticals/chemistry , Xenograft Model Antitumor AssaysABSTRACT
Niemann-Pick C disease (NPC) is an autosomal recessive lysosomal storage disorder resulting from mutations in the NPC1 (95% of cases) or NPC2 genes. Disturbance of copper homeostasis has been reported in NPC1 disease. In this study we have used whole-body positron emission tomography (PET) and brain electronic autoradiography with copper-64 (64Cu), in the form of the copper(II) bis(thiosemicarbazonato) complex 64Cu-GTSM, to image short-term changes in copper trafficking after intravenous injection in a transgenic mouse model of NPC1 disease. 64Cu-GTSM is taken up in all tissues and dissociates rapidly inside cells, allowing monitoring of the subsequent efflux and redistribution of 64Cu from all tissues. Significantly enhanced retention of 64Cu radioactivity was observed in brain, lungs and blood at 15 h post-injection in symptomatic Npc1-/- transgenic mice compared to wildtype controls. The enhanced retention of 64Cu in brain was confirmed by electronic autoradiography, particularly in the midbrain, thalamus, medulla and pons regions. Positron emission tomography imaging with 64Cu in selected chemical forms could be a useful diagnostic and research tool for the management and understanding of NPC1 disease.
Subject(s)
Copper Radioisotopes/metabolism , Copper Radioisotopes/pharmacokinetics , Disease Models, Animal , Niemann-Pick Disease, Type C/metabolism , Positron-Emission Tomography , Animals , Coordination Complexes/administration & dosage , Coordination Complexes/chemistry , Coordination Complexes/metabolism , Coordination Complexes/pharmacokinetics , Copper Radioisotopes/administration & dosage , Injections, Intravenous , Mice , Mice, Inbred BALB C , Mice, Knockout , Mice, Transgenic , Thiosemicarbazones/administration & dosage , Thiosemicarbazones/chemistry , Thiosemicarbazones/metabolism , Thiosemicarbazones/pharmacokineticsABSTRACT
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.
Subject(s)
B7-H1 Antigen/metabolism , Molecular Imaging/methods , Molecular Probes/administration & dosage , Neoplasms/diagnostic imaging , Positron Emission Tomography Computed Tomography/methods , Adult , Aged , Animals , B7-H1 Antigen/immunology , Cell Line, Tumor/transplantation , Copper Radioisotopes/administration & dosage , Copper Radioisotopes/chemistry , Disease Models, Animal , Female , Fibronectin Type III Domain/genetics , Humans , Male , Mice , Molecular Probes/chemistry , Molecular Probes/genetics , Neoplasms/pathology , Protein Binding , Protein Engineering , Radiopharmaceuticals/administration & dosage , Radiopharmaceuticals/chemistry , Recombinant Proteins/administration & dosage , Recombinant Proteins/genetics , Tissue DistributionABSTRACT
The receptor for advanced glycation end-products (RAGE) is central to multiple disease states, including diabetes-related conditions such as peripheral arterial disease (PAD). Despite RAGE's importance in these pathologies, there remains a need for a molecular imaging agent that can accurately assess RAGE levels in vivo. Therefore, we have developed a multimodal nanoparticle-based imaging agent targeted at RAGE with the well-characterized RAGE ligand, carboxymethyllysine (CML)-modified human serum albumin (HSA). Methods: A multimodal tracer (64Cu-Rho-G4-CML) was developed using a generation-4 (G4) polyamidoamine (PAMAM) dendrimer, conjugated with both rhodamine and copper-64 (64Cu) chelator (NOTA) for optical and PET imaging, respectively. First, 64Cu-Rho-G4-CML and its non-targeted analogue (64Cu-Rho-G4-HSA) were evaluated chemically using techniques such as dynamic light scattering (DLS), electron microscopy and nuclear magnetic resonance (NMR). The tracers' binding capabilities were examined at the cellular level and optimized using live and fixed HUVEC cells grown in 5.5-30 mM glucose, followed by in vivo PET-CT imaging, where the probes' kinetics, biodistribution, and RAGE targeting properties were examined in a murine model of hindlimb ischemia. Finally, histological assessment of RAGE levels in both ischemic and non-ischemic tissues was performed. Conclusions: Our RAGE-targeted probe demonstrated an average size of 450 nm, a Kd of 340-390 nM, rapid blood clearance, and a 3.4 times greater PET uptake in ischemic RAGE-expressing hindlimbs than their non-ischemic counterpart. We successfully demonstrated increased RAGE expression in a murine model of hindlimb ischemia and the feasibility for non-invasive examination of cellular, tissue, and whole-body RAGE levels with a molecularly targeted tracer.
Subject(s)
Antigens, Neoplasm/analysis , Glycation End Products, Advanced/metabolism , Lysine/analogs & derivatives , Mitogen-Activated Protein Kinases/analysis , Molecular Imaging/methods , Multimodal Imaging/methods , Nanoparticles/metabolism , Serum Albumin, Human/metabolism , Animals , Copper Radioisotopes/administration & dosage , Human Umbilical Vein Endothelial Cells , Humans , Lysine/metabolism , MiceABSTRACT
BACKGROUND: Hybrid positron emission tomography (PET)-magnetic resonance imaging (MRI) systems have been taken in use as new clinical diagnostic tools including detection and therapy planning of cancer. To reduce the amount of contrast agents injected in patients while fully benefitting both modalities, dual-modality probes are required. MATERIAL AND METHODS: This study was first aimed at developing a hybrid PET-MRI probe by labeling superparamagnetic iron oxide nanoparticles (SPIONs) with 64Cu using a fast and chelator-free conjugation method, and second, to demonstrate the ability of the agent to target sentinel lymph nodes (SLNs) in vivo using simultaneous PET-MRI imaging. RESULTS: High labeling efficiency of 97% produced within 10-15 min was demonstrated at room temperature. 64Cu-SPIONs were chemically stable in mouse serum for 24 h and after intradermal injection in the hind paw of C57BL/6J mice, demonstrated specific accumulation in the SLN. Simultaneous PET-MRI clearly demonstrated visualization of 64Cu-SPIONs, in dynamic and static imaging sequences up to 24 h after administration. CONCLUSION: The use of a single hybrid probe and simultaneous hybrid imaging provides an efficient, complementary integration of quantitation and is expected to improve preoperative planning and intraoperative guidance of cancer treatments.
Subject(s)
Contrast Media/administration & dosage , Lymphatic Vessels/diagnostic imaging , Magnetic Resonance Imaging/methods , Multimodal Imaging/methods , Positron-Emission Tomography/methods , Animals , Contrast Media/chemistry , Contrast Media/pharmacokinetics , Copper Radioisotopes/administration & dosage , Copper Radioisotopes/pharmacokinetics , Drug Evaluation, Preclinical , Female , Image Processing, Computer-Assisted , Magnetite Nanoparticles/administration & dosage , Magnetite Nanoparticles/chemistry , Mice, Inbred C57BL , Radioactive Tracers , Tissue DistributionABSTRACT
PURPOSE: To evaluate the diagnostic performance of 64Cu-PSMA-617 positron emission tomography (PET) with computed tomography (CT) for restaging prostate cancer after biochemical recurrence (BCR) and to compare it with 18F-choline PET/CT in a per-patient analysis. PATIENTS AND METHODS: An observational study was performed of 43 patients with BCR after laparoscopic radical prostatectomy who underwent 64Cu-PSMA-617 PET/CT and subsequently 18F-choline PET/CT for restaging. The detection rates (DR) of 64Cu-PSMA-617 PET/CT and of 18F-choline PET/CT were calculated by standardized maximum uptake value (SUVmax) at 4 hours and SUVmax at 1 hour as reference, respectively. Furthermore, univariate logistic regression analysis was carried out to identify independent predictive factors of positivity with 64Cu-PSMA-617 PET/CT. RESULTS: An overall positivity with 64Cu-PSMA-617 PET/CT was found in 32 patients (74.4%) versus 19 (44.2%) with 18F-choline PET/CT. Specifically, after stratifying for prostate-specific antigen (PSA) values, we found a good performance of 64Cu-PSMA-617 PET/CT at low PSA levels compared to 18F-choline PET/CT, with a DR of 57.1% versus 14.3% for PSA 0.2-0.5 ng/mL (P = .031), and of 60% versus 30% with PSA 0.5-1 ng/mL. At univariate binary logistic regression analysis, PSA level was the only independent predictor of 64Cu-PSMA-617 PET/CT positivity. No significant difference in terms of DR for both 64Cu-PSMA-617 PET/CT and 18F-choline PET/CT was found according to different Gleason score subgroups. CONCLUSION: In our study cohort, a better performance was observed for 64Cu-PSMA-617 PET/CT compared to 18F-choline PET/CT in restaging after BCR, especially in patients with low PSA values.
Subject(s)
Choline/analogs & derivatives , Copper Radioisotopes/administration & dosage , Dipeptides/metabolism , Heterocyclic Compounds, 1-Ring/metabolism , Neoplasm Recurrence, Local/diagnostic imaging , Prostatic Neoplasms/diagnostic imaging , Aged , Aged, 80 and over , Choline/administration & dosage , Early Detection of Cancer , Humans , Male , Middle Aged , Neoplasm Recurrence, Local/metabolism , Neoplasm Recurrence, Local/pathology , Neoplasm Recurrence, Local/surgery , Neoplasm Staging , Positron Emission Tomography Computed Tomography , Prostate-Specific Antigen , Prostatectomy , Prostatic Neoplasms/metabolism , Prostatic Neoplasms/pathology , Prostatic Neoplasms/surgery , Retrospective Studies , Sensitivity and SpecificityABSTRACT
The goal of this study was to establish the feasibility of integrating focused ultrasound (FUS)-mediated delivery of 64Cu-integrated gold nanoclusters (64Cu-AuNCs) to the pons for in vivo quantification of the nanocluster brain uptake using positron emission tomography (PET) imaging. FUS was targeted at the pons for the blood-brain barrier (BBB) disruption in the presence of systemically injected microbubbles, followed by the intravenous injection of 64Cu-AuNCs. The spatiotemporal distribution of the 64Cu-AuNCs in the brain was quantified using in vivo microPET/CT imaging at different time points post injection. Following PET imaging, the accumulation of radioactivity in the pons was further confirmed using autoradiography and gamma counting, and the gold concentration was quantified using inductively coupled plasma-mass spectrometry (ICP-MS). We found that the noninvasive and localized BBB opening by the FUS successfully delivered the 64Cu-AuNCs to the pons. We also demonstrated that in vivo real-time microPET/CT imaging was a reliable method for monitoring and quantifying the brain uptake of 64Cu-AuNCs delivered by the FUS. This drug delivery platform that integrates FUS, radiolabeled nanoclusters, and PET imaging provides a new strategy for noninvasive and localized nanoparticle delivery to the pons with concurrent in vivo quantitative imaging to evaluate delivery efficiency. The long-term goal is to apply this drug delivery platform to the treatment of pontine gliomas.
Subject(s)
Brain/metabolism , Copper Radioisotopes/administration & dosage , Gold/administration & dosage , Nanostructures/administration & dosage , Ultrasonic Waves , Animals , Brain/diagnostic imaging , Copper Radioisotopes/pharmacokinetics , Gold/pharmacokinetics , Male , Mice, Inbred C57BL , Microbubbles , Positron Emission Tomography Computed TomographyABSTRACT
Peripheral arterial disease (PAD) consists of a persistent obstruction of lower-extremity arteries further from the aortic bifurcation attributable to atherosclerosis. PAD is correlated with an elevated risk of morbidity and mortality as well as of deterioration of the quality of life with claudication and chronic leg ischemia being the most frequent complications. Therapeutic angiogenesis is a promising therapeutic strategy that aims to restore the blood flow to the ischemic limb. In this context, assessing the efficacy of pro-angiogenic treatment using a reliable noninvasive imaging technique would greatly benefit the implementation of this therapeutic approach. Herein, we describe the angiogenesis and perfusion recovery characteristics of a mouse model of PAD via in vivo positron emission tomography (PET) imaging of CD146 expression. For that, ischemia was generated by ligation and excision of the right femoral artery of Balb/C mice and confirmed through laser Doppler imaging. The angiogenic process, induced by ischemia, was noninvasively monitored and quantified through PET imaging of CD146 expression in the injured leg using a 64Cu-labeled anti-CD146 monoclonal antibody, 64Cu-NOTA-YY146, at post-operative days 3, 10, and 17. The CD146-specific character of 64Cu-NOTA-YY146 was verified via a blocking study performed in another cohort at day 10 after surgery. Tracer uptake was correlated with in situ CD146 expression by histological analysis. PET scan results indicated that 64Cu-NOTA-YY146 uptake in the injured leg was significantly higher, with the highest uptake with a value of 14.1 ± 2.0 %ID/g at post-operative day 3, compared to the normal contralateral hindlimb, at all time points (maximum uptake of 2.2 ± 0.2 %ID/g). The pre-injection of a blocking dose resulted in a significantly lower tracer uptake in the ischemic hindlimb on day 10 after surgery, confirming tracer specificity. CD146/CD31 immunofluorescent co-staining showed an excellent correlation between the high uptake of the tracer with in situ CD146 expression levels and a marked co-localization of CD146 and CD31 signals. In conclusion, persistent and CD146-specific tracer accumulation in the ischemic hindlimb was observed, confirming the feasibility of 64Cu-NOTA-YY146 to be used as an imaging agent to monitor the progression of angiogenesis and recovery in future PAD research.
Subject(s)
Antibodies, Monoclonal/administration & dosage , Ischemia/diagnostic imaging , Peripheral Arterial Disease/diagnostic imaging , Animals , Antibodies, Monoclonal/chemistry , CD146 Antigen/antagonists & inhibitors , CD146 Antigen/metabolism , Copper Radioisotopes/administration & dosage , Copper Radioisotopes/chemistry , Disease Models, Animal , Female , Femoral Artery/diagnostic imaging , Heterocyclic Compounds, 1-Ring/administration & dosage , Heterocyclic Compounds, 1-Ring/chemistry , Hindlimb/blood supply , Humans , Ischemia/etiology , Ischemia/pathology , Laser-Doppler Flowmetry , Mice , Mice, Inbred BALB C , Molecular Imaging/methods , Peripheral Arterial Disease/etiology , Peripheral Arterial Disease/pathology , Positron-Emission Tomography/methods , X-Ray MicrotomographyABSTRACT
Deposition of liposomal drugs into solid tumors is a potentially rate-limiting step for drug delivery and has substantial variability that may influence probability of response. Tumor deposition is a shared mechanism for liposomal therapeutics such that a single companion diagnostic agent may have utility in predicting response to multiple nanomedicines. Methods: We describe the development, characterization and preclinical proof-of-concept of the positron emission tomography (PET) agent, MM-DX-929, a drug-free untargeted 100 nm PEGylated liposome stably entrapping a chelated complex of 4-DEAP-ATSC and 64Cu (copper-64). MM-DX-929 is designed to mimic the biodistribution of similarly sized therapeutic agents and enable quantification of deposition in solid tumors. Results: MM-DX-929 demonstrated sufficient in vitro and in vivo stability with PET images accurately reflecting the disposition of liposome nanoparticles over the time scale of imaging. MM-DX-929 is also representative of the tumor deposition and intratumoral distribution of three different liposomal drugs, including targeted liposomes and those with different degrees of PEGylation. Furthermore, stratification using a single pre-treatment MM-DX-929 PET assessment of tumor deposition demonstrated that tumors with high MM-DX-929 deposition predicted significantly greater anti-tumor activity after multi-cycle treatments with different liposomal drugs. In contrast, MM-DX-929 tumor deposition was not prognostic in untreated tumor-bearing xenografts, nor predictive in animals treated with small molecule chemotherapeutics. Conclusions: These data illustrate the potential of MM-DX-929 PET as a companion diagnostic strategy to prospectively select patients likely to respond to liposomal drugs or nanomedicines of similar molecular size.
Subject(s)
Copper Radioisotopes/administration & dosage , Copper Radioisotopes/chemistry , Liposomes/chemistry , Nanoparticles/administration & dosage , Nanoparticles/chemistry , Neoplasms/drug therapy , Animals , Cell Line, Tumor , Drug Delivery Systems/methods , Female , HT29 Cells , Humans , Mice , Nanomedicine/methods , Neoplasms/metabolism , Polyethylene Glycols/chemistry , Positron-Emission Tomography/methods , Tissue Distribution/physiologyABSTRACT
Radiolabeled antibodies, polyethylene glycol-conjugated (PEGylated) peptides, liposomes, and other materials were investigated as positron-emission tomography (PET) probes. These substances accumulate in tumors but often remain too long in circulation. We investigated the combination of intravenous urokinase injection and its substrate linker as a triggered radioisotope clearance enhancement system to improve imaging contrast. To this end, we synthesized a four-arm PEGylated 64Cu-bombesin analog tetramer with a urokinase substrate linker. In mouse blood, it was almost perfectly cleaved and degraded into smaller radioactive fragments in vitro with urokinase (≥20,000â¯IU/mL). In mouse blood circulation, â¼50-65% of the probe was rapidly degraded after the urokinase injection and the radioactive fragments were eliminated mainly from the kidney. In contrast, tumor radioactivity levels did not change, and therefore, the tumors were clearly visualized. The tumor/blood ratio, an indicator of imaging contrast, increased 2.5 times, while elimination of the radioisotope from the blood was enhanced. This approach has the potential to improve imaging contrast using various PET probes. It could also shorten the time required to obtain sufficient contrast and decrease patient radiation exposure.
Subject(s)
Bombesin/administration & dosage , Coordination Complexes/administration & dosage , Copper Radioisotopes/administration & dosage , Copper/chemistry , Polyethylene Glycols/administration & dosage , Positron-Emission Tomography/methods , Prostatic Neoplasms/diagnostic imaging , Radiopharmaceuticals/administration & dosage , Urokinase-Type Plasminogen Activator/administration & dosage , Animals , Bombesin/analogs & derivatives , Bombesin/chemistry , Bombesin/pharmacokinetics , Cell Line, Tumor , Coordination Complexes/chemistry , Coordination Complexes/pharmacokinetics , Copper Radioisotopes/chemistry , Humans , Injections, Intravenous , Male , Mice, Inbred BALB C , Mice, Nude , Polyethylene Glycols/chemistry , Predictive Value of Tests , Prostatic Neoplasms/metabolism , Proteolysis , Radiopharmaceuticals/chemistry , Radiopharmaceuticals/pharmacokinetics , Urokinase-Type Plasminogen Activator/metabolismABSTRACT
Receptor-mediated internalization followed by trafficking and degradation of antibody-conjugates (ACs) via the endosomal-lysosomal pathway is the major mechanism for delivering molecular payloads inside target tumor cells. Although a mainstay for delivering payloads with clinically approved ACs in cancer treatment and imaging, tumor cells are often able to decrease intracellular payload concentrations and thereby reduce the effectiveness of the desired application. Thus, increasing payload intracellular accumulation has become a focus of attention for designing next-generation ACs. We developed a composite compound (ChAcNLS) that enables ACs to escape endosome entrapment and route to the nucleus resulting in the increased intracellular accumulation as an interleukin-5 receptor α-subunit (IL-5Rα)-targeted agent for muscle invasive bladder cancer (MIBC). We constructed 64Cu-A14-ChAcNLS, 64Cu-A14-NLS, and 64Cu-A14 and evaluated their performance by employing mechanistic studies for endosome escape coupled to nuclear routing and determining whether this delivery system results in improved 64Cu cellular accumulation. ACs consisting of â¼20 ChAcNLS or NLS moieties per 64Cu-A14 were prepared in good yield, high monomer content, and maintaining high affinity for IL-5Rα. Confocal microscopy analysis demonstrated ChAcNLS mediated efficient endosome escape and nuclear localization. 64Cu-A14-ChAcNLS increased 64Cu cellular accumulation in HT-1376 and HT-B9 cells relative to 64Cu-A14 and 64Cu-A14-NLS. In addition, we tested 64Cu-A14-ChAcNLS in vivo to evaluate its tissue distribution properties and, ultimately, tumor uptake and targeting. A model of human IL-5Rα MIBC was developed by implanting NOD/SCID mice with subcutaneous HT-1376 or HT-B9MIBC tumors, which grow containing high and low IL-5Rα-positive tumor cell densities, respectively. ACs were intravenously injected, and daily blood sampling, biodistribution at 48 and 96 h, and positron emission tomography (PET) at 24 and 48 h were performed. Region of interest (ROI) analysis was also performed on reconstructed PET images. Pharmacokinetic analysis and biodistribution studies showed that 64Cu-A14-ChAcNLS had faster clearance rates from the blood and healthy organs relative to 64Cu-A14. However, 64Cu-A14-ChAcNLS maintained comparable tumor accumulation relative to 64Cu-A14. This resulted in 64Cu-A14-ChAcNLS having superior tumor/normal tissue ratios at both 48 and 96 h biodistribution time points. Visualization of AC distribution by PET and ROI analysis confirmed that 64Cu-A14-ChAcNLS had improved targeting of MIBC tumor relative to 64Cu-A14. In addition, 64Cu-A14 modified with only NLS had poor tumor targeting. This was a result of poor tumor uptake due to extremely rapid clearance. Thus, the overall findings in this model of human IL-5Rα-positive MIBC describe an endosome escape-nuclear localization cholic-acid-linked peptide that substantially enhances AC cellular accumulation and tumor targeting.
Subject(s)
Cholic Acid/chemistry , Cholic Acid/pharmacokinetics , Immunoconjugates/chemistry , Immunoconjugates/pharmacokinetics , Interleukin-5 Receptor alpha Subunit/analysis , Urinary Bladder Neoplasms/diagnostic imaging , Animals , Cell Line, Tumor , Cholic Acid/administration & dosage , Copper Radioisotopes/administration & dosage , Copper Radioisotopes/chemistry , Copper Radioisotopes/pharmacokinetics , Drug Delivery Systems , Humans , Immunoconjugates/administration & dosage , Immunoconjugates/immunology , Interleukin-5 Receptor alpha Subunit/immunology , Mice, Inbred NOD , Mice, SCID , Positron-Emission Tomography/methods , Tissue Distribution , Urinary Bladder Neoplasms/immunology , Urinary Bladder Neoplasms/therapyABSTRACT
Wilson's disease is a genetic disorder that causes excessive accumulation of copper in the body, leading to toxic damage, especially in the liver and nervous system. The current treatment cause burdensome side effects. We describe the use of chemically modified biopolymer carriers based on microcrystalline cellulose and chitosan containing the highly specific copper chelator 8-hydroxyquinoline as a new type of therapy for Wilson's disease. The chelators can scavenges copper ions released from food during digestion and copper ions present in secretions in the gastrointestinal tract. Because the chelator is covalently bound to indigestible biopolymer carriers (crosslinked chitosan or modified cellulose), it is not taken up by the gastrointestinal tract and it can be eliminated through the feces, avoiding unwanted side effects. This concept was tested on Wistar rats, which received a radioactive 64CuCl2 solution together with the polymers with covalently bound 8-hydroxyquinoline through a gastric probe. 64Copper complex uptake from the gastrointestinal tract was significantly inhibited by both chelating polymers. With the modified polymers, the presence of 64Cu was detected mostly in the gastrointestinal tract, not in the internal organs. These findings indicate modified cellulose and crosslinked chitosan, with covalently bound 8-hydroxyquinoline exhibited the potential to be excellent therapeutics for treating Wilson's disease.
Subject(s)
Cellulose/administration & dosage , Chitosan/administration & dosage , Hepatolenticular Degeneration/drug therapy , Oxyquinoline/administration & dosage , Animals , Cellulose/pharmacokinetics , Chitosan/pharmacokinetics , Copper , Copper Radioisotopes/administration & dosage , Copper Radioisotopes/pharmacokinetics , Gastrointestinal Tract/metabolism , Hepatolenticular Degeneration/metabolism , Oxyquinoline/pharmacokinetics , Rats, WistarABSTRACT
To develop a radioactive metal complex platform for tumor theranostics, we introduced three radiopharmaceutical derivatives of 1,4,7,10-tetraazacyclododecane-1,4,7-trisacetic acid-benzothiazole aniline (DO3A-BTA, L1) labeled with medical radioisotopes for diagnosis (68Ga/64Cu) and therapy (177Lu). The tumor-targeting ability of these complexes was demonstrated in a cellular uptake experiment, in which 177Lu-L1 exhibited markedly higher uptake in HeLa cells than the 177Lu-1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid complex. According to in vivo positron emission tomography imaging, high accumulation of 68Ga-L1 and 64Cu-L1 was clearly visualized in the tumor site, while 177Lu-L1 showed therapeutic efficacy in therapy experiments. Consequently, this molecular platform represents a useful approach in nuclear medicine toward tumor-theranostic radiopharmaceuticals when 68Ga-L1 or 64Cu-L1 is used for diagnosis, 177Lu-L1 is used for therapy, or two of the compounds are used in conjunction with each other.
Subject(s)
Aniline Compounds/administration & dosage , Benzothiazoles/administration & dosage , Heterocyclic Compounds, 1-Ring/administration & dosage , Radiopharmaceuticals/administration & dosage , Theranostic Nanomedicine/methods , Aniline Compounds/chemistry , Animals , Benzothiazoles/chemistry , Copper Radioisotopes/administration & dosage , Copper Radioisotopes/chemistry , Female , Gallium Radioisotopes/administration & dosage , Gallium Radioisotopes/chemistry , HEK293 Cells , HeLa Cells , Heterocyclic Compounds, 1-Ring/chemistry , Humans , Lutetium/administration & dosage , Lutetium/chemistry , Mice , Mice, Inbred BALB C , Mice, Nude , Positron-Emission Tomography/methods , Radioisotopes/administration & dosage , Radioisotopes/chemistry , Radiopharmaceuticals/chemistry , Treatment Outcome , Uterine Cervical Neoplasms/diagnostic imaging , Uterine Cervical Neoplasms/therapy , Xenograft Model Antitumor AssaysABSTRACT
Radiation therapy may affect several important parameters in the tumor microenvironment and thereby influence the accumulation of liposomes by the enhanced permeability and retention (EPR)-effect. Here we investigate the effect of single dose radiation therapy on liposome tumor accumulation by PET/CT imaging using radiolabeled liposomes. Head and neck cancer xenografts (FaDu) and syngenic colorectal (CT26) cancer models were investigated. Radiotherapy displayed opposite effects in the two models. FaDu tumors displayed increased mean accumulation of liposomes for radiation doses up to 10 Gy, whereas CT26 tumors displayed a tendency for decreased accumulation. Tumor hypoxia was found negatively correlated to microregional distribution of liposomes. However, liposome distribution in relation to hypoxia was improved at lower radiation doses. The study reveals that the heterogeneity in liposome tumor accumulation between tumors and different radiation protocols are important factors that need to be taken into consideration to achieve optimal effect of liposome based radio-sensitizer therapy.
Subject(s)
Colorectal Neoplasms/metabolism , Gamma Rays/therapeutic use , Head and Neck Neoplasms/metabolism , Liposomes/pharmacokinetics , Animals , Colorectal Neoplasms/diagnostic imaging , Colorectal Neoplasms/pathology , Colorectal Neoplasms/radiotherapy , Copper Radioisotopes/administration & dosage , Copper Radioisotopes/pharmacokinetics , Female , Head and Neck Neoplasms/diagnostic imaging , Head and Neck Neoplasms/pathology , Head and Neck Neoplasms/radiotherapy , Humans , Hypoxia/metabolism , Liposomes/administration & dosage , Mice , Mice, Nude , Positron Emission Tomography Computed Tomography/methods , Radiation Dosage , Tissue Distribution , Treatment Outcome , Tumor Cells, Cultured , Xenograft Model Antitumor AssaysABSTRACT
Liposomes are nanoparticles used in drug delivery that distribute over several days in humans and larger animals. Radiolabeling with long-lived positron emission tomography (PET) radionuclides, such as manganese-52 (52Mn, T½=5.6days), allow the imaging of this biodistribution. We report optimized protocols for radiolabeling liposomes with 52Mn, through both remote-loading and surface labeling. For comparison, liposomes were also remote-loaded and surface labeled with copper-64 (64Cu, T½=12.7h) through conventional means. The chelator DOTA was used in all cases. The in vivo stability of radiometal chelates is widely debated but studies that mimic a realistic in vivo setting are lacking. Therefore, we employed these four radiolabeled liposome types as platforms to demonstrate a new concept for such in vivo evaluation, here of the chelates 52Mn-DOTA and 64Cu-DOTA. This was done by comparing "shielded" remote-loaded with "exposed" surface labeled variants in a CT26 tumor-bearing mouse model. Remote loading (90min at 55°C) and surface labeling (55°C for 2h) of 52Mn gave excellent radiolabeling efficiencies of 97-100% and 98-100% respectively, and the liposome biodistribution was imaged by PET for up to 8days. Liposomes with surface-conjugated 52Mn-DOTA exhibited a significantly shorter plasma half-life (T½=14.4h) when compared to the remote-loaded counterpart (T½=21.3h), whereas surface-conjugated 64Cu-DOTA cleared only slightly faster and non-significantly, when compared to remote-loaded (17.2±2.9h versus 20.3±1.2h). From our data, we conclude the successful remote-loading of liposomes with 52Mn, and furthermore that 52Mn-DOTA may be unstable in vivo whereas 64Cu-DOTA appears suitable for quantitative imaging.
Subject(s)
Chelating Agents/administration & dosage , Copper Radioisotopes/administration & dosage , Heterocyclic Compounds, 1-Ring/administration & dosage , Manganese/administration & dosage , Radioisotopes/administration & dosage , Radiopharmaceuticals/administration & dosage , Animals , Cell Line, Tumor , Chelating Agents/pharmacokinetics , Copper Radioisotopes/pharmacokinetics , Heterocyclic Compounds, 1-Ring/pharmacokinetics , Liposomes , Manganese/pharmacokinetics , Mice, Inbred BALB C , Neoplasms/diagnostic imaging , Positron-Emission Tomography , Radioisotopes/pharmacokinetics , Radiopharmaceuticals/pharmacokinetics , Tissue DistributionABSTRACT
Alzheimer's disease can involve brain copper dyshomeostasis. We aimed to determine the effect of AD-like pathology on 64Cu trafficking in mice, using positron emission tomography (PET imaging), during 24 hours after intravenous administration of ionic 64Cu (Cu(ii) acetate) and 64Cu-GTSM (GTSMH2 = glyoxalbis(thiosemicarbazone)). Copper trafficking was evaluated in 6-8-month-old and 13-15 month-old TASTPM transgenic and wild-type mice, by imaging 0-30 min and 24-25 h after intravenous administration of 64Cu tracer. Regional 64Cu distribution in brains was compared by ex vivo autoradiography to that of amyloid-ß plaque. 64Cu-acetate showed uptake in, and excretion through, liver and kidneys. There was minimal uptake in other tissues by 30 minutes, and little further change after 24 h. Radioactivity within brain was focussed in and around the ventricles and was significantly greater in younger mice. 64CuGTSM was taken up in all tissues by 30 min, remaining high in brain but clearing substantially from other tissues by 24 h. Distribution in brain was not localised to specific regions. TASTPM mice showed no major changes in global or regional 64Cu brain uptake compared to wildtype after administration of 64Cu acetate (unlike 64Cu-GTSM) but efflux of 64Cu from brain by 24 h was slightly greater in 6-8 month-old TASTPM mice than in wildtype controls. Changes in copper trafficking associated with Alzheimer's-like pathology after administration of ionic 64Cu are minor compared to those observed after administration of 64Cu-GTSM. PET imaging with 64Cu could help understand changes in brain copper dynamics in AD and underpin new clinical diagnostic imaging methods.
Subject(s)
Alzheimer Disease/metabolism , Copper Radioisotopes/pharmacokinetics , Disease Models, Animal , Positron-Emission Tomography/methods , Acetates/administration & dosage , Acetates/pharmacokinetics , Administration, Intravenous , Alzheimer Disease/diagnostic imaging , Animals , Biological Transport , Brain/metabolism , Coordination Complexes , Copper Radioisotopes/administration & dosage , Humans , Metabolic Clearance Rate , Mice, Transgenic , Organometallic Compounds/administration & dosage , Organometallic Compounds/pharmacokinetics , Thiosemicarbazones/administration & dosage , Thiosemicarbazones/pharmacokinetics , Tissue DistributionABSTRACT
Purpose: Therapeutic nanoparticles are designed to deliver their drug payloads through enhanced permeability and retention (EPR) in solid tumors. The extent of EPR and its variability in human tumors is highly debated and has been proposed as an explanation for variable responses to therapeutic nanoparticles in clinical studies.Experimental Design: We assessed the EPR effect in patients using a 64Cu-labeled nanoparticle, 64Cu-MM-302 (64Cu-labeled HER2-targeted PEGylated liposomal doxorubicin), and imaging by PET/CT. Nineteen patients with HER2-positive metastatic breast cancer underwent 2 to 3 PET/CT scans postadministration of 64Cu-MM-302 as part of a clinical trial of MM-302 plus trastuzumab with and without cyclophosphamide (NCT01304797).Results: Significant background uptake of 64Cu-MM-302 was observed in liver and spleen. Tumor accumulation of 64Cu-MM-302 at 24 to 48 hours varied 35-fold (0.52-18.5 %ID/kg), including deposition in bone and brain lesions, and was independent of systemic plasma exposure. Computational analysis quantified rates of deposition and washout, indicating peak liposome deposition at 24 to 48 hours. Patients were classified on the basis of 64Cu-MM-302 lesion deposition using a cut-off point that is comparable with a response threshold in preclinical studies. In a retrospective exploratory analysis of patient outcomes relating to drug levels in tumor lesions, high 64Cu-MM-302 deposition was associated with more favorable treatment outcomes (HR = 0.42).Conclusions: These findings provide important evidence and quantification of the EPR effect in human metastatic tumors and support imaging nanoparticle deposition in tumors as a potential means to identify patients well suited for treatment with therapeutic nanoparticles. Clin Cancer Res; 23(15); 4190-202. ©2017 AACR.
Subject(s)
Bone Neoplasms/drug therapy , Brain Neoplasms/drug therapy , Breast Neoplasms/drug therapy , Doxorubicin/analogs & derivatives , Nanoparticles/administration & dosage , Adolescent , Adult , Aged , Bone Neoplasms/diagnostic imaging , Bone Neoplasms/pathology , Bone Neoplasms/secondary , Brain Neoplasms/diagnostic imaging , Brain Neoplasms/pathology , Brain Neoplasms/secondary , Breast Neoplasms/blood , Breast Neoplasms/diagnostic imaging , Breast Neoplasms/pathology , Cell Membrane Permeability/drug effects , Cell Membrane Permeability/radiation effects , Copper Radioisotopes/administration & dosage , Copper Radioisotopes/chemistry , Cyclophosphamide/administration & dosage , Doxorubicin/administration & dosage , Doxorubicin/chemistry , Female , Humans , Liver/diagnostic imaging , Liver/drug effects , Middle Aged , Nanoparticles/chemistry , Neoplasm Metastasis , Polyethylene Glycols/administration & dosage , Polyethylene Glycols/chemistry , Positron Emission Tomography Computed Tomography , Receptor, ErbB-2/blood , Spleen/diagnostic imaging , Spleen/pathology , Trastuzumab/administration & dosageABSTRACT
Minimizing the sequestration of nanomaterials (NMs) by the reticuloendothelial system (RES) can enhance the circulation time of NMs, and thus increase their tumor-specific accumulation. Liposomes are generally regarded as safe (GRAS) agents that can block the RES reversibly and temporarily. With the help of positron emission tomography (PET), we monitored the in vivo tissue distribution of 64Cu-labeled 40 × 10 nm gold nanorods (Au NRs) after pretreatment with liposomes. We systematically studied the effectiveness of liposome administration by comparing (1) differently charged liposomes; (2) different liposome doses; and (3) varying time intervals between liposome dose and NR dose. By pre-injecting 400 µmol/kg positively charged liposomes into mice 5 h before the Au NRs, the liver and spleen uptakes of Au NRs decreased by 30% and 53%, respectively. Significantly, U87MG tumor uptake of Au NRs increased from 11.5 ± 1.1 %ID/g to 16.1 ± 1.3 %ID/g at 27 h post-injection. Quantitative PET imaging is a valuable tool to understand the fate of NMs in vivo and cationic liposomal pretreatment is a viable approach to reduce RES clearance, prolong circulation, and improve tumor uptake.
Subject(s)
Antineoplastic Agents/administration & dosage , Antineoplastic Agents/pharmacokinetics , Glioblastoma/drug therapy , Gold/administration & dosage , Liposomes/administration & dosage , Mononuclear Phagocyte System/metabolism , Nanoparticles/administration & dosage , Animals , Copper Radioisotopes/administration & dosage , Disease Models, Animal , Liver/diagnostic imaging , Mice , Positron-Emission Tomography , Spleen/diagnostic imagingABSTRACT
64Cu emits positrons as well as ß- particles and Auger and internal conversion electrons useful for radiotherapy. Our objective was to model the cellular dosimetry of 64Cu under different geometries commonly used to study the cytotoxic effects of 64Cu. METHODS: Monte Carlo N-Particle (MCNP) was used to simulate the transport of all particles emitted by 64Cu from the cell surface (CS), cytoplasm (Cy), or nucleus (N) of a single cell; monolayer in a well (radius = 0.32-1.74 cm); or a sphere (radius = 50-6,000 µm) of cells to calculate S values. The radius of the cell and N ranged from 5 to 12 µm and 2 to 11 µm, respectively. S values were obtained by MIRDcell for comparison. MCF7/HER2-18 cells were exposed in vitro to 64Cu-labeled trastuzumab. The subcellular distribution of 64Cu was measured by cell fractionation. The surviving fraction was determined in a clonogenic assay. RESULTS: The relative differences of MCNP versus MIRDcell self-dose S values (Sself) for 64Cu ranged from -0.2% to 3.6% for N to N (SNâN), 2.3% to 8.6% for Cy to N (SNâCy), and -12.0% to 7.3% for CS to N (SNâCS). The relative differences of MCNP versus MIRDcell cross-dose S values were 25.8%-30.6% for a monolayer and 30%-34% for a sphere, respectively. The ratios of SNâN versus SNâCy and SNâCy versus SNâCS decreased with increasing ratio of the N of the cell versus radius of the cell and the size of the monolayer or sphere. The surviving fraction of MCF7 /: HER2-18 cells treated with 64Cu-labeled trastuzumab (0.016-0.368 MBq/µg, 67 nM) for 18 h versus the absorbed dose followed a linear survival curve with α = 0.51 ± 0.05 Gy-1 and R2 = 0.8838. This is significantly different from the linear quadratic survival curve of MCF7 /: HER2-18 cells exposed to γ-rays. CONCLUSION: MCNP- and MIRDcell-calculated S values agreed well. 64Cu in the N increases the dose to the N in isolated single cells but has less effect in a cell monolayer or small cluster of cells simulating a micrometastasis, and little effect in a sphere analogous to a tumor xenograft compared with 64Cu in the Cy or on the CS. The dose deposited by 64Cu is less effective for cell killing than γ-rays.