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Lipid nanodiscs (LNDs), comprising a phospholipid bilayer encircled by two molecules of a recombinant membrane scaffold protein, can be targeted to tumors with covalently attached antibodies (Abs) or their fragments. Antibody attachment to click chemistry based PEGylated lipids on LNDs including DOTA allowed PET imaging with the positron emitter 64Cu. Carcinoembryonic antigen (CEA) positive tumors in CEA transgenic mice were chosen as a tumor target. Fab' fragments, that otherwise are rapidly cleared by the kidney due to their small size, were retained in circulation when conjugated to LNDs. Untargeted PET imaging of 64Cu-DOTA-LNDs revealed low tumor uptake (4-5% ID/g) in the range expected for the enhanced permeability retention (EPR) effect with high liver uptake (17-21% ID/g) indicating gut clearance. Fab' targeted LNDs showed little improvement over untargeted LNDs, but intact IgG targeted LNDs gave high tumor uptake (40% ID/g) with low liver (8% ID/g), demonstrating that tumor targeting with antibody conjugated LNDs is feasible.
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Antígeno Carcinoembrionário/metabolismo , Compostos Heterocíclicos com 1 Anel/química , Imunoconjugados/química , Neoplasias Mamárias Experimentais/diagnóstico por imagem , Fosfolipídeos/química , Polietilenoglicóis/química , Tomografia por Emissão de Pósitrons/métodos , Animais , Azidas/química , Linhagem Celular Tumoral , Radioisótopos de Cobre , Feminino , Neoplasias Mamárias Experimentais/metabolismo , Camundongos , Nanoestruturas/químicaRESUMO
As a growing number of patients with multiple myeloma (MM) respond to upfront therapies while eventually relapsing in a time frame that is often unpredictable, attention has increasingly focused on developing novel diagnostic criteria to also account for disease dissemination. Positron emission tomography/computed tomography (PET/CT) is often used as a noninvasive monitoring strategy to assess cancer cell dissemination, but because the uptake of the currently used radiotracer 18fluorodeoxyglucose (18F-FDG) is a function of the metabolic activity of both malignant and nonmalignant cells, the results frequently lack sufficient specificity. Radiolabeled antibodies targeting MM tissue may detect disease irrespective of cell metabolism. Hence, we conjugated the clinically significant CD38-directed human antibody daratumumab (Darzalex [Dara]) to the DOTA chelator and labeled it with the positron-emitting radionuclide copper 64 (64Cu; 64Cu-DOTA-Dara). Here, we show that 64Cu-DOTA-Dara can efficiently bind CD38 on the surface of MM cells and was mainly detected in the bones associated with tumor in a MM murine model. We also show that PET/CT based on 64Cu-DOTA-Dara displays a higher resolution and specificity to detect MM cell dissemination than does 18F-FDG PET/CT and was even more sensitive than were bioluminescence signals. We therefore have supporting evidence for using 64Cu-DOTA-Dara as a novel imaging agent for MM.
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Anticorpos Monoclonais , Radioisótopos de Cobre , Mieloma Múltiplo/diagnóstico , Tomografia por Emissão de Pósitrons combinada à Tomografia Computadorizada/métodos , Animais , Anticorpos Monoclonais/farmacocinética , Linhagem Celular Tumoral , Rastreamento de Células/métodos , Radioisótopos de Cobre/farmacocinética , Meia-Vida , Xenoenxertos , Humanos , Camundongos , Mieloma Múltiplo/metabolismo , Transplante de Neoplasias , Traçadores RadioativosRESUMO
BACKGROUND: Bispecific T-cell engaging antibodies (BiTES), comprising dual anti-CD3 and anti-tumor antigen scFv fragments, are important therapeutic agents for the treatment of cancer. The dual scFv construct for BiTES requires proper protein folding while their small molecular size leads to rapid kidney clearance. METHODS: An intact (150 kDa) anti-tumor antigen antibody to CEA was joined in high yield (ca. 30%) to intact (150 kDa) anti-murine and anti-human CD3 antibodies using hinge region specific Click chemistry to form dual-specific, bivalent BiTES (dbBiTES, 300 kDa). dbBiTEs were tested in vitro by EM, flow cytometry and cell cytoxicity and in vivo by PET tumor imaging and redirected T-cell therapy. RESULTS: The interlocked hinge regions are compatible with a structural model that fits the electron micrographs of 300 kDa particles. Compared to intact anti-CEA antibody, dbBiTES exhibit high in vitro cytotoxicity, high in vivo tumor targeting as demonstrated by PET imaging, and redirected dbBiTE coated T-cells (1 microgram/10 million cells) that kill CEA+ target cells in vivo in CEA transgenic mice. CONCLUSION: dbBiTE redirected T-cell therapy is a promising, efficient approach for targeting and killing cancer cells.
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Anticorpos Biespecíficos/uso terapêutico , Antígeno Carcinoembrionário/imunologia , Terapia Baseada em Transplante de Células e Tecidos/métodos , Neoplasias do Colo/terapia , Imunoterapia/métodos , Linfócitos T/imunologia , Animais , Complexo CD3/imunologia , Antígeno Carcinoembrionário/genética , Linhagem Celular Tumoral , Neoplasias do Colo/diagnóstico por imagem , Citotoxicidade Imunológica , Humanos , Ativação Linfocitária/imunologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Simulação de Dinâmica Molecular , Tomografia por Emissão de Pósitrons , Dobramento de Proteína , Anticorpos de Cadeia Única/imunologia , Transfecção , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
Umbilical cord mesenchymal stem cell-derived extracellular vesicles (UC-MSC-EVs) have become an emerging strategy for treating various autoimmune and metabolic disorders, particularly diabetes. Delivery of UC-MSC-EVs is essential to ensure optimal efficacy of UC-MSC-EVs. To develop safe and superior EVs-based delivery strategies, we explored nuclear techniques including positron emission tomography (PET) to evaluate the delivery of UC-MSC-EVs in vivo. In this study, human UC-MSC-EVs were first successfully tagged with I-124 to permit PET determination. Intravenous (I.V.) and intra-arterial (I.A.) administration routes of [124I]I-UC-MSC-EVs were compared and evaluated by in vivo PET-CT imaging and ex vivo biodistribution in a non-diabetic Lewis (LEW) rat model. For I.A. administration, [124I]I-UC-MSC-EVs were directly infused into the pancreatic parenchyma via the celiac artery. PET imaging revealed that the predominant uptake occurred in the liver for both injection routes, and further imaging characterized clearance patterns of [124I]I-UC-MSC-EVs. For biodistribution, the uptake (%ID/gram) in the spleen was significantly higher for I.V. administration compared to I.A. administration (1.95 ± 0.03 and 0.43 ± 0.07, respectively). Importantly, the pancreas displayed similar uptake levels between the two modalities (0.20 ± 0.06 for I.V. and 0.24 ± 0.03 for I.A.). Therefore, our initial data revealed that both routes had similar delivery efficiency for [124I]I-UC-MSC-EVs except in the spleen and liver, considering that higher spleen uptake could enhance immunomodulatory application of UC-MSC-EVs. These findings could guide the development of safe and efficacious delivery strategies for UC-MSC-EVs in diabetes therapies, in which a minimally invasive I.V. approach would serve as a better delivery strategy. Further confirmation studies are ongoing.
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Background: The aim of the study was to perform PET imaging and radiotherapy with a novel neurotensin derivative for neurotensin receptor 1 (NTSR-1)-positive tumors in an animal model. Materials and Methods: A di-DOTA analog of NT(6-13) with three unnatural amino acids was synthesized and radiolabeled with either 64Cu or 68Ga and tested for serum stability and tumor imaging in mice bearing NTSR-1-positive PC3, and HT29 xenografts. A dose-response therapy study was performed with 18.5, 37, and 74 kBq of 225Ac-di-DOTA-α,É-Lys-NT(6-13). Results: 68Ga-di-DOTA-α,É-Lys-NT(6-13) was >99% stable in serum for 48 h, had an IC50 of 5 nM using 125I labeled NT(8-13) for binding to HT-29 cells, and high uptake in tumor models expressing NTSR-1. 68Ga-di-DOTA-α,É-Lys-NT(6-13) had an average %ID/g (n = 4) at 2 h of 4.0 for tumor, 0.5 for blood, 12.0 for kidney, and <1 for other tissues, resulting in a favorable T/B of 8. Mean survivals of tumor-bearing mice treated with 18.5 or 37 kBq of 225Ac-di-DOTA-α,É-Lys-NT(6-13) were 81 and 93 d, respectively, versus 53 d for controls. Whole-body toxicity was seen for the 74 kBq dose. Conclusions: Based on the results of the animal model, di-DOTA-α,É-Lys-NT(6-13) is a useful imaging agent for NTSR-1-positive tumors when radiolabeled with 68Ga, and when radiolabeled with 225Ac, a potent therapeutic agent.
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Compostos Heterocíclicos com 1 Anel/farmacologia , Neoplasias , Neurotensina , Tomografia por Emissão de Pósitrons/métodos , Receptores de Neurotensina/metabolismo , Animais , Quelantes/farmacologia , Modelos Animais de Doenças , Radioisótopos de Gálio , Células HT29 , Xenoenxertos , Humanos , Camundongos , Neoplasias/diagnóstico por imagem , Neoplasias/metabolismo , Neoplasias/terapia , Neurotensina/análogos & derivados , Neurotensina/metabolismo , Avaliação de Resultados em Cuidados de SaúdeRESUMO
Radioimmunotherapy, an approach using radiolabeled antibodies, has had minimal success in the clinic with several ß-emitting radionuclides for the treatment of ovarian cancer. Alternatively, radioimmunotherapy with α-emitters offers the advantage of depositing much higher energy over shorter distances but was thought to be inappropriate for the treatment of solid tumors, for which antibody penetration is limited to a few cell diameters around the vascular system. However, the deposition of high-energy α-emitters to tumor markers adjacent to a typical leaky tumor vascular system may have large antitumor effects at the tumor vascular level, and their reduced penetration in normal tissue would be expected to lower off-target toxicity. Methods: To evaluate this concept, DOTAylated-huCC49 was labeled with the α-emitter 225Ac to target tumor-associated glycoprotein 72-positive xenografts in a murine model of ovarian cancer. Results:225Ac-labeled DOTAylated-huCC49 radioimmunotherapy significantly reduced tumor growth in a dose-dependent manner (1.85, 3.7, and 7.4 kBq), with the 7.4-kBq dose extending survival by more than 3-fold compared with the untreated control. Additionally, a multitreatment regime (1.85 kBq followed by 5 weekly doses of 0.70 kBq for a total of 5.4 kBq) extended survival almost 3-fold compared with the untreated control group, without significant off-target toxicity. Conclusion: These results establish the potential for antibody-targeted α-radionuclide therapy for ovarian cancer, which may be generalized to α-radioimmunotherapy in other solid tumors.
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Actínio/uso terapêutico , Partículas alfa/uso terapêutico , Anticorpos Monoclonais/imunologia , Antígenos de Neoplasias/metabolismo , Compostos Heterocíclicos com 1 Anel/química , Neoplasias Ovarianas/radioterapia , Radioimunoterapia/métodos , Animais , Anticorpos Monoclonais/química , Anticorpos Monoclonais/farmacocinética , Linhagem Celular Tumoral , Transformação Celular Neoplásica , Feminino , Humanos , Marcação por Isótopo , Camundongos , Terapia de Alvo Molecular , Neoplasias Ovarianas/imunologia , Neoplasias Ovarianas/metabolismo , Neoplasias Ovarianas/patologia , Distribuição TecidualRESUMO
Theranostics is a re-emerging field of medicine that aims to create targeted agents that can be used for diagnostic and/or therapeutic indications. In the past, theranostics has been used to treat neoplasms, such as thyroid cancer and neuroblastomas. More recently, theranostics has seen a resurgence with advent of new therapeutic antibodies and small molecules which can be transformed into Theranostic agents through radioconjugating with a radioactive isotope. Positron emitting radioisotopes can be used for diagnostic purposes while alpha- and beta-emitting radioisotopes can be used for therapy. The technique of radiolabeling an existing therapeutic agent (small molecule or antibody) leverages the existing qualities of that drug, and potentiates therapeutic effect by conjugating it with a cytotoxic-energy bearing radioisotope (e.g., 131-iodine, 177-lutetium). Theranostics have been used for a few decades now, starting with 131-iodine for therapy of autoimmune thyroiditis (Graves' disease, Hashimoto's thyroiditis) as well as for thyroid cancer. Additionally, 131-iodine-meta-iodobenzylguanidine (131-I-MIBG) initially had been used for gastroenteropancreatic neuroendocrine (carcinoid) tumors. However, recently clinical trials have start enrolling patients to evaluate efficacy of 131-I-MIBG in patients with small cell carcinoma of the lung. In the era of precision medicine and personalized targeted therapeutics, Theranostics can play a key pivotal in improving diagnostic and therapeutic specificity by increasing potency of these targeted small molecules and antibodies with radioisotopes. In this review, we will review various clinically relevant Theranostics agent and their utility in thoracic disorders, notably within oncology.
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INTRODUCTION: Ovarian cancer has only a 17% 5-year survival rate in patients diagnosed with late stage disease. Tumor-associated glycoprotein-72 (TAG72), expressed in 88% of all stages of ovarian cancer, is an excellent candidate for antibody-targeted therapy, as it is not expressed in normal human adult tissues, except in the secretory endometrium. METHODS: Using the clinically relevant anti-TAG72 murine monoclonal antibody CC49, we evaluated antibody drug conjugates (ADCs) incorporating the highly potent, synthetic antimitotic agent monomethylauristatin E (MMAE). MMAE was conjugated to CC49 via reduced disulfides in the hinge region, using three different types of linker chemistry, vinylsulfone (VS-MMAE), bromoacetamido (Br-MMAE), and maleimido (mal-MMAE). RESULTS: The drug antibody ratios (DARs) of the three ADCs were 2.3 for VS-MMAE, 10 for Br-MMAE, and 9.5 for mal-MMAE. All three ADCs exhibited excellent tumor to blood ratios on PET imaging, but the absolute uptake of CC49-mal-MMAE (3.3%ID/g) was low compared to CC49-Br-MMAE (6.43%ID/g), at 142 hours. Blood clearance at 43 hours was 38% for intact CC49, about 24% for both CC49-VS-MMAE and CC49-Br-MMAE, and 7% for CC49-mal-MMAE. CC49-VS-MMAE was not further studied due to its low DAR, while CC49-mal-MMAE was ineffective in the OVCAR3 xenograft likely due to its rapid blood clearance. In contrast, CC49-Br-MMAE treated mice exhibited an average of a 15.6 day tumor growth delay and a 40% increase in survival vs controls with four doses of 7.5 or 15 mg/kg of CC49-Br-MMAE. CONCLUSION: We conclude that CC49-Br-MMAE with a high DAR and stable linker performs well in a difficult to treat solid tumor model.
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Anticorpos Antineoplásicos/uso terapêutico , Antígenos de Neoplasias/imunologia , Imunoconjugados/uso terapêutico , Fatores Imunológicos/uso terapêutico , Oligopeptídeos/uso terapêutico , Neoplasias Ovarianas/terapia , Acetatos/química , Animais , Anticorpos Antineoplásicos/sangue , Anticorpos Antineoplásicos/metabolismo , Antígenos de Neoplasias/metabolismo , Linhagem Celular Tumoral , Feminino , Células HT29 , Humanos , Fatores Imunológicos/farmacocinética , Camundongos , Oligopeptídeos/sangue , Oligopeptídeos/química , Oligopeptídeos/metabolismo , Neoplasias Ovarianas/sangue , Neoplasias Ovarianas/diagnóstico por imagem , Neoplasias Ovarianas/metabolismo , Tomografia por Emissão de Pósitrons , Distribuição Aleatória , Sulfonas/químicaRESUMO
While anti-CEA antibodies have no direct effect on CEA-positive tumors, they can be used to direct potent anti-tumor effects as an antibody-IL-2 fusion protein (immunocytokine, ICK), and at the same time reduce the toxicity of IL-2 as a single agent. Using a fusion protein of humanized anti-CEA with human IL-2 (M5A-IL-2) in a transgenic murine model expressing human CEA, we show high tumor uptake of the ICK to CEA-positive tumors with additional lymph node targeting. ICK treated CEA-positive tumors exhibit significant tumor eradication. Analysis of tumor-infiltrating lymphocytes shows a high frequency of both CD8+ and CD4+ T cells along with CD11b positive myeloid cells in ICK treated mice. The frequency of tumor-infiltrating FoxP3+ CD4+ T cells (Tregs) is significantly reduced vs anti-CEA antibody-treated controls, indicating that ICK did not preferentially stimulate migration or proliferation of Tregs to the tumor. Combination therapy with anti-PD-1 antibody did not improve tumor reduction over ICK therapy alone. Since stereotactic tumor irradiation (SRT), commonly used in cancer therapy has immunomodulatory effects, we tested combination SRT+ICK therapy in two tumor model systems. Use of fractionated vs single high dose SRT in combination with ICK resulted in greater tumor inhibition and immunity to tumor rechallenge. In particular, tumor microenvironment and myeloid cell composition appear to play a significant role in the response rate to ICK+SRT combination therapy.
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Neoplasias , Radiocirurgia , Animais , Anticorpos Monoclonais , Interleucina-2 , Linfócitos do Interstício Tumoral , Camundongos , Microambiente TumoralRESUMO
[68Ga]DO3A-VS-Cys40-Exendin-4, a glucagon-like peptide 1 receptor agonist, was evaluated as a potential PET tracer for the quantitation of human islets transplanted to the liver. The short-lived PET radionuclide 68Ga, available on a regular basis from a 68Ge/68Ga generator, is an attractive choice. Human C-peptide was measured to evaluate human islet function post-transplantation and prior to microPET imaging. [68Ga]DO3A-VS-Cys40-Exendin-4 was radiosynthesized and evaluated for PET imaging of transplanted human islets in the liver of healthy NOD/SCID mice. The biodistribution of the tracer was evaluated to determine the uptake into various organs, and qPCR of liver samples was conducted to confirm engrafted islet numbers after PET imaging. Measurement of human C-peptide indicated that higher engrafted islet mass resulted in higher human C-peptide levels in post-transplantation. The microPET imaging yielded high resolution images of liver-engrafted islets and also showed significant retention in mouse livers at 8 weeks post-transplantation. Biodistribution studies in mice revealed that liver uptake of [68Ga]DO3A-VS-Cys40-Exendin-4 was approximately 6-fold higher in mice that received 1000 islet equivalent (IEQ) than in non-transplanted mice. qPCR analysis of insulin expression suggested that islet engraftment numbers were close to 1000 IEQ transplanted. In conclusion, human islets transplanted into the livers of mice exhibited significant uptake of [68Ga]DO3A-VS-Cys40-Exendin-4 compared to the livers of untreated mice; and imaging of the mice using PET showed the human islets clearly with high contrast against liver tissue, enabling accurate quantitation of islet mass. Further validation of [68Ga]DO3A-VS-Cys40-Exendin-4 as an islet imaging probe for future clinical application is ongoing.
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Exenatida/farmacocinética , Radioisótopos de Gálio/farmacocinética , Compostos Heterocíclicos com 1 Anel/farmacocinética , Ilhotas Pancreáticas/diagnóstico por imagem , Fígado/diagnóstico por imagem , Tomografia por Emissão de Pósitrons/métodos , Compostos de Vinila/farmacocinética , Animais , Humanos , Ilhotas Pancreáticas/metabolismo , Transplante das Ilhotas Pancreáticas , Camundongos , Camundongos Endogâmicos NOD , Camundongos SCID , Distribuição TecidualRESUMO
Real-time intraoperative image-guided cancer surgery promises to improve oncologic outcomes. Tumor-specific antibodies conjugated with near-infrared (NIR) fluorophores have demonstrated the potential to enhance visualization of solid tumor margins and metastatic disease; however, multiple challenges remain, including improvement in probe development for clinical utility. We have developed an NIR-IR800 dye on a PEGylated linker (sidewinder) conjugated to the humanized anti-carcinoembryonic antigen (CEA) antibody (M5A) with extended in vivo serum and tumor persistence. The anti-CEA M5A-sidewinder has a high dye-to-antibody ratio (average of 7 per antibody) that allows, in an orthotopic implanted human pancreatic cancer mouse model increased tumor fluorescence, higher tumor-to-background ratio and extends the surgical scheduling window compared to current antibody dye conjugates. These preclinical results demonstrate the potential of this probe for fluorescence-guided surgery of CEA-positive gastrointestinal cancers.
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Anticorpos Antineoplásicos/análise , Antígeno Carcinoembrionário/análise , Corantes Fluorescentes/administração & dosagem , Neoplasias Pancreáticas , Espectroscopia de Luz Próxima ao Infravermelho/métodos , Cirurgia Assistida por Computador/métodos , Animais , Modelos Animais de Doenças , Camundongos , Camundongos Nus , Neoplasias Pancreáticas/diagnóstico por imagem , Neoplasias Pancreáticas/cirurgiaRESUMO
Molecular imaging is becoming essential for precision targeted radiation therapy, yet progress is hindered from a lack of integrated imaging and treatment systems. We report the development of a prototype positron emission tomography (PET) scanner integrated into a commercial cone beam computed tomography (CBCT) based small animal irradiation system for molecular-image-guided, targeted external beam radiation therapy. The PET component consists of two rotating Hamamatsu time-of-flight PET modules positioned with a bore diameter of 101.6 mm and a radial field-of-view of 53.1 mm. The measured energy resolution after linearity correction at 511 KeV was 12.9% and the timing resolution was 283.6 ps. The measured spatial resolutions at the field-of-view center and 5 mm off the radial center were 2.6 mm × 2.6 mm × 1.6 mm and 2.6 mm × 2.6 mm × 2.7 mm respectively. 18F-Fluorodeoxyglucose-based PET imaging of a NEMA NU 4-2008 phantom resolved cylindrical volumes with diameters as small as 3 mm. To validate the system in-vivo, we performed 64Cu-DOTA-M5A PET and computed tomography (CT) imaging of carcinoembryonic antigen (CEA)-positive colorectal cancer in athymic nude mice and compared the results with a commercially available Siemens Inveon PET/CT system. The prototype PET system performed comparably to the Siemens system for identifying the location, size, and shape of tumors. Regions of heterogeneous 64Cu-DOTA-M5A uptake were observed. Using 64Cu-DOTA-M5A PET and CT images, a Monte Carlo-based radiation treatment plan was created to escalate the dose to the 64Cu-DOTA-M5A-based, highly active, biological target volume while largely sparing the normal tissue. Results demonstrate the feasibility of molecular-image-guided treatment plans using the prototype theranostic system.
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PURPOSE: Acute myeloid leukemia (AML) is a highly aggressive form of leukemia, which results in poor survival outcomes. Currently, diagnosis and prognosis are based on invasive single-point bone marrow biopsies (iliac crest). There is currently no AML-specific noninvasive imaging method to detect disease, including in extramedullary organs, representing an unmet clinical need. About 85% to 90% of human myeloid leukemia cells express CD33 cell surface receptors, highlighting CD33 as an ideal candidate for AML immunoPET. EXPERIMENTAL DESIGN: We evaluated whether [64Cu]Cu-DOTA-anti-CD33 murine mAb can be used for immunoPET imaging of AML in a preclinical model. MicroCT was adjusted to detect spatial/anatomical details of PET activity. For translational purposes, a humanized anti-CD33 antibody was produced; we confirmed its ability to detect disease and its distribution. We reconfirmed/validated CD33 antibody-specific targeting with an antibody-drug conjugate (ADC) and radioimmunotherapy (RIT). RESULTS: [64Cu]Cu-DOTA-anti-CD33-based PET-CT imaging detected CD33+ AML in mice with high sensitivity (95.65%) and specificity (100%). The CD33+ PET activity was significantly higher in specific skeletal niches [femur (P < 0.00001), tibia (P = 0.0001), humerus (P = 0.0014), and lumber spine (P < 0.00001)] in AML-bearing mice (over nonleukemic control mice). Interestingly, the hybrid PET-CT imaging showed high disease activity in the epiphysis/metaphysis of the femur, indicating regional spatial heterogeneity. Anti-CD33 therapy using newly developed humanized anti-CD33 mAb as an ADC (P = 0.02) and [225Ac]Ac-anti-CD33-RIT (P < 0.00001) significantly reduced disease burden over that of respective controls. CONCLUSIONS: We have successfully developed a novel anti-CD33 immunoPET-CT-based noninvasive modality for AML and its spatial distribution, indicating a preferential skeletal niche.
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Radioisótopos de Cobre/química , Compostos Heterocíclicos com 1 Anel/química , Imunoconjugados/farmacocinética , Leucemia Mieloide Aguda/diagnóstico por imagem , Tomografia por Emissão de Pósitrons combinada à Tomografia Computadorizada/métodos , Compostos Radiofarmacêuticos/farmacocinética , Lectina 3 Semelhante a Ig de Ligação ao Ácido Siálico/imunologia , Animais , Anticorpos Monoclonais Humanizados/farmacocinética , Linhagem Celular Tumoral , Modelos Animais de Doenças , Leucemia Mieloide Aguda/tratamento farmacológico , Leucemia Mieloide Aguda/patologia , Camundongos , Camundongos Endogâmicos NOD , Camundongos SCID , Lectina 3 Semelhante a Ig de Ligação ao Ácido Siálico/antagonistas & inibidores , Distribuição Tecidual , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
Chimeric antigen receptor (CAR) T cell therapy is a promising clinical approach for reducing tumor progression and prolonging patient survival. However, improvements in both the safety and the potency of CAR T cell therapy demand quantitative imaging techniques to determine the distribution of cells after adoptive transfer. The purpose of this study was to optimize 89Zr-oxine labeling of CAR T cells and evaluate PET as a platform for imaging adoptively transferred CAR T cells. Methods: CAR T cells were labeled with 0-1.4 MBq of 89Zr-oxine per 106 cells and assessed for radioactivity retention, viability, and functionality. In vivo trafficking of 89Zr-oxine-labeled CAR T cells was evaluated in 2 murine xenograft tumor models: glioblastoma brain tumors with intracranially delivered IL13Rα2-targeted CAR T cells, and subcutaneous prostate tumors with intravenously delivered prostate stem cell antigen (PSCA)-targeted CAR T cells. Results: CAR T cells were efficiently labeled (75%) and retained more than 60% of the 89Zr over 6 d. In vitro cytokine production, migration, and tumor cytotoxicity, as well as in vivo antitumor activity, were not significantly reduced when labeled with 70 kBq/106 cells. IL13Rα2-CAR T cells delivered intraventricularly were detectable by PET for at least 6 d throughout the central nervous system and within intracranial tumors. When intravenously administered, PSCA-CAR T cells also showed tumor tropism, with a 9-fold greater tumor-to-muscle ratio than for CAR-negative T cells. Conclusion:89Zr-oxine can be used for labeling and imaging CAR T cells while maintaining cell viability and function. On the basis of these studies, we conclude that 89Zr-oxine is a clinically translatable platform for real-time assessment of cell therapies.
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Imunoterapia Adotiva , Oxiquinolina/metabolismo , Radioisótopos , Linfócitos T/imunologia , Zircônio , Animais , Linhagem Celular Tumoral , Humanos , Marcação por Isótopo , Masculino , Camundongos , Oxiquinolina/farmacocinética , Linfócitos T/metabolismo , Distribuição TecidualRESUMO
The development of improved breast cancer screening methods is hindered by a lack of cancer-specific imaging agents and effective small-animal models to test them. The purpose of this study was to evaluate 64Cu-DOTA-alendronate as a mammary microcalcification-targeting PET imaging agent, using an ideal rat model. Our long-term goal is to develop 64Cu-DOTA-alendronate for the detection and noninvasive differentiation of malignant versus benign breast tumors with PET. Methods: DOTA-alendronate was synthesized, radiolabeled with 64Cu, and administered to normal or tumor-bearing aged, female, retired breeder Sprague-Dawley rats for PET imaging. Mammary tissues were subsequently labeled and imaged with light, confocal, and electron microscopy to verify microcalcification targeting specificity of DOTA-alendronate and elucidate the histologic and ultrastructural characteristics of the microcalcifications in different mammary tumor types. Tumor uptake, biodistribution, and dosimetry studies were performed to evaluate the efficacy and safety of 64Cu-DOTA-alendronate. Results:64Cu-DOTA-alendronate was radiolabeled with a 98% yield. PET imaging using aged, female, retired breeder rats showed specific binding of 64Cu-DOTA-alendronate in mammary glands and mammary tumors. The highest uptake of 64Cu-DOTA-alendronate was in malignant tumors and the lowest uptake in benign tumors and normal mammary tissue. Confocal analysis with carboxyfluorescein-alendronate confirmed the microcalcification binding specificity of alendronate derivatives. Biodistribution studies revealed tissue alendronate concentrations peaking within the first hour, then decreasing over the next 48 h. Our dosimetric analysis demonstrated a 64Cu effective dose within the acceptable range for clinical PET imaging agents and the potential for translation into human patients. Conclusion:64Cu-DOTA-alendronate is a promising PET imaging agent for the sensitive and specific detection of mammary tumors as well as the differentiation of malignant versus benign tumors based on absolute labeling uptake.
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Alendronato/química , Calcinose/diagnóstico por imagem , Radioisótopos de Cobre , Compostos Heterocíclicos com 1 Anel/química , Neoplasias Mamárias Experimentais/diagnóstico por imagem , Tomografia por Emissão de Pósitrons , Animais , Calcinose/metabolismo , Linhagem Celular Tumoral , Modelos Animais de Doenças , Feminino , Neoplasias Mamárias Experimentais/metabolismo , Ratos , Ratos Sprague-Dawley , Distribuição TecidualRESUMO
INTRODUCTION: Single chain (scFv) antibodies are ideal targeting ligands due to their modular structure, high antigen specificity and affinity. These monovalent ligands display rapid tumor targeting but have limitations due to their fast urinary clearance. METHODS: An anti-prostate membrane antigen (PSMA) scFv with a site-specific cysteine was expressed and evaluated in a prostate cancer xenograft model by Cu-64 PET imaging. To enhance tumor accumulation, the scFv-cys was conjugated to the co-polymer DSPE-PEG-maleimide that spontaneously assembled into a homogeneous multivalent lipid nanoparticle (LNP). RESULTS: The targeted LNP exhibited a 2-fold increase in tumor uptake compared to the scFv alone using two different thiol ester chemistries. The anti-PSMA scFv-LNP exhibited a 1.6 fold increase in tumor targeting over the untargeted LNP. CONCLUSIONS: The targeted anti-PSMA scFv-LNP showed enhanced tumor accumulation over the scFv alone or the untargeted DOTA-micelle providing evidence for the development of this system for drug delivery. ADVANCES IN KNOWLEDGE AND IMPLICATIONS FOR PATIENT CARE: Anti-tumor scFv antibody fragments have not achieved their therapeutic potential due to their fast blood clearance. Conjugation to an LNP enables multivalency to the tumor antigen as well as increased molecular size for chemotherapy drug delivery.
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Antígenos de Superfície/imunologia , Radioisótopos de Cobre , Glutamato Carboxipeptidase II/imunologia , Compostos Heterocíclicos com 1 Anel/química , Nanopartículas/química , Tomografia por Emissão de Pósitrons , Neoplasias da Próstata/diagnóstico por imagem , Anticorpos de Cadeia Única/química , Animais , Linhagem Celular Tumoral , Transformação Celular Neoplásica , Portadores de Fármacos/química , Humanos , Masculino , Camundongos , Neoplasias da Próstata/patologia , Anticorpos de Cadeia Única/imunologiaRESUMO
The blood clearance of chemotherapeutic drugs such as doxorubicin (Dox) can be extended by incorporation into lipid nanoparticles (LNPs) and further improved by tumor targeting with antibody fragments. We used positron emission tomography (PET) imaging in a murine prostate cancer model to evaluate tumor targeting of LNPs incorporating Dox and antiprostate-specific membrane antigen (PSMA) diabodies. Dox-LNPs were generated by mixing or covalent attachment to water soluble distearoylphosphatidyl ethanolamine-polyethylene glycol (DSPE-PEG)2000. Cu-64 PET imaging was performed with DOTA-conjugated Dox, PEG-LNP, or an anti-PSMA site-specific cysteine-diabody. Since the mixture Dox+PEG-LNP was unstable in serum, further studies utilized Dox covalently bound to LNP ± covalently bound DOTA-cys-diabody (cys-DB)-LNP. Blood clearance of covalent Dox-PEG-LNP was slower than Dox alone or Dox+PEG-LNP. PET imaging of 64Cu-DOTA-Dox-PEG-LNP reached a maximum of 10% ID/g in tumors compared with 3% ID/g of 64Cu-DOTA-Dox, due to the prolonged blood clearance. Mixing 64Cu-DOTA-cys-DB-PEG-LNP with covalent Dox-PEG-LNP gave LNPs containing both drug and tumor targeting cys-DB. The mixed LNPs exhibited increased tumor uptake (15% ID/g) versus untargeted 64Cu-DOTA-Dox-PEG-LNPs (10% ID/g) demonstrating feasibility of the approach. Based on these results, a therapy study with mixed LNPs containing cys-DB-LNP and either Dox-LNP or the antitubulin drug auristatin-LNP showed significant reduction of tumor growth with the auristatin-diabody-LNP mixture, but not the Dox-diabody-LNP mixture.
Assuntos
Lipídeos/administração & dosagem , Lipídeos/química , Nanopartículas/administração & dosagem , Nanopartículas/química , Neoplasias da Próstata/tratamento farmacológico , Aminobenzoatos/química , Aminobenzoatos/farmacologia , Animais , Antígenos de Superfície/química , Linhagem Celular Tumoral , Radioisótopos de Cobre/química , Modelos Animais de Doenças , Doxorrubicina/química , Doxorrubicina/farmacologia , Glutamato Carboxipeptidase II/química , Humanos , Masculino , Camundongos , Camundongos Endogâmicos NOD , Camundongos SCID , Transplante de Neoplasias/métodos , Oligopeptídeos/química , Oligopeptídeos/farmacologia , Fosfatidiletanolaminas/química , Polietilenoglicóis/química , Tomografia por Emissão de Pósitrons/métodos , Próstata/efeitos dos fármacos , Compostos Radiofarmacêuticos/química , Distribuição TecidualRESUMO
A series of anti-tumor/anti-chelate bispecific antibody formats were developed for pre-targeted radioimmunotherapy. Based on the anti-carcinoembryonic antigen humanized hT84.66-M5A monoclonal antibody and the anti-DOTA C8.2.5 scFv antibody fragment, this cognate series of bispecific antibodies were radioiodinated to determine their tumor targeting, biodistribution and pharmacokinetic properties in a mouse xenograft tumor model. The in vivo biodistribution studies showed that all the bispecific antibodies exhibited specific high tumor uptake but the tumor targeting was approximately one-half of the parental anti-CEA mAb due to faster blood clearance. Serum stability and FcRn studies showed no apparent reason for the faster blood clearance. A dual radiolabel biodistribution study revealed that the (111)In-DOTA bispecific antibody had increased liver and spleen uptake, not seen for the (125)I-version due to metabolism and release of the radioiodine from the cells. These data suggest increased clearance of the antibody fusion formats by the mononuclear phagocyte system. Importantly, a pre-targeted study showed specific tumor uptake of (177)Lu-DOTA and a tumor : blood ratio of 199 : 1. This pre-targeted radiotherapeutic and substantial reduction in the radioactive exposure to the bone marrow should enhance the therapeutic potential of RIT.