RESUMEN
Vascular contributions to dementia and Alzheimer's disease are increasingly recognized1-6. Recent studies have suggested that breakdown of the blood-brain barrier (BBB) is an early biomarker of human cognitive dysfunction7, including the early clinical stages of Alzheimer's disease5,8-10. The E4 variant of apolipoprotein E (APOE4), the main susceptibility gene for Alzheimer's disease11-14, leads to accelerated breakdown of the BBB and degeneration of brain capillary pericytes15-19, which maintain BBB integrity20-22. It is unclear, however, whether the cerebrovascular effects of APOE4 contribute to cognitive impairment. Here we show that individuals bearing APOE4 (with the ε3/ε4 or ε4/ε4 alleles) are distinguished from those without APOE4 (ε3/ε3) by breakdown of the BBB in the hippocampus and medial temporal lobe. This finding is apparent in cognitively unimpaired APOE4 carriers and more severe in those with cognitive impairment, but is not related to amyloid-ß or tau pathology measured in cerebrospinal fluid or by positron emission tomography23. High baseline levels of the BBB pericyte injury biomarker soluble PDGFRß7,8 in the cerebrospinal fluid predicted future cognitive decline in APOE4 carriers but not in non-carriers, even after controlling for amyloid-ß and tau status, and were correlated with increased activity of the BBB-degrading cyclophilin A-matrix metalloproteinase-9 pathway19 in cerebrospinal fluid. Our findings suggest that breakdown of the BBB contributes to APOE4-associated cognitive decline independently of Alzheimer's disease pathology, and might be a therapeutic target in APOE4 carriers.
Asunto(s)
Apolipoproteína E4/genética , Barrera Hematoencefálica/patología , Disfunción Cognitiva/genética , Disfunción Cognitiva/patología , Alelos , Enfermedad de Alzheimer/genética , Enfermedad de Alzheimer/patología , Péptidos beta-Amiloides/líquido cefalorraquídeo , Péptidos beta-Amiloides/metabolismo , Capilares/patología , Ciclofilina A/líquido cefalorraquídeo , Ciclofilina A/metabolismo , Femenino , Heterocigoto , Hipocampo/irrigación sanguínea , Humanos , Masculino , Metaloproteinasa 9 de la Matriz/líquido cefalorraquídeo , Metaloproteinasa 9 de la Matriz/metabolismo , Giro Parahipocampal/irrigación sanguínea , Pericitos/patología , Tomografía de Emisión de Positrones , Receptor beta de Factor de Crecimiento Derivado de Plaquetas/líquido cefalorraquídeo , Lóbulo Temporal/irrigación sanguínea , Proteínas tau/líquido cefalorraquídeo , Proteínas tau/metabolismoRESUMEN
Tau pathology and vascular dysfunction are important contributors to Alzheimer's disease (AD), but vascular-tau associations and their effects on cognition are poorly understood. We investigated these associations in male and female humans by conducting voxelwise comparisons between cerebral blood flow (CBF) and tau positron emission tomography (PET) images in independent discovery [cognitively normal (CN), 19; mild cognitive impairment (MCI) risk, 43; MCI, 6] and replication (CN,73; MCI, 45; AD, 20) cohorts. In a subgroup, we assessed relationships between tau and soluble platelet-derived growth factor ß (sPDGFRß), a CSF marker of pericyte injury. We tested whether CBF/sPDGFRß-tau relationships differed based on Montreal Cognitive Assessment (MoCA) global cognition performance, or based on amyloid burden. Mediation analyses assessed relationships among CBF/sPDGFRß, tau, and cognition. Negative CBF-tau correlations were observed predominantly in temporal-parietal regions. In the replication cohort, early negative CBF-tau correlations increased in spatial extent and in strength of correlation with increased disease severity. Stronger CBF-tau and sPDGFRß-tau correlations were observed in participants with greater amyloid burden and lower MoCA scores. Importantly, when stratifying by amyloid status, stronger CBF-tau relationships in individuals with lower MoCA scores were driven by amyloid+ participants. Tau PET was a significant mediator CBF/sPDGFRß-MoCA relationships in numerous regions. Our results demonstrate vascular-tau associations across the AD spectrum and suggest that early vascular-tau associations are exacerbated in the presence of amyloid, consistent with a two-hit model of AD on cognition. Combination treatments targeting vascular health, as well as amyloid-ß and tau levels, may preserve cognitive function more effectively than single-target therapies.SIGNIFICANCE STATEMENT Emerging evidence demonstrates a role for vascular dysfunction as a significant contributor to Alzheimer's pathophysiology. However, associations between vascular dysfunction and tau pathology, and their effects on cognition remain poorly understood. Multimodal neuroimaging data from two independent cohorts were analyzed to provide novel in vivo evidence of associations between cerebral blood flow (CBF), an MRI measure of vascular health, and tau pathology using PET. CBF-tau associations were related to cognition and driven in part by amyloid burden. Soluble platelet-derived growth factor ß, an independent CSF vascular biomarker, confirmed vascular-tau associations in a subgroup analysis. These results suggest that combination treatments targeting vascular health, amyloid-ß, and tau levels may more effectively preserve cognitive function than single-target therapies.
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Amiloide/metabolismo , Vasos Sanguíneos/diagnóstico por imagen , Cognición , Proteínas tau/metabolismo , Anciano , Enfermedad de Alzheimer/metabolismo , Circulación Cerebrovascular , Disfunción Cognitiva/diagnóstico por imagen , Disfunción Cognitiva/fisiopatología , Estudios de Cohortes , Femenino , Humanos , Imagen por Resonancia Magnética , Masculino , Pruebas de Estado Mental y Demencia , Persona de Mediana Edad , Tomografía de Emisión de Positrones , Receptor beta de Factor de Crecimiento Derivado de Plaquetas/líquido cefalorraquídeoRESUMEN
This review explores recent work directed toward the development of nanoparticles (NPs) for multimodality cancer imaging and targeted cancer therapy. In the growing era of precision medicine, theranostics, or the combined use of targeted molecular probes in diagnosing and treating diseases is playing a particularly powerful role. There is a growing interest, particularly over the past few decades, in the use of NPs as theranostic tools due to their excellent performance in receptor target specificity and reduction in off-target effects when used as therapeutic agents. This review discusses recent advances, as well as the advantages and challenges of the application of NPs in cancer imaging and therapy.
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Imagen Multimodal/métodos , Nanopartículas/química , Neoplasias/diagnóstico por imagen , Neoplasias/terapia , Animales , Diagnóstico por Imagen/métodos , Humanos , Imagen por Resonancia Magnética/métodos , Nanopartículas del Metal/química , Ratones , Imagen Molecular/métodos , Trasplante de Neoplasias , Seguridad del Paciente , Fototerapia/métodos , Tomografía de Emisión de Positrones/métodos , Medicina de Precisión/métodos , Radioterapia/métodos , Solubilidad , Nanomedicina TeranósticaRESUMEN
Heat shock protein 90 (Hsp90) plays a vital role in the progress of malignant disease and elevated Hsp90 expression has been reported in pancreatic cancer. In this study, we radiolabeled a dimeric Sansalvamide A derivative (Di-San A1) with 64Cu, and evaluated the feasibility of using 64Cu-Di-San A1 for PET imaging of Hsp90 expression in a mouse model of pancreatic cancer. A macrocyclic chelator NOTA (1,4,7-triazacyclononane-1,4,7-trisacetic acid) was conjugated to Di-San A1. 64Cu-Di-San A1 was successfully prepared in a radiochemical yield > 97% with a radiochemical purity > 98%. 64Cu-Di-San A1 is stable in PBS and mouse serum with > 92% of parent probe intact after 4 h incubation. The cell binding and uptake revealed that 64Cu-Di-San A1 binds to Hsp90-positive PL45 pancreatic cancer cells, and the binding can be effectively blocked by an Hsp90 inhibitor (17AAG). For microPET study, 64Cu-Di-San A1 shows good in vivo performance in terms of tumor uptake in nude mice bearing PL45 tumors. The Hsp90-specific tumor activity accumulation of 64Cu-Di-San A1 was further demonstrated by significant reduction of PL45 tumor uptake with a pre-injected blocking dose of 17AAG. The ex vivo PET imaging and biodistribution results were consistent with the quantitative analysis of PET imaging, demonstrating good tumor-to-muscle ratio (5.35 ± 0.46) of 64Cu-Di-San A1 at 4 h post-injection in PL45 tumor mouse xenografts. 64Cu-Di-San A1 allows PET imaging of Hsp90 expression in PL45 tumors, which may provide a non-invasive method to quantitatively characterize Hsp90 expression in pancreatic cancer.
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Cobre/farmacología , Depsipéptidos/farmacología , Regulación Neoplásica de la Expresión Génica , Proteínas HSP90 de Choque Térmico/biosíntesis , Proteínas de Neoplasias/biosíntesis , Neoplasias Pancreáticas , Tomografía de Emisión de Positrones , Animales , Línea Celular Tumoral , Cobre/química , Depsipéptidos/química , Femenino , Xenoinjertos , Humanos , Ratones , Ratones Desnudos , Trasplante de Neoplasias , Neoplasias Pancreáticas/diagnóstico por imagen , Neoplasias Pancreáticas/metabolismo , Multimerización de ProteínaRESUMEN
Heparin anticoagulation therapy is an indispensable feature of clinical care yet has a narrow therapeutic window and is the second most common intensive care unit (ICU) medication error. The active partial thromboplastin time (aPTT) monitors heparin but suffers from long turnaround times, a variable reference range, limited utility with low molecular weight heparin, and poor correlation to dose. Here, we describe a photoacoustic imaging technique to monitor heparin concentration using methylene blue as a simple and Federal Drug Administration-approved contrast agent. We found a strong correlation between heparin concentration and photoacoustic signal measured in phosphate buffered saline (PBS) and blood. Clinically relevant heparin concentrations were detected in blood in 32 s with a detection limit of 0.28 U/mL. We validated this imaging approach by correlation to the aPTT (Pearson's r = 0.86; p < 0.05) as well as with protamine sulfate treatment. This technique also has good utility with low molecular weight heparin (enoxaparin) including a blood detection limit of 72 µg/mL. We then used these findings to create a nanoparticle-based hybrid material that can immobilize methylene blue for potential applications as a wearable/implantable heparin sensor to maintain drug levels in the therapeutic window. To the best of our knowledge, this is the first use of photoacoustics to image anticoagulation therapy with significant potential implications to the cardiovascular and surgical community.
RESUMEN
Fibroblast activation protein (FAP) is highly expressed in the tumor-associated fibroblasts (TAFs) of most human epithelial cancers. FAP plays a critical role in tumorigenesis and cancer progression, which makes it a promising target for novel anticancer therapy. However, mere abrogation of FAP enzymatic activity by small molecules is not very effective in inhibiting tumor growth. In this study, we have evaluated a novel immune-based approach to specifically deplete FAP-expressing TAFs in a mouse 4T1 metastatic breast cancer model. Depletion of FAP-positive stromal cells by FAP-targeting immunotoxin αFAP-PE38 altered levels of various growth factors, cytokines, chemokines and matrix metalloproteinases, decreased the recruitment of tumor-infiltrating immune cells in the tumor microenvironment and suppressed tumor growth. In addition, combined treatment with αFAP-PE38 and paclitaxel potently inhibited tumor growth in vivo. Our findings highlight the potential use of immunotoxin αFAP-PE38 to deplete FAP-expressing TAFs and thus provide a rationale for the use of this immunotoxin in cancer therapy.
Asunto(s)
Antineoplásicos/farmacología , Neoplasias de la Mama/patología , Fibroblastos/metabolismo , Gelatinasas/antagonistas & inhibidores , Inmunotoxinas/farmacología , Proteínas de la Membrana/antagonistas & inhibidores , Animales , Antineoplásicos/farmacocinética , Células 3T3 BALB , Modelos Animales de Enfermedad , Endopeptidasas , Femenino , Citometría de Flujo , Técnica del Anticuerpo Fluorescente , Humanos , Inmunohistoquímica , Inmunotoxinas/farmacocinética , Ratones , Reacción en Cadena en Tiempo Real de la Polimerasa , Serina EndopeptidasasRESUMEN
The fast kinetics and bioorthogonal nature of the tetrazine trans-cyclooctene (TCO) ligation makes it a unique tool for PET probe construction. In this study, we report the development of an (18)F-labeling system based on a CF3-substituted diphenyl-s-tetrazine derivative with the aim of maintaining high reactivity while increasing in vivo stability. c(RGDyK) was tagged by a CF3-substituted diphenyl-s-tetrazine derivative via EDC-mediated coupling. The resulting tetrazine-RGD conjugate was combined with a (19)F-labeled TCO derivative to give HPLC standards. The analogous (18)F-labeled TCO derivative was combined with the diphenyl-s-tetrazine-RGD at µM concentration. The resulting tracer was subjected to in vivo metabolic stability assessment, and microPET studies in murine U87MG xenograft models. The diphenyl-s-tetrazine-RGD combines with an (18)F-labeled TCO in high yields (>97% decay-corrected on the basis of TCO) using only 4 equiv of tetrazine-RGD relative to the (18)F-labeled TCO (concentration calculated based on product's specific activity). The radiochemical purity of the (18)F-RGD peptides was >95% and the specific activity was 111 GBq/µmol. Noninvasive microPET experiments demonstrated that (18)F-RGD had integrin-specific tumor uptake in subcutaneous U87MG glioma. In vivo metabolic stability of (18)F-RGD in blood, urine, and major organs showed two major peaks: one corresponded to the Diels-Alder conjugate and the other was identified as the aromatized analog. A CF3-substituted diphenyl-s-tetrazine displays excellent speed and efficiency in (18)F-PET probe construction, providing nearly quantitative (18)F labeling within minutes at low micromolar concentrations. The resulting conjugates display improved in vivo metabolic stability relative to our previously described system.
Asunto(s)
Ciclooctanos/metabolismo , Radioisótopos de Flúor/metabolismo , Compuestos Heterocíclicos con 1 Anillo/metabolismo , Tomografía de Emisión de Positrones , Animales , Línea Celular Tumoral , Ciclooctanos/química , Femenino , Radioisótopos de Flúor/química , Compuestos Heterocíclicos con 1 Anillo/química , Humanos , Ratones , Ratones Desnudos , Tomografía de Emisión de Positrones/métodos , Ensayos Antitumor por Modelo de Xenoinjerto/métodosRESUMEN
In this study, we synthesized a novel Cy5.5-labeled dimeric NGR peptide (Cy5.5-NGR2) via bioorthogonal click chemistry, and evaluated the utility of Cy5.5-NGR2 for near-infrared fluorescence imaging of CD13 receptor expression in vivo. The dimeric NGR peptide (NGR2) was conjugated with an alkyne-containing PEG unit followed by mixing with an azide-terminated Cy5.5 fluorophore (Cy5.5-N3) to afford Cy5.5-NGR2. The probe was subject to in vitro and in vivo evaluations. The bioorthogonal click chemistry provided a rapid conjugation of the alkyne-containing NGR2 with Cy5.5-N3 in a quantitative yield within 15 min. The laser confocal microscopy revealed that binding of Cy5.5-NGR2 to CD13 receptor is target-specific as demonstrated in CD13-positive HT-1080 cells, CD13-negative MCF-7 cells, and a blocking study in HT-1080 cells. For in vivo optical imaging, Cy5.5-NGR2 exhibited rapid HT-1080 tumor targeting at 0.5 h postinjection (pi), and highest tumor-to-background contrast at 2 h pi. The CD13-specific tumor accumulation of Cy5.5-NGR2 was accomplished by a blocking study with unlabeled NGR peptide in HT-1080 tumor bearing mice. The tumor-to-muscle ratio of Cy5.5-NGR2 at 2 h pi reached 2.65 ± 0.13 in the non-blocking group vs. 1.05 ± 0.06 in the blocking group. The results from ex vivo imaging were consistent with the in vivo findings. We concluded that Cy5.5-NGR2 constructed by bioorthogonal click chemistry is a promising molecular probe, not only allowing the NIR optical imaging of CD13 overexpressed tumors, but also having the potential to facilitate noninvasive monitoring of CD13-targeted tumor therapy.
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Carbocianinas , Colorantes Fluorescentes , Oligopéptidos , Animales , Línea Celular Tumoral , Humanos , Células MCF-7 , Ratones , Microscopía Confocal , Espectroscopía Infrarroja Corta/métodosRESUMEN
Pathological angiogenesis is crucial in tumor growth, invasion and metastasis. Previous studies demonstrated that the vascular endothelial growth inhibitor (VEGI), a member of the tumor necrosis factor superfamily, can be used as a potent endogenous inhibitor of tumor angiogenesis. Molecular probes containing the asparagine-glycine-arginine (NGR) sequence can specifically bind to CD13 receptor which is overexpressed on neovasculature and several tumor cells. Near-infrared fluorescence (NIRF) optical imaging for targeting tumor vasculature offers a noninvasive method for early detection of tumor angiogenesis and efficient monitoring of response to anti-tumor vasculature therapy. The aim of this study was to develop a new NIRF imaging probe on the basis of an NGR-VEGI protein for the visualization of tumor vasculature. The NGR-VEGI fusion protein was prepared from prokaryotic expression, and its function was characterized in vitro. The NGR-VEGI protein was then labeled with a Cy5.5 fluorophore to afford Cy5.5-NGR-VEGI probe. Using the NIRF imaging technique, we visualized and quantified the specific delivery of Cy5.5-NGR-VEGI protein to subcutaneous HT-1080 fibrosarcoma tumors in mouse xenografts. The Cy5.5-NGR-VEGI probe exhibited rapid HT-1080 tumor targeting, and highest tumor-to-background contrast at 8 h post-injection (pi). Tumor specificity of Cy5.5-NGR-VEGI was confirmed by effective blocking of tumor uptake in the presence of unlabeled NGR-VEGI (20 mg/kg). Ex vivo NIRF imaging further confirmed in vivo imaging findings, demonstrating that Cy5.5-NGR-VEGI displayed an excellent tumor-to-muscle ratio (18.93 ± 2.88) at 8 h pi for the non-blocking group and significantly reduced ratio (4.92 ± 0.75) for the blocking group. In conclusion, Cy5.5-NGR-VEGI provided highly sensitive, target-specific, and longitudinal imaging of HT-1080 tumors. As a novel theranostic protein, Cy5.5-NGR-VEGI has the potential to improve cancer treatment by targeting tumor vasculature.
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Sistemas de Liberación de Medicamentos/métodos , Neoplasias/tratamiento farmacológico , Neovascularización Patológica/tratamiento farmacológico , Oligopéptidos/administración & dosificación , Miembro 15 de la Superfamilia de Ligandos de Factores de Necrosis Tumoral/administración & dosificación , Animales , Apoptosis , Carbocianinas/química , Sistemas de Liberación de Medicamentos/instrumentación , Femenino , Colorantes Fluorescentes/química , Humanos , Ratones , Ratones Desnudos , Imagen Molecular , Sondas Moleculares/química , Neoplasias/química , Neoplasias/patología , Neoplasias/fisiopatología , Neovascularización Patológica/patología , Neovascularización Patológica/fisiopatología , Oligopéptidos/química , Oligopéptidos/genética , Oligopéptidos/metabolismo , Espectroscopía Infrarroja Corta , Miembro 15 de la Superfamilia de Ligandos de Factores de Necrosis Tumoral/química , Miembro 15 de la Superfamilia de Ligandos de Factores de Necrosis Tumoral/genética , Miembro 15 de la Superfamilia de Ligandos de Factores de Necrosis Tumoral/metabolismoRESUMEN
CD13 receptor as a tumor vasculature biomarker has attracted great attention in cancer research. Through phage display screening, NGR-containing peptides have been characterized as specific ligands binding to CD13 receptor. In this study, a (64)Cu-labeled dimeric NGR peptide based on sarcophagine cage was synthesized and evaluated for micropositron emission tomography (PET) imaging of CD13 expression in vivo. Macrocyclic chelating agent (sarcophagine cage) was conjugated with two azide moieties, followed by mixing with an alkyne-containing NGR peptide to rapidly provide the Sar-NGR2 peptide by click chemistry. Radiolabeling of Sar-NGR2 with (64)Cu was achieved in >90% decay-corrected yield with radiochemical purity of >99%. The cell uptake study showed that (64)Cu-Sar-NGR2 binds to CD13-positive HT-1080 cells, but not to CD13-negative MCF-7 cells. MicroPET imaging results revealed that (64)Cu-Sar-NGR2 exhibits good tumor uptake in CD13-positive HT-1080 xenografts and significantly lower tumor uptake in CD13-negative MCF-7 xenografts. The CD13-specific binding of (64)Cu-Sar-NGR2 was further verified by significant reduction of tumor uptake in HT-1080 tumor xenografts with coinjection of a nonradiolabeled NGR peptide. The biodistribution results demonstrated good tumor/muscle ratio (8.28 ± 0.37) of (64)Cu-Sar-NGR2 at 24 h pi in HT-1080 tumor xenografts, which is in agreement with the quantitative analysis of microPET imaging. In conclusion, sarcophagine cage has been successfully applied to the construction of a (64)Cu-labeled dimeric NGR-containing peptide. In vitro and in vivo studies demonstrated that (64)Cu-Sar-NGR2 is a promising PET probe for imaging CD13 expression in living mice.
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4-Butirolactona/análogos & derivados , Antígenos CD13/química , Radioisótopos de Cobre/química , Oligopéptidos/química , Tomografía de Emisión de Positrones , 4-Butirolactona/química , Animales , Catálisis , Línea Celular Tumoral , Cromatografía Líquida de Alta Presión , Química Clic , Dimerización , Femenino , Regulación de la Expresión Génica , Humanos , Hígado/diagnóstico por imagen , Células MCF-7 , Ratones , Ratones Desnudos , Trasplante de Neoplasias , Neovascularización Patológica , Péptidos/química , Distribución TisularRESUMEN
Accumulating experimental evidence indicates that overexpression of the oncogenic receptor tyrosine kinase, Axl, plays a key role in the tumorigenesis and metastasis of various types of cancer. The objective of this study is to design a novel imaging probe based on the monoclonal antibody, h173, for microPET imaging of Axl expression in human lung cancer. A bifunctional chelator, DOTA, was conjugated to h173, followed by radiolabeling with (64)Cu. The binding of DOTA-h173 to the Axl receptor was first evaluated by a cell uptake assay and flow cytometry analysis using human lung cancer cell lines. The probe (64)Cu-DOTA-h173 was further evaluated by microPET imaging, and ex vivo histology studies in the Axl-positive A549 tumors. In vitro cellular study showed that Axl probe, (64)Cu-DOTA-h173, was highly immuno-reactive with A549 cells. Western blot analysis confirmed that Axl is highly expressed in the A549 cell line. For microPET imaging, the A549 xenografts demonstrated a significantly higher (64)Cu-DOTA-h173 uptake compared to the NCI-H249 xenograft (a negative control model). Furthermore, (64)Cu-DOTA-h173 uptake in A549 is significantly higher than that of (64)Cu-DOTA-hIgG. Immuno-fluorescence staining was consistent with the in vivo micro-PET imaging results. In conclusion, (64)Cu-DOTA-h173 could be potentially used as a probe for noninvasive imaging of Axl expression, which could collect important information regarding tumor response to Axl-targeted therapeutic interventions.
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Anticuerpos Monoclonales Humanizados , Carcinoma de Pulmón de Células no Pequeñas/diagnóstico por imagen , Radioisótopos de Cobre , Diseño de Fármacos , Tomografía de Emisión de Positrones/métodos , Proteínas Proto-Oncogénicas/inmunología , Radiofármacos , Proteínas Tirosina Quinasas Receptoras/inmunología , Animales , Anticuerpos Monoclonales Humanizados/química , Anticuerpos Monoclonales Humanizados/farmacocinética , Western Blotting , Carcinoma de Pulmón de Células no Pequeñas/enzimología , Carcinoma de Pulmón de Células no Pequeñas/inmunología , Carcinoma de Pulmón de Células no Pequeñas/patología , Radioisótopos de Cobre/farmacocinética , Citometría de Flujo , Técnica del Anticuerpo Fluorescente , Humanos , Neoplasias Pulmonares/diagnóstico por imagen , Neoplasias Pulmonares/enzimología , Neoplasias Pulmonares/inmunología , Neoplasias Pulmonares/patología , Ratones , Ratones Desnudos , Proteínas Proto-Oncogénicas/antagonistas & inhibidores , Radiofármacos/farmacocinética , Proteínas Tirosina Quinasas Receptoras/antagonistas & inhibidores , Distribución Tisular , Células Tumorales Cultivadas , Ensayos Antitumor por Modelo de Xenoinjerto , Tirosina Quinasa del Receptor AxlRESUMEN
Recombinant protein therapeutics have increased in number and frequency since the introduction of human insulin, 25 years ago. Presently, proteins and peptides are commonly used in the clinic. However, the incorporation of peptides into clinically approved nanomedicines has been limited. Reasons for this include the challenges of decorating pharmaceutical-grade nanoparticles with proteins by a process that is robust, scalable, and cost-effective. As an alternative to covalent bioconjugation between a protein and nanoparticle, we report that biologically active proteins may themselves mediate the formation of small multimers through steric stabilization by large protein polymers. Unlike multistep purification and bioconjugation, this approach is completed during biosynthesis. As proof-of-principle, the disintegrin protein called vicrostatin (VCN) was fused to an elastin-like polypeptide (A192). A significant fraction of fusion proteins self-assembled into multimers with a hydrodynamic radius of 15.9 nm. The A192-VCN fusion proteins compete specifically for cell-surface integrins on human umbilical vein endothelial cells (HUVECs) and two breast cancer cell lines, MDA-MB-231 and MDA-MB-435. Confocal microscopy revealed that, unlike linear RGD-containing protein polymers, the disintegrin fusion protein undergoes rapid cellular internalization. To explore their potential clinical applications, fusion proteins were characterized using small animal positron emission tomography (microPET). Passive tumor accumulation was observed for control protein polymers; however, the tumor accumulation of A192-VCN was saturable, which is consistent with integrin-mediated binding. The fusion of a protein polymer and disintegrin results in a higher intratumoral contrast compared to free VCN or A192 alone. Given the diversity of disintegrin proteins with specificity for various cell-surface integrins, disintegrin fusions are a new source of biomaterials with potential diagnostic and therapeutic applications.
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Neoplasias de la Mama/tratamiento farmacológico , Polímeros/química , Polímeros/farmacología , Proteínas Recombinantes de Fusión/química , Proteínas Recombinantes de Fusión/farmacología , Animales , Materiales Biocompatibles/química , Materiales Biocompatibles/farmacología , Línea Celular Tumoral/efectos de los fármacos , Membrana Celular/efectos de los fármacos , Membrana Celular/metabolismo , Desintegrinas/química , Desintegrinas/farmacología , Elastina/química , Células Endoteliales de la Vena Umbilical Humana/efectos de los fármacos , Humanos , Integrinas/metabolismo , Ratones Desnudos , Microscopía Confocal , Microscopía Electrónica de Transmisión , Nanopartículas/química , Péptidos/química , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
A new class of multifunctionalized sarcophagine derivatives was synthesized for 64Cu chelation. The platform developed in this study could have broad applications in 64Cu-radiopharmaceuticals.
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Quelantes/síntesis química , Complejos de Coordinación/síntesis química , Dipéptidos/química , Quelantes/química , Complejos de Coordinación/química , Radioisótopos de Cobre/química , Marcaje Isotópico/métodos , Tomografía de Emisión de Positrones , Radiofármacos/químicaRESUMEN
We present a 49-year-old woman requiring living donor liver transplantation after liver metastasis from a pancreatic solid pseudopapillary tumor. After identifying a pancreatic mass and liver lesions, she underwent extensive surgical resection. Pathology revealed a solid pseudopapillary neoplasm of the head and body of the pancreas, extending into the peripancreatic soft tissues and confirmed to have spread to the liver. Subsequently, she underwent adjuvant chemotherapy and radiofrequency ablations of the new liver lesions. Despite immunotherapy and chemotherapy, there was a progression of the lesions. With interval growth of liver lesions, without evidence of extrahepatic disease, she underwent living donor liver transplantation.
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BACKGROUND AND OBJECTIVE: Positron emission tomography (PET) imaging has been useful in delineating tumor volumes and allowing for improved radiation treatment. The field of PET-guided radiotherapy is rapidly growing and will have significant impact on radiotherapy delivery in the future. This narrative review provides an overview of the current state of PET-guided radiotherapy as well as the future directions of the field. METHODS: For this narrative review, PubMed was searched for articles from 2010-2023. A total of 18 keywords or phrases were searched to provide an overview of PET-guided radiotherapy, radiotracers, the role of PET-guided radiotherapy in oligometastatic disease, and biology-guided radiotherapy (BgRT). The first 300 results for each keyword were searched and relevant articles were extracted. The references of these articles were also reviewed for relevant articles. KEY CONTENT AND FINDINGS: In radiotherapy, 18F-2-fluoro-2-deoxy-D-glucose (F-FDG or FDG) is the major radiotracer for PET and when combined with computed tomography (CT) scan allows for anatomic visualization of metabolically active malignancy. Novel radiotracers are being explored to delineate certain cell types and numerous tumor metrics including metabolism, hypoxia, vascularity, and cellular proliferation. This molecular and functional imaging will provide improved tumor characterization. Through these radiotracers, radiation plans can employ dose painting by creating different dose levels based upon specific risk factors of the target volume. Additionally, biologic imaging during radiotherapy can allow for adaptation of the radiation plan based on response to treatment. Dose painting and adaptive radiotherapy should improve the therapeutic ratio through more selective dose delivery. The novel PET-linear accelerator hopes to combine these techniques and more by using radiotracers to deliver BgRT. The areas of radiotracer uptake will serve as fiducials to guide radiotherapy to themselves. This technique may prove promising in the growing area of oligometastatic radiation treatment. CONCLUSIONS: Significant challenges exist for the future of PET-guided radiotherapy. However, with the advancements being made, PET imaging is set to change the delivery of radiotherapy.
Asunto(s)
Tomografía de Emisión de Positrones , Radioterapia Guiada por Imagen , Humanos , Tomografía de Emisión de Positrones/métodos , Radioterapia Guiada por Imagen/métodos , Neoplasias/radioterapia , Neoplasias/diagnóstico por imagenRESUMEN
18F positron emission tomography (PET) has a number of attributes that make it clinically attractive, including nearly 100% positron efficiency, very high specific radioactivity, and a short half-life of ≈ 110 minutes. However, the short half-life of 18F and the poor nucleophilicity of fluoride introduce challenges for the incorporation of 18F into complex molecules. Recently, the tetrazine-trans-cyclooctene ligation was introduced as a novel 18F labeling method that proceeds with fast reaction rates without catalysis. Herein we report an efficient method for 18F labeling of free cysteines of peptides and proteins based on sequential ligation with a bifunctional tetrazinyl-maleimide and an 18F-labeled trans-cyclooctene. The newly developed method was tested for site-specific labeling of both c(RGDyC) peptide and vascular endothelial growth factor (VEGF)-SH protein. Starting with 4 mCi of 18F-trans-cyclooctene and only 10 µg of tetrazine-RGD (80-100 µM) or 15 µg of tetrazine-VEGF (6.0 µM), 18F-labeled RGD peptide and VEGF protein could be obtained within 5 minutes in 95% yield and 75% yield, respectively. The obtained tracers were then evaluated in mice. In conclusion, a highly efficient method has been developed for site-specific 18F labeling of cysteine-containing peptides and proteins. The special characteristics of the tetrazine-trans-cyclooctene ligation provide unprecedented opportunities to synthesize 18F-labeled probes with high specific activity for PET applications.
Asunto(s)
Ciclooctanos/química , Cisteína/química , Radioisótopos de Flúor , Péptidos/química , Tomografía de Emisión de Positrones/métodos , Proteínas/química , Línea Celular Tumoral , HumanosRESUMEN
The NGR-containing peptides have been shown to bind specifically to CD13/aminopeptidase N (APN) receptor, one of the attractive tumor vasculature biomarkers. In this study, we evaluated (64)Cu-labeled monomeric and dimeric NGR peptides for microPET imaging of CD13 receptor expression in vivo. Western blot analysis and immunofluorescence staining were performed to identify CD13-positive and CD13-negative cell lines. NGR-containing peptides were conjugated with 1,4,7,10-tetraazadodecane-N,N',Nâ³,Nâ´-tetraacetic acid (DOTA) and labeled with (64)Cu (t(1/2) = 12.7 h) in ammonium acetate buffer. The resulting monomeric ((64)Cu-DOTA-NGR1) and dimeric ((64)Cu-DOTA-NGR2) peptides were then subjected to in vitro stability, cell uptake and efflux, small animal micorPET, and biodistribution studies. In vitro studies demonstrated that CD13 receptors are overexpressed in human fibrosarcoma HT-1080 cells and negative in human colon adenocarcinoma HT-29 cells. The binding affinity of (64)Cu-DOTA-NGR2 to HT-1080 cells was measured to be within low nanomolar range and about 2-fold higher than that of (64)Cu-DOTA-NGR1. For small animal microPET studies, (64)Cu-DOTA-NGR2 displayed more favorable in vivo performance in terms of higher tumor uptake and slower tumor washout in CD13-positive HT-1080 tumor xenografts as compared to (64)Cu-DOTA-NGR1. As expected, significantly lower tumor uptake and poorer tumor/normal organ contrast were observed for both (64)Cu-DOTA-NGR1 and (64)Cu-DOTA-NGR2 in CD13-negative HT-29 tumor xenografts in comparison with those in the HT-1080 tumor xenografts. The CD13-specific tumor activity accumulation of both (64)Cu-DOTA-NGR1 and (64)Cu-DOTA-NGR2 was further demonstrated by significant reduction of tumor uptake in HT-1080 tumor xenografts with a coinjected blocking dose of cyclic NGR peptide [c(CNGRC)]. The biodistribution results were consistent with the quantitative analysis of microPET imaging. We concluded that both (64)Cu-DOTA-NGR1 and (64)Cu-DOTA-NGR2 have good and specific tumor uptake in CD13-positive HT-1080 tumor xenografts. (64)Cu-DOTA-NGR2 showed higher tumor uptake and better tumor retention than (64)Cu-DOTA-NGR1, presumably due to bivalency effect and increase in apparent molecular size. (64)Cu-DOTA-NGR2 is a promising PET probe for noninvasive detection of CD13 receptor expression in vivo.
Asunto(s)
Antígenos CD13/metabolismo , Radioisótopos de Cobre/química , Neoplasias/diagnóstico por imagen , Oligopéptidos/química , Tomografía de Emisión de Positrones/métodos , Radiofármacos/química , Animales , Línea Celular Tumoral , Femenino , Células HT29 , Humanos , Ratones , Ratones Desnudos , Neoplasias/diagnóstico , Neoplasias/metabolismo , Distribución TisularRESUMEN
Accumulating evidence suggests that overexpression of the tyrosine kinase receptor EphB4, a mediator of vascular development, is a novel target for tumor diagnosis, prognosis and therapy. Noninvasive imaging of EphB4 expression could therefore be valuable for evaluating disease course and therapeutic efficacy at the earliest stages of anti-EphB4 treatment. In this study, we systematically investigated the use of anti-EphB4 antibody h131 (150 kDa) and its fragments (h131-F(ab')2, 110 kDa; h131-Fab, 50 kDa) for near-infrared fluorescence (NIRF) imaging of EphB4 expression in vivo. h131-F(ab')2 and h131-Fab were produced through pepsin and papain digestion of h131 respectively, whose purity was confirmed by FPLC and SDS-PAGE. After conjugation with Cy5.5, in vivo characteristics of h131, h131-F(ab')2 and h131-Fab were evaluated in EphB4-positive HT29 tumor model. Although h131-Cy5.5 demonstrated highest tumor uptake among these probes, its optimal tumor uptake level was obtained at 2 days post injection (p.i.). For h131-Fab-Cy5.5, maximum tumor uptake was achieved at 4 h p.i. However, no significant difference was observed between h131-Fab-Cy5.5 and hIgG-Fab-Cy5.5, indicating the tumor accumulation was mainly caused by passive targeting. In contrast, h131-F(ab')2-Cy5.5 demonstrated prominent tumor uptake at 6 h p.i. The target specificity was confirmed by hIgG-F(ab')2-Cy5.5 control and immunofluorescent staining. Collectively, h131-F(ab')2 exhibited prominent and specific tumor uptake at early time points, which suggests it is a promising agent for EphB4-targeted imaging.
Asunto(s)
Anticuerpos Monoclonales/inmunología , Neoplasias/diagnóstico , Neoplasias/inmunología , Receptor EphB4/inmunología , Línea Celular Tumoral , Diagnóstico por Imagen/métodos , Células HT29 , Humanos , Distribución Tisular/inmunologíaRESUMEN
Accumulating evidence suggests that EphB4 plays key roles in cancer progression in numerous cancer types. In fact, therapies focusing on EphB4 have become potentially important components of various cancer treatment strategies. However, tumor sensitivity to EphB4 suppression may not be uniform for different cancers. In this study, we developed near-infrared fluorescence (NIRF) probes for EphB4 targeted imaging, based on EphB4-specific humanized monoclonal antibody hAb47. NIRF dye Cy5.5 was introduced to hAb47 either through the reaction with amino groups (named hAb47-Cy5.5) or sulfhydryl groups (named hAb47-Cy5.5-Mal). The resulting probes were evaluated in both HT-29 xenograft and the mAb131 (anti-EphB4) treated models. Although these methods lead to modifications of both the heavy chain and light chain of the antibody, the majority of the EphB4 binding affinity was maintained (81.62 ± 2.08% for hAb47-Cy5.5 and 77.14 ± 2.46% for hAb47-Cy5.5-Mal, respectively). hAb47-Cy5.5 was then chosen for in vivo NIRF imaging of EphB4 expression. In HT29 colorectal tumor xenografts, hAb47-Cy5.5 demonstrated significantly higher tumor uptake compared with that of the hIgG-Cy5.5 control, which was further confirmed by immunofluorescent staining. Moreover, hAb47-Cy5.5 successfully imaged the decreased EphB4 expression (confirmed by Western blot) in EphB4-targeted immunotherapy using another EphB4-specific antibody, mAb131. Collectively, hAb47-Cy5.5 could be used as a specific NIRF contrast agent for noninvasive imaging of EphB4 expression, which may predict whether an individual tumor would likely respond to EphB4 targeted interventions, as well as monitor the therapeutic response.
Asunto(s)
Carbocianinas/farmacocinética , Diagnóstico por Imagen/métodos , Colorantes Fluorescentes/farmacocinética , Neoplasias/diagnóstico , Neoplasias/metabolismo , Receptor EphB4/metabolismo , Animales , Anticuerpos Monoclonales/farmacología , Anticuerpos Monoclonales Humanizados/metabolismo , Línea Celular Tumoral , Monitoreo del Ambiente/métodos , Femenino , Células HT29 , Humanos , Ratones , Ratones Endogámicos BALB C , Ratones Desnudos , Neoplasias/terapia , Espectroscopía Infrarroja Corta/métodos , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
Positron emission tomography (PET) imaging of receptor integrin αv ß3 expression may play a key role in the early detection of cancer and cardiovascular diseases, monitoring disease progression, evaluating therapeutic response, and aiding anti-angiogenic drugs discovery and development. The last decade has seen the development of new PET tracers for in vivo imaging of integrin αv ß3 expression along with advances in PET chemistry. In this review, we will focus on the radiochemistry development of PET tracers based on arginine-glycine-aspartic acid (RGD) peptide, present an overview of general strategies for preparing RGD-based PET tracers, and review the recent advances in preparations of (18) F-labeled, (64) Cu-labeled, and (68) Ga-labeled RGD tracers, RGD-based PET multivalent probes, and RGD-based PET multimodality probes for imaging receptor integrin αv ß3 expression.