RESUMEN
PURPOSE: Exploring synaptic density changes during brain growth is crucial to understanding brain development. Previous studies in nonhuman primates report a rapid increase in synapse number between the late gestational period and the early neonatal period, such that synaptic density approaches adult levels by birth. Prenatal synaptic development may have an enduring impact on postnatal brain development, but precisely how synaptic density changes in utero are unknown because current methods to quantify synaptic density are invasive and require post-mortem brain tissue. METHODS: We used synaptic vesicle glycoprotein 2A (SV2A) positron emission tomography (PET) radioligands [11C]UCB-J and [18F]Syn-VesT-1 to conduct the first assessment of synaptic density in the developing fetal brain in gravid rhesus monkeys. Eight pregnant monkeys were scanned twice during the third trimester at two imaging sites. Fetal post-mortem samples were collected near term in a subset of subjects to quantify SV2A density by Western blot. RESULTS: Image-derived fetal brain SV2A measures increased during the third trimester. SV2A concentrations were greater in subcortical regions than in cortical regions at both gestational ages. Near term, SV2A density was higher in primary motor and visual areas than respective associative regions. Post-mortem quantification of SV2A density was significantly correlated with regional SV2A PET measures. CONCLUSION: While further study is needed to determine the exact relationship of SV2A and synaptic density, the imaging paradigm developed in the current study allows for the effective in vivo study of SV2A development in the fetal brain.
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Encéfalo , Glicoproteínas de Membrana , Proteínas del Tejido Nervioso , Animales , Encéfalo/diagnóstico por imagen , Encéfalo/metabolismo , Macaca mulatta/metabolismo , Glicoproteínas de Membrana/metabolismo , Proteínas del Tejido Nervioso/metabolismo , Tomografía de Emisión de Positrones/métodosRESUMEN
Women exposed to a variety of viral and bacterial infections during pregnancy have an increased risk of giving birth to a child with autism, schizophrenia or other neurodevelopmental disorders. Preclinical maternal immune activation (MIA) models are powerful translational tools to investigate mechanisms underlying epidemiological links between infection during pregnancy and offspring neurodevelopmental disorders. Our previous studies documenting the emergence of aberrant behavior in rhesus monkey offspring born to MIA-treated dams extends the rodent MIA model into a species more closely related to humans. Here we present novel neuroimaging data from these animals to further explore the translational potential of the nonhuman primate MIA model. Nine male MIA-treated offspring and 4 controls from our original cohort underwent in vivo positron emission tomography (PET) scanning at approximately 3.5-years of age using [18F] fluoro-l-m-tyrosine (FMT) to measure presynaptic dopamine levels in the striatum, which are consistently elevated in individuals with schizophrenia. Analysis of [18F]FMT signal in the striatum of these nonhuman primates showed that MIA animals had significantly higher [18F]FMT index of influx compared to control animals. In spite of the modest sample size, this group difference reflects a large effect size (Cohen's d = 0.998). Nonhuman primates born to MIA-treated dams exhibited increased striatal dopamine in late adolescence-a hallmark molecular biomarker of schizophrenia. These results validate the MIA model in a species more closely related to humans and open up new avenues for understanding the neurodevelopmental biology of schizophrenia and other neurodevelopmental disorders associated with prenatal immune challenge.
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Conducta Animal/efectos de los fármacos , Cuerpo Estriado/fisiología , Dopamina/fisiología , Neostriado/patología , Animales , Cuerpo Estriado/diagnóstico por imagen , Modelos Animales de Enfermedad , Femenino , Macaca mulatta , Masculino , Neostriado/inmunología , Poli I-C/farmacología , Tomografía de Emisión de Positrones , Embarazo , Efectos Tardíos de la Exposición Prenatal/inmunología , Efectos Tardíos de la Exposición Prenatal/patología , Esquizofrenia/inmunología , Conducta EstereotipadaRESUMEN
The central component of the complement cascade, C3, is involved in various biological functions, including opsonization of foreign bodies, clearance of waste material, activation of immune cells, and triggering of pathways controlling development. Given its broad role in immune responses, particularly in phagocytosis and the clearance of microbes, a deficiency in complement C3 in humans is often associated with multiple bacterial infections. Interestingly, an increased susceptibility to infections appears to occur mainly in the first two years of life and then wanes throughout adulthood. In view of the well-established connection between C3 deficiency and infections, therapeutic inhibition of complement at the level of C3 is often considered with caution or disregarded. We therefore set out to investigate the immune and biochemical profile of non-human primates under prolonged treatment with the C3 inhibitor compstatin (Cp40 analog). Cynomolgus monkeys were dosed subcutaneously with Cp40, resulting in systemic inhibition of C3, for 1â¯week, 2â¯weeks, or 3â¯months. Plasma concentrations of both C3 and Cp40 were measured periodically and complete saturation of plasma C3 was confirmed. No differences in hematological, biochemical, or immunological parameters were identified in the blood or tissues of animals treated with Cp40 when compared to those injected with vehicle alone. Further, skin wounds showed no signs of infection in those treated with Cp40. In fact, Cp40 treatment was associated with a trend toward accelerated wound healing when compared with the control group. In addition, a biodistribution study in a rhesus monkey indicated that the distribution of Cp40 in the body is associated with the presence of C3, concentrating in organs that accumulate blood and produce C3. Overall, our data suggest that systemic C3 inhibition in healthy adult non-human primates is not associated with a weakened immune system or susceptibility to infections.
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Complemento C3/antagonistas & inhibidores , Inactivadores del Complemento/toxicidad , Péptidos Cíclicos/toxicidad , Cicatrización de Heridas/inmunología , Infección de Heridas/epidemiología , Animales , Complemento C3/inmunología , Complemento C3/metabolismo , Inactivadores del Complemento/farmacocinética , Macaca fascicularis , Macaca mulatta , Péptidos Cíclicos/farmacocinética , Factores de Tiempo , Distribución Tisular , Heridas y Lesiones/inmunologíaRESUMEN
Lymphatic dysfunction is associated with the progression of many cardiovascular disorders due to their role in maintaining tissue fluid homeostasis. Promoting new lymphatic vessels (lymphangiogenesis) is a promising strategy to reverse these cardiovascular disorders via restoring lymphatic function. Vascular endothelial growth factor (VEGF) members VEGF-C and VEGF-D are both potent candidates for stimulating lymphangiogenesis, though maintaining spatial and temporal control of these factors represents a challenge to developing efficient therapeutic lymphangiogenic applications. Injectable alginate hydrogels have been useful for the controlled delivery of many angiogenic factors, including VEGF-A, to stimulate new blood vasculature. However, the utility of these tunable hydrogels for delivering lymphangiogenic factors has never been closely examined. Thus, the objective of this study was to utilize ionically cross-linked alginate hydrogels to deliver VEGF-C and VEGF-D for potential lymphangiogenic applications. We demonstrated that lymphatic endothelial cells (LECs) are sensitive to temporal presentation of VEGF-C and VEGF-D but with different responses between the factors. The greatest LEC mitogenic and sprouting response was observed for constant concentrations of VEGF-C and a high initial concentration that gradually decreased over time for VEGF-D. Additionally, alginate hydrogels provided sustained release of radiolabeled VEGF-C and VEGF-D. Finally, VEGF-C and VEGF-D released from these hydrogels promoted a similar number of LEC sprouts as exogenously added growth factors and new vasculature in vivo via a chick chorioallantoic membrane (CAM) assay. Overall, these findings demonstrate that alginate hydrogels can provide sustained and bioactive release of VEGF-C and VEGF-D which could have applications for therapeutic lymphangiogenesis.
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Células Endoteliales/efectos de los fármacos , Hidrogeles , Linfangiogénesis/efectos de los fármacos , Neovascularización Fisiológica/efectos de los fármacos , Factor C de Crecimiento Endotelial Vascular/administración & dosificación , Factor D de Crecimiento Endotelial Vascular/administración & dosificación , Alginatos , Células Cultivadas , Preparaciones de Acción Retardada , Células Endoteliales/metabolismo , Ácido Glucurónico , Ácidos Hexurónicos , HumanosRESUMEN
Pair bonding leads to increases in dopamine D1 receptor (D1R) binding in the nucleus accumbens of monogamous prairie voles. In the current study, we hypothesized that there is similar up-regulation of D1R in a monogamous primate, the titi monkey (Callicebus cupreus). Receptor binding of the D1R antagonist [11 C]-SCH23390 was measured in male titi monkeys using PET scans before and after pairing with a female. We found that within-subject analyses of pairing show significant increases in D1R binding in the lateral septum, but not the nucleus accumbens, caudate, putamen, or ventral pallidum. The lateral septum is involved in a number of processes that may contribute to social behavior, including motivation, affect, reward, and reinforcement. This region also plays a role in pair bonding and paternal behavior in voles. Our observations of changes in D1R in the lateral septum, but not the nucleus accumbens, suggest that there may be broadly similar dopaminergic mechanisms underlying pair bonding across mammalian species, but that the specific changes to neural circuitry differ. This study is the first research to demonstrate neuroplasticity of the dopamine system following pair bonding in a non-human primate; however, substantial variability in the response to pairing suggests the utility of further research on the topic.
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Apareamiento , Pitheciidae , Receptores de Dopamina D1 , Conducta Social , Animales , Femenino , Masculino , Apego a ObjetosRESUMEN
BACKGROUND: Acute intoxication with organophosphorus (OP) cholinesterase inhibitors can trigger convulsions that progress to life-threatening status epilepticus. Survivors face long-term morbidity including mild-to-severe decline in memory. It is posited that neuroinflammation plays a key role in the pathogenesis of OP-induced neuropsychiatric deficits. Rigorous testing of this hypothesis requires preclinical models that recapitulate relevant phenotypic outcomes. Here, we describe a rat model of acute intoxication with the OP diisopropylfluorophosphate (DFP) that exhibits persistent neuroinflammation and cognitive impairment. METHODS: Neuroinflammation, neurodegeneration, and cognitive function were compared in adult male Sprague Dawley rats injected with an acutely toxic dose of DFP vs. vehicle controls at multiple time points up to 36 days post-exposure. Neuroinflammation was quantified using immunohistochemical biomarkers of microglia (ionized calcium-binding adapter molecule 1, IBA1) and activated astrocytes (glial fibrillary acidic protein, GFAP) and positron emission tomography (PET) imaging of [11C]-(R)-PK11195, a ligand for the 18-kDa mitochondrial membrane translocator protein (TSPO). FluoroJade-B staining was used to assess neurodegeneration; Pavlovian conditioning, to assess cognitive function. RESULTS: Animals exhibited moderate-to-severe seizures within minutes of DFP injection that continued for up to 6 h post-injection. As indicated by IBA1 and GFAP immunoreactivity and by PET imaging of TSPO, acute DFP intoxication triggered neuroinflammation in the hippocampus and cortex during the first 3 days that peaked at 7 days and persisted to 21 days post-exposure in most animals. Neurodegeneration was detected in multiple brain regions from 1 to 14 days post-exposure. All DFP-intoxicated animals exhibited significant deficits in contextual fear conditioning at 9 and 20 days post-exposure compared to vehicle controls. Whole-brain TSPO labeling positively correlated with seizure severity score, but did not correlate with performance in the contextual fear-conditioning task. CONCLUSIONS: We describe a preclinical model in which acute DFP intoxication causes seizures, persistent neuroinflammation, neurodegeneration, and memory impairment. The extent of the neuroinflammatory response is influenced by seizure severity. However, the observation that a subset of animals with moderate seizures and minimal TSPO labeling exhibited cognitive deficits comparable to those of animals with severe seizures and significant TSPO labeling suggests that DFP may impair learning and memory circuitry via mechanisms independent of seizures or neuroinflammation.
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Inhibidores de la Colinesterasa/toxicidad , Disfunción Cognitiva/inducido químicamente , Encefalitis/inducido químicamente , Isoflurofato/toxicidad , Animales , Proteínas de Unión al Calcio/metabolismo , Proteínas Portadoras/metabolismo , Disfunción Cognitiva/diagnóstico por imagen , Condicionamiento Clásico/efectos de los fármacos , Encefalitis/diagnóstico por imagen , Conducta Exploratoria/efectos de los fármacos , Proteína Ácida Fibrilar de la Glía/metabolismo , Imagen por Resonancia Magnética , Masculino , Proteínas de Microfilamentos/metabolismo , Tomografía de Emisión de Positrones , Ratas , Ratas Sprague-Dawley , Receptores de GABA-A/metabolismo , Análisis de Regresión , Factores de TiempoRESUMEN
The neuropeptide oxytocin is part of a neuroendocrine system that has physiological effects ranging from ensuring uterine myometrial contractions at parturition and post-partum mammary gland milk ejection to the modulation of neural control of social relationships. This initial study was performed to investigate the potential use of positron emission tomography (PET) for localizing oxytocin receptors in two New World primates. Three biomarkers for PET (1-3) that are known to have high affinity and selectivity for the human oxytocin receptor were investigated in the common marmoset (Callithrix jacchus) via PET imaging. Brain penetration, and uptake in the salivary gland area were both observed with biomarkers 2 and 3. No brain penetration was observed with 1, but uptake was observed more specifically in several peripheral endocrine glands compared to 2 or 3. Biomarker 2, which displayed the best brain penetration of the three biomarkers in the marmoset, was then investigated in the monogamous coppery titi monkey (Callicebus cupreus) in a brain scan and a limited full body scan. No significant brain penetration of 2 was observed in the titi monkey, but significant uptake was observed in various locations throughout the periphery. Metabolism of 2 was suspected to have been significant based upon HPLC analysis of blood draws, but parent compound was still present near the end of the scan. Follow-up investigations will focus on next generation biomarkers bearing improved binding characteristics and brain penetrability as well as investigating tissue in regions where biomarker uptake was observed.
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Tomografía de Emisión de Positrones , Receptores de Oxitocina/análisis , Bibliotecas de Moléculas Pequeñas/farmacocinética , Animales , Biomarcadores/análisis , Biomarcadores/metabolismo , Ligandos , Estructura Molecular , Platirrinos , Receptores de Oxitocina/metabolismo , Bibliotecas de Moléculas Pequeñas/química , Bibliotecas de Moléculas Pequeñas/metabolismoRESUMEN
Multifunctional nanoparticles with combined diagnostic and therapeutic functions show great promise towards personalized nanomedicine. However, attaining consistently high performance of these functions in vivo in one single nanoconstruct remains extremely challenging. Here we demonstrate the use of one single polymer to develop a smart 'all-in-one' nanoporphyrin platform that conveniently integrates a broad range of clinically relevant functions. Nanoporphyrins can be used as amplifiable multimodality nanoprobes for near-infrared fluorescence imaging (NIRFI), magnetic resonance imaging (MRI), positron emission tomography (PET) and dual modal PET-MRI. Nanoporphyrins greatly increase the imaging sensitivity for tumour detection through background suppression in blood, as well as preferential accumulation and signal amplification in tumours. Nanoporphyrins also function as multiphase nanotransducers that can efficiently convert light to heat inside tumours for photothermal therapy (PTT), and light to singlet oxygen for photodynamic therapy (PDT). Furthermore, nanoporphyrins act as programmable releasing nanocarriers for targeted delivery of drugs or therapeutic radio-metals into tumours.
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Portadores de Fármacos/química , Nanoestructuras/química , Nanoestructuras/uso terapéutico , Porfirinas/química , Nanomedicina Teranóstica/métodos , Animales , Línea Celular Tumoral/efectos de los fármacos , Dendrímeros/química , Doxorrubicina/administración & dosificación , Doxorrubicina/química , Portadores de Fármacos/farmacocinética , Femenino , Fluorescencia , Humanos , Imagen por Resonancia Magnética/métodos , Masculino , Ratones Desnudos , Ratones Transgénicos , Nanoestructuras/administración & dosificación , Fotoquimioterapia/métodos , Porfirinas/sangre , Porfirinas/farmacocinética , Ratas Sprague-Dawley , Especies Reactivas de Oxígeno/metabolismo , Espectroscopía Infrarroja Corta/métodos , Nanomedicina Teranóstica/instrumentaciónRESUMEN
These studies focused on a new radiolabeling technique with copper ((64)Cu) and zirconium ((89)Zr) for positron emission tomography (PET) imaging using a CD45 antibody. Synthesis of (64)Cu-CD45 and (89)Zr-CD45 immunoconjugates was performed and the evaluation of the potential toxicity of radiolabeling human peripheral blood stem cells (hPBSC) was assessed in vitro (viability, population doubling times, colony forming units). hPBSC viability was maintained as the dose of (64)Cu-TETA-CD45 increased from 0 (92%) to 160 µCi/mL (76%, p>0.05). Radiolabeling efficiency was not significantly increased with concentrations of (64)Cu-TETA-CD45 >20 µCi/mL (p>0.50). Toxicity affecting both growth and colony formation was observed with hPBSC radiolabeled with ≥40 µCi/mL (p<0.05). For (89)Zr, there were no significant differences in viability (p>0.05), and a trend towards increased radiolabeling efficiency was noted as the dose of (89)Zr-Df-CD45 increased, with a greater level of radiolabeling with 160 µCi/mL compared to 0-40 µCi/mL (p<0.05). A greater than 2,000 fold-increase in the level of (89)Zr-Df-CD45 labeling efficiency was observed when compared to (64)Cu-TETA-CD45. Similar to (64)Cu-TETA-CD45, toxicity was noted when hPBSC were radiolabeled with ≥40 µCi/mL (p<0.05) (growth, colony formation). Taken together, 20 µCi/mL resulted in the highest level of radiolabeling efficiency without altering cell function. Young rhesus monkeys that had been transplanted prenatally with 25×10(6) hPBSC expressing firefly luciferase were assessed with bioluminescence imaging (BLI), then 0.3 mCi of (89)Zr-Df-CD45, which showed the best radiolabeling efficiency, was injected intravenously for PET imaging. Results suggest that (89)Zr-Df-CD45 was able to identify engrafted hPBSC in the same locations identified by BLI, although the background was high.
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Inmunoconjugados/química , Tomografía de Emisión de Positrones/métodos , Radiofármacos , Coloración y Etiquetado/métodos , Células Madre/química , Animales , Anticuerpos/química , Supervivencia Celular/efectos de los fármacos , Cobre/química , Femenino , Compuestos Heterocíclicos/administración & dosificación , Humanos , Inmunoconjugados/administración & dosificación , Inyecciones Intravenosas , Antígenos Comunes de Leucocito/química , Macaca mulatta , Masculino , Compuestos Organometálicos/administración & dosificación , Trasplante de Células Madre de Sangre Periférica , Radiofármacos/síntesis química , Radiofármacos/farmacología , Células Madre/citología , Circonio/químicaRESUMEN
PURPOSE: Oxidized low-density lipoprotein (LDL) plays an essential role in the pathogenesis of atherosclerosis. The purpose of this study was to characterize the pharmacokinetics (PK) of a human recombinant IgG1 antibody to oxidized LDL (anti-oxLDL) in cynomolgus monkey. The tissue biodistribution of anti-oxLDL was also investigated using positron emission tomography (PET) imaging. METHODS: Anti-oxLDL was conjugated with the N-hydroxysuccinimide ester of DOTA (1,4,7,10-tetraazacyclododecane 1,4,7,10-tetraacetic acid) and radiolabeled by chelation of radioactive copper-64 ((64)Cu) for detection by PET. Anti-oxLDL was administered as a single intravenous (IV) dose of 10 mg/kg (as a mixture of radiolabeled and non-labeled material) to two male and two female cynomolgus monkeys. Serum samples were collected over 29 days. Two ELISA methods were used to measure serum concentrations of anti-oxLDL; Assay A was a ligand binding assay that measured free anti-oxLDL (unbound and partially bound forms) and Assay B measured total anti-oxLDL. The biodistribution was observed over a 48-hour period following dose administration using PET imaging. RESULTS: Anti-oxLDL serum concentration-time profiles showed a biphasic elimination pattern that could be best described by a two-compartment elimination model. The serum concentrations obtained using the two ELISA methods were comparable. Clearance values ranged from 8 to 17 ml/day/kg, while beta half-life ranged from 8 to 12 days. The initial volume of distribution and volume of distribution at steady state were approximately 55 mL/kg and 150 mL/kg, respectively. PET imaging showed distribution predominantly to the blood pool, visible as the heart and great vessels in the trunk and limbs, plus diffuse signals in the liver, kidney, spleen, and bone marrow. CONCLUSIONS: The clearance of anti-oxLDL is slightly higher than typical IgG1 antibodies in cynomolgus monkeys. The biodistribution pattern appears to be consistent with an antibody that has no large, rapid antigen sink outside the blood space.
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Anticuerpos Monoclonales/farmacocinética , Lipoproteínas LDL/inmunología , Macaca fascicularis/inmunología , Tomografía de Emisión de Positrones , Animales , Anticuerpos Monoclonales/administración & dosificación , Anticuerpos Monoclonales/sangre , Femenino , Compuestos Heterocíclicos con 1 Anillo/farmacocinética , Humanos , Inyecciones Intravenosas , Macaca fascicularis/sangre , Masculino , Factores de Tiempo , Distribución TisularRESUMEN
Theranostic agents are critical for improving the diagnosis and treatment of non-Hodgkin Lymphoma (NHL). The peptidomimetic LLP2A is a novel peptide receptor radiotherapy candidate for treating NHL that expresses the activated α4ß1 integrin. Tumor-bearing dogs are an excellent model of human NHL with similar clinical characteristics, behavior, and compressed clinical course. Canine in vivo imaging studies will provide valuable biodistribution and affinity information that reflects a diverse clinical population of lymphoma. This may also help to determine potential dose-limiting radiotoxicity to organs in human clinical trials. To validate this construct in a naturally occurring model of NHL, we performed in-vivo molecular targeted imaging and biodistribution in 3 normal dogs and 5 NHL bearing dogs. (99m)Tc-LLP2A-HYNIC-PEG and (99m)Tc-LLP2A-HYNIC were successfully synthesized and had very good labeling efficiency and radiochemical purity. (99m)Tc-LLP2A-HYNIC and (99m)Tc-LLP2A-HYNIC-PEG had biodistribution in keeping with their molecular size, with (99m)Tc-LLP2A-HYNIC-PEG remaining longer in the circulation, having higher tissue uptake, and having more activity in the liver compared to (99m)Tc-LLP2A-HYNIC. (99m)Tc-LLP2A-HYNIC was mainly eliminated through the kidneys with some residual activity. Radioactivity was reduced to near-background levels at 6 hours after injection. In NHL dogs, tumor showed moderately increased activity over background, with tumor activity in B-cell lymphoma dogs decreasing after chemotherapy. This compound is promising in the development of targeted drug-delivery radiopharmaceuticals and may contribute to translational work in people affected by non-Hodgkin lymphoma.
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Linfoma no Hodgkin/diagnóstico , Compuestos de Organotecnecio/química , Radiofármacos/química , Animales , Dipéptidos/química , Dipéptidos/farmacocinética , Dipéptidos/toxicidad , Perros , Ratones , Compuestos de Organotecnecio/farmacocinética , Compuestos de Organotecnecio/toxicidad , Compuestos de Fenilurea/química , Compuestos de Fenilurea/farmacocinética , Compuestos de Fenilurea/toxicidad , Radiofármacos/farmacocinética , Radiofármacos/toxicidad , Distribución Tisular , Imagen de Cuerpo EnteroRESUMEN
We describe the synthesis and development of new reactive DOTA-metal complexes for covalently targeting engineered receptors in vivo, which have superior tumor uptake and clearance properties for biomedical applications. These probes are found to clear efficiently through the kidneys and minimally through other routes, but bind persistently in the tumor target. We also explore the new technique of Cerenkov luminescence imaging to optically monitor radiolabeled probe distribution and kinetics in vivo. Cerenkov luminescence imaging uniquely enables sensitive noninvasive in vivo imaging of a ß(-) emitter such as (90)Y with an optical imager.
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Diagnóstico por Imagen/métodos , Diagnóstico por Imagen/enfermería , Sondas Moleculares/síntesis química , Neoplasias/diagnóstico , Neoplasias/tratamiento farmacológico , Disulfuros , Luminiscencia , Sondas Moleculares/uso terapéutico , Unión ProteicaRESUMEN
Significant upregulation of the integrin alpha(v)beta(6) has been described as a prognostic indicator in several cancers, making it an attractive target for tumor imaging. This study compares variants of a PEGylated alpha(v)beta(6)-targeting peptide, bearing either an [(18)F]fluorobenzoyl prosthetic group ([(18)F]FBA-PEG-A20FMDV2) or different [(64)Cu]copper chelators (DOTA-PEG-A20FMDV2, CB-TE2A-PEG-A20FMDV2). The compounds were evaluated in vitro by enzyme-linked immunosorbent assay (against the integrin alpha(v)beta(6) and the closely related integrin alpha(v)beta(3)) and by cell labeling (alpha(v)beta(6)-positive DX3purobeta6/alpha(v)beta(6)-negative DX3puro) and in vivo using micro-positron emission tomography in a mouse model bearing paired DX3purobeta6/Dx3puro xenografts. In vitro, all three compounds showed excellent alpha(v)beta(6)-specific binding (50% inhibitory concentration [IC(50)](alpha(v)beta(6)) = 3 to 6 nmol/L; IC(50)(alpha(v)beta(3)) > 10 micromol/L). In vivo, they displayed comparable, preferential uptake for the alpha(v)beta(6)-expressing xenograft over the alpha(v)beta(6)-negative control (> 4:1 ratio at 4 hours postinjection). Whereas [(64)Cu]Cu-DOTA-PEG-A20FMDV2 resulted in increased levels of radioactivity in the liver, [(64)Cu]Cu-CB-TE2A-PEG-A20FMDV2 did not. Significantly, both (64)Cu-labeled tracers showed unexpectedly high and persistent levels of radioactivity in the kidneys (> 40% injected dose/g at 4 and 12 hours postinjection). The findings underscore the potential influence of the prosthetic group on targeted in vivo imaging of clinically relevant markers such as alpha(v)beta(6). Despite identical targeting peptide moiety and largely equal in vitro behavior, both (64)Cu-labeled tracers displayed inferior pharmacokinetics, making them in their present form less suitable candidates than the (18)F-labeled tracer for in vivo imaging of alpha(v)beta(6).
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Antígenos de Neoplasias/metabolismo , Compuestos Heterocíclicos con 1 Anillo , Integrinas/metabolismo , Neoplasias Experimentales/diagnóstico por imagen , Compuestos Organometálicos , Tomografía de Emisión de Positrones/métodos , Animales , Línea Celular , Radioisótopos de Cobre , Interpretación Estadística de Datos , Ensayo de Inmunoadsorción Enzimática , Compuestos Heterocíclicos con 1 Anillo/química , Compuestos Heterocíclicos con 1 Anillo/metabolismo , Compuestos Heterocíclicos con 1 Anillo/farmacocinética , Masculino , Ratones , Ratones Desnudos , Trasplante de Neoplasias , Neoplasias Experimentales/metabolismo , Compuestos Organometálicos/química , Compuestos Organometálicos/metabolismo , Compuestos Organometálicos/farmacocinética , Péptidos/metabolismo , Unión Proteica , Distribución TisularRESUMEN
PURPOSE: The aim of the study is to compare the tumor-specific targeting, pharmacokinetics, and biodistribution of (64)Cu-DOTA-HB22.7 when administered to xenograft-bearing mice intravenously (IV), intraperitoneally (IP), and subcutaneously (SQ). PROCEDURES: Mice bearing human non-Hodgkin's lymphoma (NHL) xenografts were injected IV, IP, or SQ with (64)Cu-DOTA-HB22.7. Xenograft targeting was evaluated by micro positron emission tomography (microPET) and confirmed by organ biodistribution studies. Blood measurements of (64)Cu were performed to determine the pharmacokinetics and clearance of (64)Cu-DOTA-HB22.7. RESULTS: (64)Cu-DOTA-HB22.7 demonstrated equivalent tumor targeting within 24-48 h, regardless of the route of administration. Organ biodistribution confirmed tumor-specific targeting. Blood pharmacokinetics demonstrated that (64)Cu-DOTA-HB22.7 accessed the bloodstream after IP and SQ administration to a similar degree as IV administration, albeit at a slower rate. CONCLUSIONS: These findings establish (64)Cu-DOTA-HB22.7 as a potential radioimmunotherapeutic and/or NHL-specific imaging agent. These findings provide evidence that IP and SQ administration can achieve results equivalent to IV administration and may lead to more efficient, reproducible treatment plans for antibody-based therapeutics.
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Anticuerpos Monoclonales/farmacocinética , Radioisótopos de Cobre/farmacocinética , Linfoma no Hodgkin/diagnóstico por imagen , Linfoma no Hodgkin/tratamiento farmacológico , Radiofármacos/farmacocinética , Animales , Anticuerpos Monoclonales/administración & dosificación , Radioisótopos de Cobre/administración & dosificación , Femenino , Citometría de Flujo , Inmunoconjugados/administración & dosificación , Inmunoconjugados/farmacocinética , Inyecciones , Ratones , Ratones Desnudos , Trasplante de Neoplasias , Tomografía de Emisión de Positrones , Radiofármacos/administración & dosificación , Radiofármacos/síntesis química , Succinimidas/administración & dosificación , Succinimidas/farmacocinética , Distribución Tisular , Trasplante Heterólogo , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
Radiolabeling of liposomes with 64Cu (t(1/2)=12.7 h) is attractive for molecular imaging and monitoring drug delivery. A simple chelation procedure, performed at a low temperature and under mild conditions, is required to radiolabel preloaded liposomes without lipid hydrolysis or the release of the encapsulated contents. Here, we report a 64Cu postlabeling method for liposomes. A 64Cu-specific chelator, 6-[p-(bromoacetamido)benzyl]-1,4,8,11-tetraazacyclotetradecane-N,N',N'',N'''-tetraacetic acid (BAT), was conjugated with an artificial lipid to form a BAT-PEG-lipid. After incorporation of 0.5% (mol/mol) BAT-PEG-lipid during liposome formulation, liposomes were successfully labeled with 64Cu in 0.1 M NH4OAc pH 5 buffer at 35 degrees C for 30-40 min with an incorporation yield as high as 95%. After 48 h of incubation of 64Cu-liposomes in 50/50 serum/PBS solution, more than 88% of the 64Cu label was still associated with liposomes. After injection of liposomal 64Cu in a mouse model, 44+/-6.9, 21+/-2.7, 15+/-2.5, and 7.4+/-1.1 (n=4) % of the injected dose per cubic centimeter remained within the blood pool at 30 min, 18, 28, and 48 h, respectively. The biodistribution at 48 h after injection verified that 7.0+/-0.47 (n=4) and 1.4+/-0.58 (n=3) % of the injected dose per gram of liposomal 64Cu and free 64Cu remained in the blood pool, respectively. Our results suggest that this fast and easy 64Cu labeling of liposomes could be exploited in tracking liposomes in vivo for medical imaging and targeted delivery.
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Radioisótopos de Cobre/química , Liposomas/química , Tomografía de Emisión de Positrones/métodos , Coloración y Etiquetado/métodos , Animales , Compuestos Heterocíclicos/química , Liposomas/farmacocinética , Masculino , Ratones , Compuestos Organometálicos/química , Péptidos/química , Polietilenglicoles/química , Especificidad por Sustrato , Factores de Tiempo , Distribución TisularRESUMEN
Complementary imaging modalities provide more information than either method alone can yield and we have developed a dual-mode imaging probe for combined magnetic resonance (MR) and positron emission tomography (PET) imaging. We have developed dual-mode PET/MRI active probes targeted to vascular inflammation and present synthesis of (1) an aliphatic amine polystyrene bead and (2) a novel superparamagnetic iron oxide nanoparticle targeted to macrophages that were both coupled to positron-emitting copper-64 isotopes. The amine groups of the polystyrene beads were directly conjugated with an amine-reactive form (isothiocyanate) of aza-macrocycle 1,4,7,10-tetraazacyclo-dodecane-1,4,7,10-tetraacetic acid (DOTA). Iron oxide nanoparticles are dextran sulfate coated, and the surface was modified to contain aldehyde groups to conjugate to an amine-activated DOTA. Incorporation of chelated Cu-64 to nanoparticles under these conditions, which is routinely used to couple DOTA to macromolecules, was unexpectedly difficult and illustrates that traditional conjugation methods do not always work in a nanoparticle environment. Therefore, we developed new methods to couple Cu-64 to nanoparticles and demonstrate successful labeling to a range of nanoparticle types. We obtained labeling yields of 24% for the amine polystyrene beads and 21% radiolabeling yield for the anionic dextran sulfate iron oxide nanoparticles. The new coupling chemistry can be generalized for attaching chelated metals to other nanoparticle platforms.
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Compuestos Férricos/síntesis química , Magnetismo , Nanopartículas/química , Coloración y Etiquetado/métodos , Aminación , Quelantes/química , Radioisótopos de Cobre , Reactivos de Enlaces Cruzados/química , Sulfato de Dextran/química , Compuestos Férricos/química , Compuestos Heterocíclicos con 1 Anillo/química , Imagen por Resonancia Magnética , Microesferas , Poliestirenos/química , Tomografía de Emisión de PositronesRESUMEN
PURPOSE: Although radioimmunotherapy alone is effective in lymphoma, its application to solid tumors will likely require a combined modality approach. In these phase I studies, paclitaxel was combined with radioimmunotherapy in patients with metastatic hormone-refractory prostate cancer or advanced breast cancer. EXPERIMENTAL DESIGN: Patients were imaged with indium-111 (111In)-1,4,7,10-tetraazacyclododecane-N,N',N'',N'''-tetraacetic acid-peptide-m170. One week later, yttrium-90 (90Y)-m170 was infused (12 mCi/m2 for prostate cancer and 22 mCi/m2 for breast cancer). Initial cohorts received radioimmunotherapy alone. Subsequent cohorts received radioimmunotherapy followed 48 hours later by paclitaxel (75 mg/m2). Cyclosporine was given to prevent development of human anti-mouse antibody. RESULTS: Bone and soft tissue metastases were targeted by 111In-m170 in 15 of the 16 patients imaged. Three prostate cancer patients treated with radioimmunotherapy alone had no grade 3 or 4 toxicity. With radioimmunotherapy and paclitaxel, two of three prostate cancer patients developed transient grade 4 neutropenia. Four breast cancer patients treated with radioimmunotherapy alone had grade 3 or 4 myelosuppression. With radioimmunotherapy and paclitaxel, both breast cancer patients developed grade 4 neutropenia. Three breast cancer patients required infusion of previously harvested peripheral blood stem cells because of neutropenic fever or bleeding. One patient in this trial developed human anti-mouse antibody in contrast to 12 of 17 patients in a prior trial using m170-radioimmunotherapy without cyclosporine. CONCLUSIONS: 111In/90Y-m170 targets prostate and breast cancer and can be combined with paclitaxel with toxicity limited to marrow suppression at the dose levels above. The maximum tolerated dose of radioimmunotherapy and fixed-dose paclitaxel with peripheral blood stem cell support has not been reached. Cyclosporine is effective in preventing human anti-mouse antibody, suggesting the feasibility of multidose, "fractionated" therapy that could enhance clinical response.
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Neoplasias de la Mama/tratamiento farmacológico , Neoplasias de la Mama/radioterapia , Paclitaxel/uso terapéutico , Neoplasias de la Próstata/tratamiento farmacológico , Neoplasias de la Próstata/radioterapia , Radioinmunoterapia/métodos , Adulto , Anciano , Anciano de 80 o más Años , Animales , Anticuerpos Monoclonales/química , Anticuerpos Monoclonales/inmunología , Anticuerpos Monoclonales/uso terapéutico , Antineoplásicos Fitogénicos/efectos adversos , Antineoplásicos Fitogénicos/uso terapéutico , Neoplasias Óseas/prevención & control , Neoplasias Óseas/secundario , Neoplasias de la Mama/patología , Catepsinas/metabolismo , Terapia Combinada , Ciclosporina/metabolismo , Ciclosporina/uso terapéutico , Relación Dosis-Respuesta a Droga , Relación Dosis-Respuesta en la Radiación , Femenino , Compuestos Heterocíclicos con 1 Anillo/química , Compuestos Heterocíclicos con 1 Anillo/farmacocinética , Humanos , Radioisótopos de Indio/química , Radioisótopos de Indio/farmacocinética , Radioisótopos de Indio/uso terapéutico , Masculino , Ratones , Persona de Mediana Edad , Neutropenia/etiología , Paclitaxel/efectos adversos , Neoplasias de la Próstata/patología , Radioinmunoterapia/efectos adversos , Neoplasias de los Tejidos Blandos/prevención & control , Neoplasias de los Tejidos Blandos/secundario , Resultado del Tratamiento , Radioisótopos de Itrio/química , Radioisótopos de Itrio/farmacocinética , Radioisótopos de Itrio/uso terapéuticoRESUMEN
PURPOSE: Radioimmunotherapy delivered by radiometal immunoconjugates and followed by marrow support is dose limited by deposition of radioactivity in normal organs. To increase elimination of radioactivity from the liver and body and, thus, minimize hepatic radiation dose, a peptide having a specific cathepsin B cleavage site was placed between the radiometal chelate DOTA (1,4,7,10-tetraazacyclododecane-N,N',N",N"'-tetraacetic acid) and the monoclonal antibody m170, and the comparative pharmacokinetics was evaluated in prostate cancer patients. EXPERIMENTAL DESIGN: (111)In-DOTA-2IT-m170 and (111)In-DOTA-peptide-(GGGF)-m170, representing the same monoclonal antibody and chelate with and without the cleavable linkage, were studied in comparable groups of prostate cancer patients (17 with In-2IT-BAD-m170 and 8 with In-DOTA-peptide-m170). Pharmacokinetics over 7 days, calculated yttrium-90 radiation dosimetry, therapeutic index, and projected maximum tolerated injected yttrium-90 dose were evaluated. RESULTS: The radioimmunoconjugates pharmacokinetics and calculated tumor and normal organ absorbed radiation dose (rads/mCi) were similar, except for a significant decrease in the mean dose to the liver (31%; P < 0.01) and lungs (31%; P < 0.01) with the DOTA-peptide immunoconjugates. Because mean tumor dose was not statistically different, this peptide linkage provided a significant increase in the therapeutic index for this tumor targeting radiopharmaceutical. If marrow support is adequate, the radiation dose historically tolerated by normal organs other than marrow would allow a 30% increase in the administered dose, resulting in a mean dose of 9500 rads to metastatic prostate cancer.
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Neoplasias de la Próstata/terapia , Radioinmunoterapia/métodos , Anticuerpos Monoclonales/uso terapéutico , Quelantes/uso terapéutico , Compuestos Heterocíclicos con 1 Anillo/uso terapéutico , Humanos , Radioisótopos de Indio/uso terapéutico , Masculino , Modelos Químicos , Péptidos/química , Radiometría , Radiofármacos/uso terapéutico , Factores de Tiempo , Distribución Tisular , Resultado del Tratamiento , Radioisótopos de Itrio/uso terapéuticoRESUMEN
CD22 is a membrane glycophosphoprotein found on nearly all healthy B-lymphocytes and most B-cell lymphomas. Recent in vitro studies have identified several anti-CD22 monoclonal antibodies (mAbs) that block the interaction of CD22 with its ligand. One of these mAbs, HB22.7, has been shown to effectively induce apoptosis in several B-cell lymphoma cell lines. Lymphoma xenograft studies with Raji-xenograft mice were used to assess the toxicity and efficacy of HB22.7 alone and with combined modality immunotherapy (CMIT) with yttrium (90)Y-DOTA-peptide-Lym-1 radioimmunotherapy (RIT). The effect of the sequence of these agents on the combined treatment was assessed by administering HB22.7 24 hours before, simultaneously with, or 24 hours after RIT. Within the groups treated with RIT alone or with RIT and HB22.7 (CMIT), the reduction in tumor volume was the greatest when HB22.7 was administered simultaneously with and 24 hours after RIT, and in the RIT treatment groups, this translated into the greatest overall response and survival, respectively. Overall survival rates at the end of the 84-day CMIT trial were 67% and 50% in the groups treated with HB22.7 simultaneously and 24 hours after RIT, respectively. This compared favorably with the untreated and the RIT alone groups, which had survival rates of 38% and 43% at the end of the trial. Surprisingly, when compared with untreated controls and all other treatment groups, the greatest cure and overall survival rates were observed in the group treated with HB22.7 alone, with 47% cured and 76% surviving at the end of the 84-day trial. RIT clearance was not affected by treatment with HB22.7. When compared with RIT alone, there was no significant additional hematologic (white blood cell, red blood cell, or platelet count) toxicity when HB22.7 was added to RIT. Nonhematologic toxicity (assessed as change in body weight) was also unchanged when HB22.7 was added to RIT. Thus the anti-CD22 ligand-blocking antibody HB22.7 has independent lymphomacidal properties and augments the efficacy of (90)Y-DOTA-peptide-Lym-1 in lymphoma xenografts without significant toxicity.
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Anticuerpos Monoclonales/uso terapéutico , Antígenos CD/inmunología , Antígenos de Diferenciación de Linfocitos B/inmunología , Antígenos de Neoplasias/inmunología , Antineoplásicos/uso terapéutico , Linfoma de Burkitt/terapia , Moléculas de Adhesión Celular , Inmunización Pasiva , Inmunoconjugados/uso terapéutico , Lectinas/inmunología , Radioinmunoterapia , Animales , Anticuerpos Monoclonales/inmunología , Anticuerpos Monoclonales/farmacocinética , Anticuerpos Monoclonales/farmacología , Anticuerpos Monoclonales de Origen Murino , Apoptosis , Linfoma de Burkitt/radioterapia , Sinergismo Farmacológico , Femenino , Humanos , Inmunoconjugados/farmacocinética , Ratones , Ratones Desnudos , Lectina 2 Similar a Ig de Unión al Ácido Siálico , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
BACKGROUND: Single-agent radioimmunotherapy (RIT), although potentially useful for slowing solid tumor growth, has not been effective in curing aggressive tumors, such as breast cancer. These cancers typically have p53 mutations and are less susceptible to apoptosis, the apparent mechanism of cell death from low dose-rate radiation. Thus, synergistic or combined modality radioimmunotherapy (CMRIT) agents are needed to increase radiosensitivity for therapeutic enhancement without additive toxicity. METHODS: To assess synergy in CMRIT in a breast cancer xenograft model, we evaluated RGD peptide EMD 121974, an inhibitor of alpha(v)beta(3) integrin; paclitaxel, an antimicrotubule; IMC-C225, a monoclonal antibody to epidermal growth factor receptor (EGFR); and bcl-2 antisense oligonucleotide G3139. Groups of mice received (90)Y-DOTA-ChL6 in combination with each agent. Tumor size, survival, and blood counts were monitored for efficacy and toxicity. Immunopathologic evaluation of apoptosis was performed at selected time points after RIT and RIT + RGD CMRIT. RESULTS: CMRIT with RGD peptide increased apoptosis and resulted in 57% cures, compared with 0% cures with RIT alone. CMRIT with paclitaxel after RIT increased cures to 88%, compared with 25% cures with RIT before paclitaxel administration. CMRIT with IMC-C225 resulted in up to 20% cures if given before RIT. A time-dependent increase in toxicity was observed with IMC-C225 after RIT. CMRIT with bcl-2 antisense G3139 resulted in no cures and an increased rate of regrowth compared with RIT alone. CONCLUSIONS: Some combined modality therapies resulted in higher numbers of cures, while others decreased cures and responses and increased toxicity compared with RIT alone. These results support the potential for CMRIT but illustrate the complexity of predicting the efficacy and toxicity and the importance of the relationship between dose and sequence of administration.