RESUMEN

The enzyme indoleamine 2,3 dioxygenase 1 (IDO1) catalyzes the rate-limiting step in the kynurenine pathway (KP) which produces both neuroprotective and neurotoxic metabolites. Neuroinflammatory signals produced as a result of pathological conditions can increase production of IDO1 and boost its enzymatic capacity. IDO1 and the KP have been implicated in behavioral recovery after human traumatic brain injury (TBI), but their roles in experimental models of TBI are for the most part unknown. We hypothesized there is an increase in KP activity in the fluid percussion injury (FPI) model of TBI, and that administration of an IDO1 inhibitor will improve neurological recovery. In this study, adult male Sprague Dawley rats were subjected to FPI or sham injury and received twice-daily oral administration of the IDO1 inhibitor PF-06840003 (100 mg/kg) or vehicle control. FPI resulted in a significant increase in KP activity, as demonstrated by an increased ratio of kynurenine: tryptophan, in the perilesional neocortex and ipsilateral hippocampus 3 days postinjury (DPI), which normalized by 7 DPI. The increase in KP activity was prevented by PF-06840003. IDO1 inhibition also improved memory performance as assessed in the Barnes maze and anxiety behaviors as assessed in open field testing in the first 28 DPI. These results suggest increased KP activity after FPI may mediate neurological dysfunction, and IDO1 inhibition should be further investigated as a potential therapeutic target to improve recovery.

Asunto(s)

Lesiones Traumáticas del Encéfalo , Indolamina-Pirrol 2,3,-Dioxigenasa , Masculino , Animales , Ratas Sprague-Dawley , Indolamina-Pirrol 2,3,-Dioxigenasa/antagonistas & inhibidores , Indolamina-Pirrol 2,3,-Dioxigenasa/metabolismo , Lesiones Traumáticas del Encéfalo/tratamiento farmacológico , Lesiones Traumáticas del Encéfalo/enzimología , Quinurenina/metabolismo , Modelos Animales de Enfermedad , Indoles/administración & dosificación , Succinimidas/administración & dosificación , Administración Oral

RESUMEN

Postoperative adhesion not only causes severe complications for patients but also increases their economic burden. Injectable bioadhesives with adhesiveness to tissues can cover irregular wounds and stay stable in situ, which is a promising barrier for antiadhesion. However, the potential tissue adhesion caused by bioadhesives' indiscriminate adhesiveness between normal and wounded tissue is still a problem. Herein, by using poly(ethylene glycol) succinimidyl succinate (PEG-SS) and gelatin, a succinyl ester-based bioadhesive (SEgel) was fabricated with self-deactivating properties for postoperative antiadhesion. Because N-hydroxysuccinimide esters (NHS-esters) were used as the adhesive group, the bioadhesives' side in contact with the tissue built covalent anchors quickly to maintain the stability, but the superficial layer facing outward withstood fast hydrolysis and then lost its adhesion within minutes, avoiding the indiscriminate adhesiveness. In addition, because of the specific degradation behavior of succinyl ester, the SEgel with proper in vivo retention was achieved without the worry of causing foreign body reactions and unexpected tissue adhesion. Both the cecum-sidewall adhesion and hepatic adhesion models showed that the SEgel markedly reduced the severity of tissue adhesion. These results, together with the ease of the preparation process and well-proven biocompatibility of raw materials, revealed that the SEgel might be a promising solution for postoperative antiadhesion.

Asunto(s)

Materiales Biocompatibles/farmacología , Ésteres/farmacología , Polietilenglicoles/farmacología , Succinimidas/farmacología , Adherencias Tisulares/tratamiento farmacológico , Adhesivos Tisulares/farmacología , Animales , Materiales Biocompatibles/administración & dosificación , Materiales Biocompatibles/química , Ésteres/administración & dosificación , Ésteres/química , Ensayo de Materiales , Ratones , Estructura Molecular , Polietilenglicoles/administración & dosificación , Polietilenglicoles/química , Succinimidas/administración & dosificación , Succinimidas/química , Adhesivos Tisulares/administración & dosificación , Adhesivos Tisulares/química

RESUMEN

Successful imaging of atherosclerosis, one of the leading global causes of death, is crucial for diagnosis and intervention. Near-infrared fluorescence (NIRF) imaging has been widely adopted along with multimodal/hybrid imaging systems for plaque detection. We evaluate two macrophage-targeting fluorescent tracers for NIRF imaging (TLR4-ZW800-1C and Feraheme-Alexa Fluor 750) in an atherosclerotic murine cohort, where the left carotid artery (LCA) is ligated to cause stenosis, and the right carotid artery (RCA) is used as a control. Imaging performed on dissected tissues revealed that both tracers had high uptake in the diseased vessel compared to the control, which was readily visible even at short exposure times. In addition, ZW800-1C's renal clearance ability and Feraheme's FDA approval puts these two tracers in line with other NIRF tracers such as ICG. Continued investigation with these tracers using intravascular NIRF imaging and larger animal models is warranted for clinical translation.

Asunto(s)

Enfermedades de las Arterias Carótidas/diagnóstico por imagen , Placa Aterosclerótica/diagnóstico por imagen , Compuestos de Amonio Cuaternario/administración & dosificación , Succinimidas/administración & dosificación , Ácidos Sulfónicos/administración & dosificación , Animales , Enfermedades de las Arterias Carótidas/inducido químicamente , Enfermedades de las Arterias Carótidas/metabolismo , Dieta Alta en Grasa/efectos adversos , Modelos Animales de Enfermedad , Óxido Ferrosoférrico/química , Humanos , Macrófagos/metabolismo , Masculino , Ratones , Imagen Molecular , Imagen Óptica , Placa Aterosclerótica/inducido químicamente , Placa Aterosclerótica/metabolismo , Compuestos de Amonio Cuaternario/química , Compuestos de Amonio Cuaternario/farmacocinética , Succinimidas/química , Succinimidas/farmacocinética , Ácidos Sulfónicos/química , Ácidos Sulfónicos/farmacocinética , Receptor Toll-Like 4/metabolismo

RESUMEN

Background PF-06840003 is a highly selective indoleamine 2, 3-dioxygenase (IDO1) inhibitor with antitumor effects in preclinical models. This first-in-human phase 1 study evaluated safety, pharmacokinetics/pharmacodynamics, and preliminary efficacy in recurrent malignant glioma to determine the maximum tolerated dose (MTD) or recommended phase 2 dose (RP2D). Methods Patients (N = 17) received oral PF-06840003 in four dose-escalation groups: 125 mg once-daily (QD; n = 2); 250 mg QD (n = 4); 250 mg twice-daily (BID; n = 3); 500 mg BID (n = 8). A modified toxicity probability interval method determined the MTD. Results Four patients experienced serious adverse events (SAEs); one with treatment-related SAEs (grade 4 alanine and aspartate aminotransferase elevations). The dose-limiting toxicity (DLT) rate at 500 mg BID was 12.5% (n = 1/8); the MTD was not reached. Following PF-06840003 dosing, median time to maximum plasma concentration for the active enantiomer PF-06840002 was 1.5-3.0 hr and mean elimination half-life was 2 to 4 hr (Cycle 1 Day 1). Urinary recovery of PF-06840002 was low (< 1%). At 500 mg BID, maximum mean percentage inhibition of 13C10 kynurenine vs endogenous kynurenine was 75% vs 24%. PF-06840002 CSF-to-plasma ratio was 1.00. Disease control occurred in eight patients (47%). Mean duration of stable disease (SD) was 32.1 (12.1-72.3) weeks. Two patients with SD discontinued the study at 450 and 561 days and continued PF-06840003 on compassionate use. Conclusion PF­06840003 up to 500 mg BID was generally well tolerated with evidence of a pharmacodynamic effect and durable clinical benefit in a subset of patients with recurrent malignant glioma. ClinicalTrials.gov, NCT02764151, registered April 2016.

Asunto(s)

Antineoplásicos/administración & dosificación , Neoplasias del Sistema Nervioso Central/tratamiento farmacológico , Glioma/tratamiento farmacológico , Indolamina-Pirrol 2,3,-Dioxigenasa/antagonistas & inhibidores , Indoles/administración & dosificación , Recurrencia Local de Neoplasia/tratamiento farmacológico , Succinimidas/administración & dosificación , Adolescente , Adulto , Anciano , Antineoplásicos/efectos adversos , Antineoplásicos/farmacocinética , Femenino , Humanos , Indoles/efectos adversos , Indoles/farmacocinética , Quinurenina/metabolismo , Masculino , Persona de Mediana Edad , Succinimidas/efectos adversos , Succinimidas/farmacocinética , Resultado del Tratamiento , Triptófano/metabolismo , Adulto Joven

RESUMEN

Breast cancer is the most common type of cancer in women worldwide. There have been many efforts for early breast cancer detection and among them molecular imaging have been extremely of high importance. Single-photon emission computed tomography (SPECT/CT) is a kind of imaging technique able to reveal crucial information with using radiopharmaceuticals. In this study, Technetium-99m-(DOTA-NHS-ester)-Methionine radiopharmaceutical was synthesized. Between 1,4,7,10-Tetraazacyclododecane-1,4,7,10-tetraacetic acid mono-N-hydroxysuccinimide ester (DOTA-HNS ester) (MACROCYCLICS, DOTA-NHS ester, Plano, Texas, USA) and methionine(marker) were conjugated. The DOTA-HNS ester-Methionine was labeled with Technetium-99m (Inter-Medical, Technetium-99m, Bergamo, Italy). The synthesized radiopharmaceutical was used in SPECT/CT imaging for breast cancer diagnosis. For radiopharmaceutical evaluation, MTT assay for cellular toxicity, biodistribution, cellular uptake and radiochemical purity were employed.Technetium-99m-(DOTA-NHS-ester)-Methionine radiochemical had less cellular toxicity in human embryonic kidney cells 293 cell line (HEK293). Cellular uptake was indicated higher percent with use of Methionine as a marker, and radiochemical purity was high. Based on the results Technetium-99m-(DOTA-NHS-ester)-Methionine radiochem may be a better option for early detection of breast cancer. Further study is recommended to confirm these findings in clinical practice.

Asunto(s)

Medios de Contraste/administración & dosificación , Metionina/administración & dosificación , Radiofármacos/administración & dosificación , Succinimidas/administración & dosificación , Tecnecio/administración & dosificación , Animales , Transporte Biológico , Supervivencia Celular/efectos de los fármacos , Medios de Contraste/farmacocinética , Ésteres , Células HEK293 , Humanos , Células MCF-7 , Masculino , Metionina/farmacocinética , Ratones , Neoplasias/metabolismo , Radiofármacos/farmacocinética , Succinimidas/farmacocinética , Tecnecio/farmacocinética , Distribución Tisular , Tomografía Computarizada de Emisión de Fotón Único

RESUMEN

BACKGROUND: The use of regional and other opioid-sparing forms of anesthesia has been associated with a decrease in the recurrence of certain malignancies. Direct suppression of human natural killer cells by opioids has been postulated to explain this observation. However, the effect of different classes of opioids on suppression of natural killer cell cytotoxicity has not been systematically characterized. METHODS: After confirming that freshly isolated natural killer cells from peripheral human blood express opioid receptors, cells were incubated with increasing concentrations of clinically used or receptor-specific opioid agonists. We also evaluated the effect of pretreatment with receptor-specific antagonists or naloxone. Treated natural killer cells were then coincubated with a carboxyfluorescein succinimidyl ester-labeled target tumor cell line, K562. Annexin V staining was used to compare the percent of tumor cell apoptosis in the presence of opioid-pretreated and untreated natural killer cells. Treated samples were compared to untreated samples using Kruskal-Wallis tests with a post hoc Dunn correction. RESULTS: Morphine, methadone, buprenorphine, loperamide, [D-Ala2, N-MePhe4, Gly-ol]-enkephalin, and U-50488 significantly decreased natural killer cell cytotoxicity. When natural killer cells were pretreated with naloxone, cyprodime, and nor-binaltorphimine before exposure to morphine, there was no difference in natural killer cytotoxicity, compared to the amount observed by untreated natural killer cells. Fentanyl, O-desmethyltramadol, and [D-Pen2,D-Pen5] enkephalin did not change natural killer cell cytotoxicity compare to untreated natural killer cells. CONCLUSIONS: Incubation of isolated natural killer cells with certain opioids causes a decrease in activity that is not observed after naloxone pretreatment. Suppression of natural killer cell cytotoxicity was observed with µ- and κ-receptor agonists but not δ-receptor agonists. These data suggest that the effect is mediated by µ- and κ-receptor agonism and that suppression is similar with many clinically used opioids.

Asunto(s)

Analgésicos Opioides/administración & dosificación , Células Asesinas Naturales/efectos de los fármacos , 3,4-Dicloro-N-metil-N-(2-(1-pirrolidinil)-ciclohexil)-bencenacetamida, (trans)-Isómero/administración & dosificación , Anestesia , Buprenorfina/administración & dosificación , Encefalina D-Penicilamina (2,5)/administración & dosificación , Fentanilo/administración & dosificación , Fluoresceínas/administración & dosificación , Humanos , Terapia de Inmunosupresión , Células K562 , Loperamida/administración & dosificación , Metadona/administración & dosificación , Morfinanos/administración & dosificación , Morfina/administración & dosificación , Naloxona/administración & dosificación , Naltrexona/administración & dosificación , Naltrexona/análogos & derivados , Succinimidas/administración & dosificación , Receptor Toll-Like 4/metabolismo , Tramadol/administración & dosificación , Tramadol/análogos & derivados

RESUMEN

The tracing of neuronal cell lineages is critical to our understanding of cellular diversity in the CNS. This protocol describes a fluorescence birth-dating technique to label, track and isolate isochronic cohorts of newborn cells in the CNS in vivo in mouse embryos. Injection of carboxyfluorescein esters (CFSEs) into the cerebral ventricle allows pulse labeling of mitotic (M phase) ventricular zone (VZ) progenitors and their progeny across the CNS, a procedure we termed FlashTag. Specificity for M-phase apical progenitors is a result of the somata of these cells transiently contacting the ventricular wall during this cell-cycle phase, exposing them to CFSE injected into the cerebrospinal fluid. Using this approach, the developmental trajectory of progenitors and their daughter neurons can be tracked. Labeled cells can be imaged ex vivo or in fixed tissue, targeted for electrophysiological experiments or isolated using FACS for cell culture or (single-cell) RNA sequencing. Multiple embryos can be labeled within 30 min. The dye is retained for several weeks, allowing labeled cells to be identified postnatally. This protocol describes the labeling procedure using in utero injection, the isolation of live cells using FACS and the processing of labeled tissue for immunohistochemistry.

Asunto(s)

Sistema Nervioso Central/citología , Embrión de Mamíferos/citología , Fluoresceínas/análisis , Colorantes Fluorescentes/análisis , Células-Madre Neurales/citología , Neuronas/citología , Succinimidas/análisis , Animales , División Celular , Rastreo Celular/métodos , Embrión de Mamíferos/ultraestructura , Citometría de Flujo/métodos , Fluoresceínas/administración & dosificación , Colorantes Fluorescentes/administración & dosificación , Ratones , Mitosis , Succinimidas/administración & dosificación

RESUMEN

Targeted nano-delivery vehicles were developed from genetically modified Cowpea chlorotic mottle virus (CCMV) capsid by ligands bioconjugation for efficient drug delivery in cancer cells. RNA binding (N 1-25aa) and ß-hexamer forming (N 27-41aa) domain of capsid was selectively deleted by genetic engineering to achieve the efficient in vitro assembly without natural cargo. Two variants of capsids were generated by truncating 41 and 26 amino acid from N terminus (NΔ41 and NΔ26) designated as F1 and F2 respectively. These capsid were optimally self-assembled in 1:2 molar ratio (F1:F2) to form a monodisperse nano-scaffold of size 28 nm along with chemically conjugated modalities for visualization (fluorescent dye), targeting (folic acid, FA) and anticancer drug (doxorubicin). The cavity of the nano-scaffold was packed with doxorubicin conjugated gold nanoparticles (10 nm) to enhance the stability, drug loading and sustained release of drug. The chimeric system was stable at pH range of 4-8. This chimeric nano-scaffold system showed highly specific receptor mediated internalization (targeting) and ~300% more cytotoxicity (with respect to FA- delivery system) to folate receptor positive Michigan Cancer Foundation-7 (MCF7) cell lines. The present system may offer a programmable nano-scaffold based platform for developing chemotherapeutics for cancer.

Asunto(s)

Antibióticos Antineoplásicos/farmacología , Proteínas de la Cápside/química , Proteínas de la Cápside/genética , Citotoxinas/farmacología , Doxorrubicina/farmacología , Sistemas de Liberación de Medicamentos/métodos , Bromovirus/genética , Liberación de Fármacos , Fluoresceínas/administración & dosificación , Fluoresceínas/química , Receptor 1 de Folato/metabolismo , Humanos , Ligandos , Células MCF-7 , Nanopartículas del Metal/administración & dosificación , Nanopartículas del Metal/química , Succinimidas/administración & dosificación , Succinimidas/química

RESUMEN

The ubiquitin-proteasome system plays a pivotal role in breast tumorigenesis by controlling transcription factors, thus promoting cell cycle growth, and degradation of tumor suppressor proteins. However, breast cancer patients have failed to benefit from proteasome inhibitor treatment partially due to proteasome heterogeneity, which is poorly understood in malignant breast neoplasm. Chemical crosslinking is an increasingly important tool for mapping protein three-dimensional structures and proteinprotein interactions. In the present study, two cross-linkers, bis (sulfosuccinimidyl) suberate (BS(3)) and its water-insoluble analog disuccinimidyl suberate (DSS), were used to map the subunit-subunit interactions in 20S proteasome core particle (CP) from MDA-MB-231 cells. Different types of gel electrophoresis technologies were used. In combination with chemical cross-linking and mass spectrometry, we applied these gel electrophoresis technologies to the study of the noncovalent interactions among 20S proteasome subunits. Firstly, the CP subunit isoforms were profiled. Subsequently, using native/SDSPAGE, it was observed that 0.5 mmol/L BS(3) was a relatively optimal cross-linking concentration for CP subunit-subunit interaction study. 2-DE analysis of the cross-linked CP revealed that α1 might preinteract with α2, and α3 might pre-interact with α4. Moreover, there were different subtypes of α1α2 and α3α4 due to proteasome heterogeneity. There was no significant difference in cross-linking pattern for CP subunits between BS(3) and DSS. Taken together, the gel-based characterization in combination with chemical cross-linking could serve as a tool for the study of subunit interactions within a multi-subunit protein complex. The heterogeneity of 20S proteasome subunit observed in breast cancer cells may provide some key information for proteasome inhibition strategy.

Asunto(s)

Neoplasias de la Mama/tratamiento farmacológico , Reactivos de Enlaces Cruzados/administración & dosificación , Complejo de la Endopetidasa Proteasomal/efectos de los fármacos , Succinimidas/administración & dosificación , Secuencia de Aminoácidos , Neoplasias de la Mama/genética , Neoplasias de la Mama/patología , Línea Celular Tumoral , Electroforesis en Gel Bidimensional , Femenino , Humanos , Espectrometría de Masas , Unión Proteica/efectos de los fármacos , Isoformas de Proteínas/genética , Subunidades de Proteína/biosíntesis , Subunidades de Proteína/genética , Proteómica

RESUMEN

A series of ß-cyclodextrin-grafted carboxymethyl chitosan hydrogels (CD-g-CMCs) were prepared from carboxymethyl chitosan (CMC) and carboxymethyl ß-chitosan (CMCD) using a water-soluble carbodiimide as a crosslinker in the presence of N-hydroxysuccinimide. Details of the hydrogel structures were determined via FTIR and solid-state NMR spectroscopic analyses. Increasing the feed ratio of CMCD to CMC in the reaction mixture led to an increase in CD grafting within the gel networks comprising CMC; this was confirmed by SEM observations and rheological analysis of the swollen hydrogels. The prepared CD-g-CMC hydrogels exhibited absorption properties toward acetylsalicylic acid (ASA, or Aspirin) due to the presence of CD in the structure; the amount of ASA absorbed into the hydrogels was enhanced with an increase in the amount of CD incorporated within the hydrogels. In addition, CD-g-CMC hydrogels provided a slower release of the entrapped ASA in comparison to the ASA release profile of a solely CMC-containing hydrogel. From these results, CD-g-CMC hydrogels have the potential to function as a biodegradable active material with controlled drug release ability.

Asunto(s)

Quitosano/química , Ciclodextrinas/química , Sistemas de Liberación de Medicamentos , Hidrogeles/química , Aspirina/administración & dosificación , Aspirina/química , CME-Carbodiimida/química , Quitosano/administración & dosificación , Ciclodextrinas/administración & dosificación , Liberación de Fármacos , Humanos , Hidrogeles/administración & dosificación , Reología , Succinimidas/administración & dosificación , Succinimidas/química

RESUMEN

Administration of drugs via inhalation is an attractive route for pulmonary and systemic drug delivery. The therapeutic outcome of inhalation therapy depends not only on the dose of the lung-delivered drug, but also on its bioactivity and regional distribution. Fluorescence imaging has the potential to monitor these aspects already during preclinical development of inhaled drugs, but quantitative methods of analysis are lacking. In this proof-of-concept study, we demonstrate that Cryoslicing Imaging allows for 3D quantitative fluorescence imaging on ex vivo murine lungs. Known amounts of fluorescent substance (nanoparticles or fluorophore-drug conjugate) were instilled in the lungs of mice. The excised lungs were measured by Cryoslicing Imaging. Herein, white light and fluorescence images are obtained from the face of a gradually sliced frozen organ block. A quantitative representation of the fluorescence intensity throughout the lung was inferred from the images by accounting for instrument noise, tissue autofluorescence and out-of-plane fluorescence. Importantly, the out-of-plane fluorescence correction is based on the experimentally determined effective light attenuation coefficient of frozen murine lung tissue (10.0 ± 0.6 cm(-1) at 716 nm). The linear correlation between pulmonary total fluorescence intensity and pulmonary fluorophore dose indicates the validity of this method and allows direct fluorophore dose assessment. The pulmonary dose of a fluorescence-labeled drug (FcγR-Alexa750) could be assessed with an estimated accuracy of 9% and the limit of detection in ng regime. Hence, Cryoslicing Imaging can be used for quantitative assessment of dose and 3D distribution of fluorescence-labeled drugs or drug carriers in the lungs of mice.

Asunto(s)

Pulmón/química , Succinimidas/análisis , Administración por Inhalación , Animales , Crioultramicrotomía , Relación Dosis-Respuesta a Droga , Femenino , Humanos , Masculino , Ratones , Microscopía Fluorescente , Nanopartículas/administración & dosificación , Receptores de IgG/administración & dosificación , Receptores de IgG/análisis , Succinimidas/administración & dosificación

RESUMEN

RATIONALE: To determine the role of the endocannabinoid, 2-arachodonyl glycerol (2-AG), in the regulation of nausea and vomiting. OBJECTIVE: We evaluated the effectiveness of the potent selective monoacylglycerol lipase (MAGL) inhibitor, MJN110, which selectively elevates the endocannabinoid 2-AG, to suppress acute nausea and vomiting, as well as anticipatory nausea in rat and shrew models. METHODS: The rat gaping models were used to evaluate the potential of MJN110 (5, 10, and 20 mg/kg, intraperitoneally [IP]) to suppress acute nausea produced by LiCl and of MJN110 (10 and 20 mg/kg, IP) to suppress anticipatory nausea elicited by a LiCl-paired context. The potential as well of MJN110 (10 and 20 mg/kg, IP) to suppress vomiting and contextually elicited gaping in the Suncus murinus was evaluated. RESULTS: MJN110 suppressed acute nausea in rats, LiCl-induced vomiting in shrews and contextually-elicited anticipatory nausea in both rats (accompanied by elevation of 2-AG in the visceral insular cortex) and shrews. These effects were reversed by the CB1 antagonist/inverse agonist, SR141716. The MAGL inhibitor did not modify locomotion at any dose. An activity-based protein profiling analysis of samples of tissue collected from the visceral insular cortex in rats and whole brain tissues in shrews revealed that MJN110 selectively inhibited MAGL and the alternative 2-AG hydrolase, ABHD6. CONCLUSIONS: MAGL inhibition by MJN110 which selectively elevates endogenous 2-AG has therapeutic potential in the treatment of acute nausea and vomiting as well as anticipatory nausea, a distressful symptom that is resistant to currently available treatments.

Asunto(s)

Carbamatos/farmacología , Monoacilglicerol Lipasas/antagonistas & inhibidores , Náusea/tratamiento farmacológico , Succinimidas/farmacología , Vómitos/tratamiento farmacológico , Animales , Carbamatos/administración & dosificación , Modelos Animales de Enfermedad , Femenino , Masculino , Ratas , Ratas Sprague-Dawley , Musarañas , Succinimidas/administración & dosificación

RESUMEN

We developed a new robust reduction-responsive polymersome based on the amphiphilic block copolymer PEG-SS-PAChol. The stability and robustness were achieved by the smectic physical cross-linking of cholesterol-containing liquid crystal polymer PAChol in the hydrophobic layer. The reduction-sensitivity was introduced by the disulfide bridge (-S-S-) that links the hydrophilic PEG block and the hydrophobic PAChol block. We used a versatile synthetic strategy based on atom transfer radical polymerization (ATRP) to synthesize the reduction-responsive amphiphilic block copolymers. The reductive cleavage of the disulfide bridge in the block copolymers was first evidenced in organic solution. The partial destruction of PEG-SS-PAChol polymersomes in the presence of a reducing agent was then demonstrated by cryo-electron microscopy. Finally, the calcein release from PEG-SS-PAChol polymersomes triggered by glutathione (GSH) was observed both in PBS suspension and in vitro inside the macrophage cells. High GSH concentrations (≥35 mM in PBS or artificially enhanced in macrophage cells by GSH-OEt pretreatment) and long incubation time (in the order of hours) were, however, necessary to get significant calcein release. These polymersomes could be used as drug carriers with very long circulation profiles and slow release kinetics.

Asunto(s)

Colesterol/química , Portadores de Fármacos/química , Sistemas de Liberación de Medicamentos/métodos , Polietilenglicoles/química , Succinimidas/química , Animales , Línea Celular , Colesterol/administración & dosificación , Portadores de Fármacos/administración & dosificación , Macrófagos/efectos de los fármacos , Ratones , Polietilenglicoles/administración & dosificación , Succinimidas/administración & dosificación

RESUMEN

BACKGROUND: The aim of this study was to determine the effects of N-(morpholinomethyl)-p-isopropoxy-phenylsuccinimide (MMIPPS) on the protective action of four classical antiepileptic drugs (AEDs: carbamazepine [CBZ], phenobarbital [PB], phenytoin [PHT] and valproate [VPA]) against maximal electroshock (MES)-induced seizures in mice. METHODS: Tonic hind limb extension (seizure activity) was evoked in adult male albino Swiss mice by a current (sine-wave, 25 mA, 500 V, 50 Hz, 0.2 s stimulus duration) delivered via auricular electrodes. Total brain concentrations of AEDs were measured to determine the characteristics of interaction between MMIPPS and classical AEDs in the mouse MES model. RESULTS: MMIPPS administered intraperitoneally (ip) at 100 mg/kg significantly elevated the threshold for electroconvulsions in mice (p < 0.01). MMIPPS at doses of 25 and 50 mg/kg had no impact on the threshold for electroconvulsions in mice. Moreover, MMIPPS (50 mg/kg) significantly enhanced the anticonvulsant activity of PB and VPA (p < 0.05), but not that of CBZ or PHT, in the MES test in mice. Pharmacokinetic studies revealed that MMIPPS (50 mg/kg) did not alter total brain concentrations of PB, but significantly elevated total brain concentrations of VPA in mice (p < 0.05). CONCLUSIONS: The enhanced anticonvulsant action of PB by MMIPPS in the mouse MES model and lack of any pharmacokinetic interaction between drugs make the combination of MMIPPS with PB of pivotal importance for further experimental and clinical studies. Pharmacokinetic increase in total brain VPA concentration seems to be responsible for the enhanced anticonvulsant action of VPA by MMIPPS in the mouse MES model. The combinations of MMIPPS with CBZ and PHT are neutral from a preclinical viewpoint.

Asunto(s)

Anticonvulsivantes/farmacología , Encéfalo/metabolismo , Morfolinas/farmacología , Convulsiones/prevención & control , Succinimidas/farmacología , Animales , Anticonvulsivantes/administración & dosificación , Anticonvulsivantes/farmacocinética , Modelos Animales de Enfermedad , Relación Dosis-Respuesta a Droga , Interacciones Farmacológicas , Electrochoque , Inyecciones Intraperitoneales , Masculino , Ratones , Morfolinas/administración & dosificación , Succinimidas/administración & dosificación , Distribución Tisular

RESUMEN

Block copolymer of poly(ethylene glycol)-block-poly{N-[N-(2-aminoethyl)-2-aminoethyl]aspartamide} (PEG-P[Asp(DET)]) has been originally introduced as a promising gene carrier by forming a nanomicelle with plasmid DNA. In this study, the polyplex micelle of PEG-SS-P[Asp(DET)], which disulfide linkage (SS) between PEG and cationic polymer can detach the surrounding PEG chains upon intracellular reduction, was firstly evaluated with respect to in vivo transduction efficiency and toxicity in comparison to that of PEG-P[Asp(DET)] in peritoneally disseminated cancer model. Intraperitoneal (i.p.) administration of PEG-SS-P[Asp(DET)] polyplex micelles showed a higher (P<0.05) transgene expression compared with PEG-P[Asp(DET)] in tumors. In contrast, the delivered distribution of the micelles was not different between the two polyplex micelles. PEG-SS-P[Asp(DET)] micelle encapsulating human tumor necrosis factor α (hTNF-α) gene exhibits a higher antitumor efficacy against disseminated cancer compared with PEG-P[Asp(DET)] or saline control. No hepatic and renal toxicities were observed by the administration of polyplex micelles. In conclusion, PEG-detachable polyplex micelles may represent an advantage in gene transduction in vivo over PEG-undetachable polyplex micelles after i.p. administration for peritoneal dissemination of cancer.

Asunto(s)

Neoplasias/terapia , Polietilenglicoles/administración & dosificación , Proteínas Recombinantes de Fusión/administración & dosificación , Proteínas Recombinantes de Fusión/genética , Succinimidas/administración & dosificación , Animales , Línea Celular Tumoral , Femenino , Terapia Genética/métodos , Humanos , Inyecciones Intraperitoneales , Hígado/metabolismo , Luciferasas/genética , Luciferasas/metabolismo , Ratones , Ratones Endogámicos BALB C , Micelas , Trasplante de Neoplasias , Neoplasias/metabolismo , Polietilenglicoles/química , Proteínas Recombinantes de Fusión/metabolismo , Bazo/metabolismo , Succinimidas/química , Transgenes/genética

RESUMEN

Lipid-based oil-filled nanoparticles (NPs) with a high concentration of surface-chelated nickel (Ni-NPs) were successfully prepared using a Brij 78-NTA-Ni conjugate synthesized with Brij 78 (Polyoxyethylene (20) stearyl ether) and nitrilotriacetic acid (NTA). The facile incorporation of the Brij 78-NTA-Ni conjugate into the NP formulation allowed up to 90% Ni incorporation, which was a significant improvement over the previously used standard agent DOGS-NTA-Ni which led to ~6% Ni incorporation. The Ni-NPs were targeted to the highly epidermal growth factor receptor (EGFR)-overexpressing epidermoid carcinoma cells A431. This was accomplished using a novel high affinity histidine×6-tagged EGFR-binding Z domain (heptameric Z(EGFR) domain). In vitro cell uptake studies showed enhanced internalization (up to 90%) of the targeted Ni-NPs in A431 cells with only ≤10% internalization of the untargeted Ni-NPs. ICP-MS analysis used to quantify the amount of Ni in the cells were in close agreement with flow cytometry studies, which showed a dose dependent increase in the amount of Ni with the targeted Ni-NPs. Cell uptake competition studies showed that internalization of the targeted Ni-NPs within the cells was competed off with free heptameric Z(EGFR) domain at concentrations of 8.75ng/mL or higher. In vivo studies were carried out in nude mice bearing A431 tumors to determine the biodistribution and intracellular delivery. Near Infrared (NIR) optical imaging studies using Alexa750-labeled heptameric Z(EGFR) domain showed localization of 19% of the total detected fluorescence intensity in the tumor tissue, 28% in the liver and 42% in the kidneys 16h post i.v. injection. ICP-MS analysis showed almost a two-fold increase in the amount of intracellular Ni with the targeted Ni-NPs. These new Ni-NPs could be a very useful tool for targeting and drug delivery to a wide range of EGFR positive cancers.

Asunto(s)

Portadores de Fármacos/administración & dosificación , Receptores ErbB/metabolismo , Nanopartículas del Metal/administración & dosificación , Neoplasias/metabolismo , Níquel/administración & dosificación , Animales , Línea Celular Tumoral , Portadores de Fármacos/química , Receptores ErbB/química , Humanos , Nanopartículas del Metal/química , Ratones , Ratones Desnudos , Níquel/química , Ácido Nitrilotriacético/administración & dosificación , Ácido Nitrilotriacético/química , Polietilenglicoles/administración & dosificación , Polietilenglicoles/química , Estructura Terciaria de Proteína , Succinimidas/administración & dosificación , Succinimidas/química , Tensoactivos/administración & dosificación , Tensoactivos/química , Distribución Tisular , Triglicéridos/administración & dosificación , Triglicéridos/química

RESUMEN

The objective of the present study was to evaluate the effect of oleic acid modified polymeric bilayered nanoparticles (NPS) on combined delivery of two anti-inflammatory drugs, spantide II (SP) and ketoprofen (KP) on the skin permeation. NPS were prepared using poly(lactic-co-glycolic acid) (PLGA) and chitosan. SP and KP were encapsulated in different layers alone or/and in combination (KP-NPS, SP-NPS and SP+KP-NPS). The surface of NPS was modified with oleic acid (OA) ('Nanoease' technology) using an established procedure in the laboratory (KP-NPS-OA, SP-NPS-OA and SP+KP-NPS-OA). Fluorescent dyes (DiO and DID) containing surface modified (DiO-NPS-OA and DID-NPS-OA) and unmodified NPS (DiO-NPS and DID-NPS) were visualized in lateral rat skin sections using confocal microscopy and Raman confocal spectroscopy after skin permeation. In vitro skin permeation was performed in dermatomed human skin and HPLC was used to analyze the drug levels in different skin layers. Further, allergic contact dermatitis (ACD) model was used to evaluate the response of KP-NPS, SP-NPS, SP+KP-NPS, KP-NPS-OA, SP-NPS-OA and SP+KP-NPS-OA treatment in C57BL/6 mice. The fluorescence from OA modified NPS was observed up to a depth of 240µm and was significantly higher as compared to non-modified NPS. The amount of SP and KP retained in skin layers from OA modified NPS increased by several folds compared to unmodified NPS and control solution. In addition, the combination index value calculated from ACD response for solution suggested an additive effect and moderate synergism for NPS-OA. Our results strongly suggest that surface modification of bilayered nanoparticles with oleic acid improved drug delivery to the deeper skin layers.

Asunto(s)

Analgésicos/administración & dosificación , Antiinflamatorios no Esteroideos/administración & dosificación , Cetoprofeno/administración & dosificación , Nanopartículas/administración & dosificación , Ácido Oléico/administración & dosificación , Sustancia P/análogos & derivados , Administración Cutánea , Analgésicos/química , Animales , Antiinflamatorios no Esteroideos/química , Dermatitis Alérgica por Contacto/tratamiento farmacológico , Dermatitis Alérgica por Contacto/etiología , Dinitrofluorobenceno , Humanos , Cetoprofeno/química , Ácido Láctico/administración & dosificación , Ácido Láctico/química , Masculino , Ratones , Ratones Endogámicos C57BL , Nanopartículas/química , Ácido Oléico/química , Ácido Poliglicólico/administración & dosificación , Ácido Poliglicólico/química , Copolímero de Ácido Poliláctico-Ácido Poliglicólico , Ratas , Piel/metabolismo , Absorción Cutánea/efectos de los fármacos , Sustancia P/administración & dosificación , Sustancia P/química , Succinimidas/administración & dosificación , Succinimidas/química

RESUMEN

PURPOSE: We investigated the use of hyperthermia to improve the anti-cancer efficacy of doxorubicin (DOX)-loaded mesoporous silica nanocontainer Si-SS-CD-PEG. The hypothesis was that heat stimulates glutathione-mediated degradation of cyclodextrin gatekeeper, thereby causing the release of DOX from the carrier and DOX-induced cell death. MATERIALS AND METHODS: The release of DOX from DOX-loaded Si-SS-CD-PEG suspended in PBS containing glutathione (GSH) was studied by assessing the changes in DOX fluorescence intensity. The effect of heating at 42°C on the release of DOX from the intracellular carriers was determined with confocal microscopy. The extents of clonogenic and apoptotic cell death caused by DOX-loaded Si-SS-CD-PEG were determined. RESULTS: The release of DOX from DOX-loaded Si-SS-CD-PEG in PBS occurred only when GSH presented in the suspension, and heating at 42°C slightly increased the release of DOX from the carriers. Heating significantly elevated the GSH content in A549 cells and increased the release of DOX from the internalised carriers. Heating the cancer cells treated with the carriers at 42°C markedly increased the clonogenic death and apoptosis. The GSH content in A549 cells was greater than that in L-132 cells, and A549 cells were far more sensitive than L-132 cells to DOX-loaded Si-SS-CD-PEG at both 37°C and 42°C. CONCLUSIONS: Hyperthermia increased the GSH-mediated release of DOX from DOX-loaded Si-SS-CD-PEG. Furthermore, hyperthermia markedly elevated the GSH content in cancer cells, thereby increasing the release of DOX from the internalised carriers and potentiating the DOX-induced clonogenic and apoptotic cell death.

Asunto(s)

Adenocarcinoma/tratamiento farmacológico , Antibióticos Antineoplásicos/administración & dosificación , Doxorrubicina/administración & dosificación , Hipertermia Inducida , Neoplasias Pulmonares/tratamiento farmacológico , Nanoestructuras/administración & dosificación , Adenocarcinoma del Pulmón , Línea Celular Tumoral , Ciclodextrinas/administración & dosificación , Portadores de Fármacos , Glutatión/metabolismo , Humanos , Polietilenglicoles/administración & dosificación , Succinimidas/administración & dosificación

RESUMEN

Twenty-two new 1-[2-oxo-2-(4-phenylpiperazin-1-yl)ethyl]pyrrolidine-2,5-diones were synthesized and tested for anticonvulsant activity. Initial anticonvulsant screening was performed using standard maximal electroshock (MES) and subcutaneous pentylenetetrazole (scPTZ) screens in mice. Several compounds were tested additionally in the 6-Hz psychomotor seizure model. The neurotoxicity was determined applying the rotarod test. Excluding one compound, all other molecules were found to be effective in at least one seizure model. The most active were 1-(2-oxo-2-{4-[3-(trifluoromethyl)phenyl]piperazin-1-yl}ethyl)pyrrolidine-2,5-dione (14), 1-{2-[4-(4-chlorophenyl)piperazin-1-yl]-2-oxoethyl}-3-methylpyrrolidine-2,5-dione (17), 1-{2-[4-(4-chlorophenyl)piperazin-1-yl]-2-oxoethyl}-3,3-dimethylpyrrolidine-2,5-dione (23) and 3,3-dimethyl-1-(2-oxo-2-{4-[3-(trifluoromethyl)phenyl]piperazin-1-yl}ethyl)pyrrolidine-2,5-dione (26). These compounds showed high activity in the 6-Hz psychomotor seizure test as well as were active in the maximal electroshock and subcutaneous pentylenetetrazole (14 and 23) screens. Initial SAR studies for anticonvulsant activity have been discussed.

Asunto(s)

Anticonvulsivantes/síntesis química , Anticonvulsivantes/farmacología , Diseño de Fármacos , Piperazinas/síntesis química , Piperazinas/farmacología , Piperazinas/toxicidad , Convulsiones/prevención & control , Succinimidas/síntesis química , Succinimidas/farmacología , Succinimidas/toxicidad , Administración Oral , Animales , Anticonvulsivantes/administración & dosificación , Anticonvulsivantes/química , Convulsivantes , Relación Dosis-Respuesta a Droga , Evaluación Preclínica de Medicamentos , Electrochoque , Etosuximida/farmacología , Inyecciones Intraperitoneales , Ratones , Estructura Molecular , Actividad Motora/efectos de los fármacos , Pentilenotetrazol , Fenitoína/farmacología , Piperazinas/administración & dosificación , Piperazinas/química , Ratas , Convulsiones/tratamiento farmacológico , Relación Estructura-Actividad , Succinimidas/administración & dosificación , Factores de Tiempo

RESUMEN

A comprehensive knowledge of the in vivo fate of polymers is essential for their potential application in humans. In this study, the body distribution, accumulation, and elimination processes of intraperitoneally (ip) administered poly(vinyl alcohol) (PVA) in mice were investigated in detail. Two derivatives of PVA (195 kDa) having covalently bound fluorescent dye labels were synthesized and used to follow PVA in vivo by noninvasive multispectral fluorescence imaging over several months. Detailed ex vivo fluorescence imaging was performed additionally and combined with tissue accumulation studies using confocal microscopy. Filtration and confocal imaging at appropriate synthetic membranes, used as models for glomerular filtration, confirmed a considerable PVA permeation. This investigation yields new scientific findings about the fate of PVA in vivo. PVA accumulated in fat tissue at high levels, which suggests that PVA is suitable not only for abdominal surgeries but also for controlled release applications after ip or subcutaneous injection.

Asunto(s)

Colorantes Fluorescentes/farmacocinética , Imagen Molecular/métodos , Alcohol Polivinílico/farmacocinética , Coloración y Etiquetado/métodos , Succinimidas/farmacocinética , Grasa Abdominal/efectos de los fármacos , Animales , Femenino , Fluorescencia , Colorantes Fluorescentes/administración & dosificación , Colorantes Fluorescentes/química , Tasa de Filtración Glomerular/fisiología , Humanos , Inyecciones Intraperitoneales , Inyecciones Subcutáneas , Riñón/fisiología , Cinética , Masculino , Membranas Artificiales , Ratones , Ratones Desnudos , Microscopía Confocal , Alcohol Polivinílico/administración & dosificación , Alcohol Polivinílico/análogos & derivados , Succinimidas/administración & dosificación , Succinimidas/química , Distribución Tisular