Your browser doesn't support javascript.
loading
Mostrar: 20 | 50 | 100
Resultados 1 - 10 de 10
Filtrar
1.
Pharmaceutics ; 13(12)2021 Nov 26.
Artigo em Inglês | MEDLINE | ID: mdl-34959304

RESUMO

The aim of this study was to get insight into the internalization and transport of PEGylat-ed mixed micelles loaded by vitamin K, as mediated by Scavenger Receptor B1 (SR-B1) that is abundantly expressed by intestinal epithelium cells as well as by differentiated Caco-2 cells. Inhibition of SR-B1 reduced endocytosis and transport of vitamin-K-loaded 0%, 30% and 50% PEGylated mixed micelles and decreased colocalization of the micelles with SR-B1. Confocal fluorescence microscopy, fluorescence-activated cell sorting (FACS) analysis, and surface plasmon resonance (SPR) were used to study the interaction between the mixed micelles of different compositions (varying vitamin K loading and PEG content) and SR-B1. Interaction of PEGylated micelles was independent of the vitamin K content, indicating that the PEG shell prevented vitamin K exposure at the surface of the micelles and binding with the receptor and that the PEG took over the micelles' ability to bind to the receptor. Molecular docking calculations corroborated the dual binding of both vita-min K and PEG with the binding domain of SR-B1. In conclusion, the improved colloidal stability of PEGylated mixed micelles did not compromise their cellular uptake and transport due to the affinity of PEG for SR-B1. SR-B1 is able to interact with PEGylated nanoparticles and mediates their subsequent internalization and transport.

3.
Mol Pharm ; 15(9): 3786-3795, 2018 09 04.
Artigo em Inglês | MEDLINE | ID: mdl-30063364

RESUMO

The aim of the study is to investigate the uptake by and transport through Caco-2 cells of two mixed micelle formulations (based on egg phosphatidylcholine and glycocholic acid) of vitamin K, i.e., with and without DSPE-PEG2000. The uptake of vitamin K and fluorescently labeled mixed micelles with and without PEG coating showed similar kinetics and their uptake ratio remained constant over time. Together with the fact that an inhibitor of scavenger receptor B1 (BLT-1) decreased cellular uptake of vitamin K by ∼80% compared to the uptake in the absence of this inhibitor, we conclude that both types of micelles loaded with vitamin K can be taken up intactly by Caco-2 cells via this scavenger receptor. The amount of vitamin K in chylomicrons fraction from Caco-2 cell monolayers further indicates that mixed micelles (with or without PEGylation) are likely packed into chylomicrons after internalization by Caco-2 cells. Uptake of vitamin K from PEGylated mixed micelles increased four- to five-fold at simulated gastrointestinal conditions. In conclusion, PEGylated mixed micelles are stable upon exposure to simulated gastric conditions, and as a result, they do show overall a higher cellular uptake efficiency of vitamin K as compared to mixed micelles without PEG coating.


Assuntos
Micelas , Fosfatidiletanolaminas/química , Polietilenoglicóis/química , Vitamina K/química , Vitamina K/farmacologia , Transporte Biológico/efeitos dos fármacos , Células CACO-2 , Humanos , Receptores Depuradores Classe B/metabolismo
4.
Int J Pharm ; 550(1-2): 190-199, 2018 Oct 25.
Artigo em Inglês | MEDLINE | ID: mdl-30130606

RESUMO

Thermosensitive liposomes grafted with cholesterol-conjugated poly(N-(2-hydroxypropyl) methacrylamide mono/dilactate) (chol-pHPMAlac) have been developed for heat-induced release of doxorubicin (DOX). These liposomes release DOX completely during mild hyperthermia, but their interaction with blood cells and cancer cells has not been studied. Following intravenous administration, liposomes may interact with plasma proteins and various types of cells (e.g., endothelial cells, platelets, and macrophages), which would reduce their disposition in the tumor stroma. Interaction between liposomes and platelets may further cause platelet activation and thrombosis, which could lead to vascular occlusion and thromboembolic complications. The aim was to investigate DOX release kinetics in the presence of serum, stability, in vitro uptake by and toxicity to cancer cells and somatic cells, and platelet activating potential of the chol-pHPMAlac liposomes. DOX release was determined spectrofluorometrically. Liposome stability was determined in buffer and serum by dynamic light scattering and nanoparticle tracking analysis. Association with/uptake by and toxicity of empty liposomes to AML-12, HepG2 (both hepatocyte-derived cancer cells), RAW 264.7 (macrophages), and HUVEC (endothelial) cells was assayed in vitro. Platelet activation was determined by analysis of P-selectin expression and fibrinogen binding. DOPE:EPC liposomes (diameter = 135 nm) grafted with 5% chol-pHPMAlac (cloud point (CP) = 16 °C; Mn = 8.5 kDa) released less than 10% DOX at 37 °C in 30 min, whereas complete release took place at 47 °C or higher within 10 min. The size of these liposomes remained stable in buffer and serum during 24 h at 37 °C. Fluorescently labeled but DOX-lacking chol-pHPMAlac-liposomes exhibited poor association with/uptake by all cells under investigation, were not cytotoxic, and did not activate platelets in both buffered solution and whole blood. In conclusion, thermosensitive chol-pHPMAlac-grafted liposomes rapidly release DOX during mild hyperthermia. The liposomes are stable in a physiological milieu, are not taken up by cells that are encountered in an in vivo setting, and are non-antagonistic towards platelets. Chol-pHPMAlac-grafted liposomes are therefore good candidates for DOX delivery to tumors and temperature-triggered release in tumor stroma.


Assuntos
Acrilamidas , Antineoplásicos/administração & dosagem , Doxorrubicina/administração & dosagem , Portadores de Fármacos , Hipertermia Induzida , Lactatos , Lipossomos , Animais , Linhagem Celular Tumoral , Avaliação Pré-Clínica de Medicamentos , Humanos , Hipertermia Induzida/métodos , Lipossomos/química , Camundongos , Neoplasias/tratamento farmacológico
5.
Biomacromolecules ; 19(7): 2841-2848, 2018 07 09.
Artigo em Inglês | MEDLINE | ID: mdl-29750866

RESUMO

Ultrasmall gold atom clusters (<2 nm in diameter) or gold nanoclusters exhibit emergent photonic properties (near-infrared absorption and emission) compared to larger plasmonic gold particles because of the significant quantization of their conduction band. Although single gold nanocluster properties and applications are being increasingly investigated, little is still known about their behavior and properties when assembled into suprastructures, and even fewer studies are investigating their use for biomedical applications. Here, a simple synthetic pathway combines gold nanoclusters with thermosensitive diblock copolymers of poly(ethylene glycol) (PEG) and poly( N-isopropylacrylamide) (PNIPAm) to form a new class of gold-polymer, micelle-forming, hybrid nanoparticle. The nanohybrids' design is uniquely centered on enabling the temperature-dependent self-assembly of gold nanoclusters into the hydrophobic cores of micelles. This nonbulk assembly not only preserves but also enhances the attractive near-infrared photonics of the gold nanoclusters by significantly increasing their native fluorescent signal. In parallel to the fundamental insights into gold nanocluster ordering and assembly, the gold-polymer nanohybrids also demonstrated great potential as fluorescent live-imaging probes in vitro. This innovative material design based on the temperature-dependent, self-assembly of gold nanoclusters within a polymeric micelle's core shows great promise toward bioassays, nanosensors, and nanomedicine.


Assuntos
Substâncias Luminescentes/química , Nanopartículas Metálicas/química , Resinas Acrílicas/química , Ouro/química , Células Hep G2 , Humanos , Interações Hidrofóbicas e Hidrofílicas , Micelas , Polietilenoglicóis/química , Polimerização
6.
Eur J Pharm Sci ; 107: 112-125, 2017 Sep 30.
Artigo em Inglês | MEDLINE | ID: mdl-28679107

RESUMO

Selective elimination of macrophages by photodynamic therapy (PDT) is a new and promising therapeutic modality for the reduction of atherosclerotic plaques. m-Tetra(hydroxyphenyl)chlorin (mTHPC, or Temoporfin) may be suitable as photosensitizer for this application, as it is currently used in the clinic for cancer PDT. In the present study, mTHPC was encapsulated in polymeric micelles based on benzyl-poly(ε-caprolactone)-b-methoxy poly(ethylene glycol) (Ben-PCL-mPEG) using a film hydration method, with loading capacity of 17%. Because of higher lipase activity in RAW264.7 macrophages than in C166 endothelial cells, the former cells degraded the polymers faster, resulting in faster photosensitizer release and higher in vitro photocytotoxicity of mTHPC-loaded micelles in those macrophages. However, we observed release of mTHPC from the micelles in 30min in blood plasma in vitro which explains the observed similar in vivo pharmacokinetics of the mTHPC micellar formulation and free mTHPC. Therefore, we could not translate the beneficial macrophage selectivity from in vitro to in vivo. Nevertheless, we observed accumulation of mTHPC in atherosclerotic lesions of mice aorta's which is probably the result of binding to lipoproteins upon release from the micelles. Therefore, future experiments will be dedicated to increase the stability and thus allow accumulation of intact mTHPC-loaded Ben-PCL-mPEG micelles to macrophages of atherosclerotic lesions.


Assuntos
Doenças Cardiovasculares/tratamento farmacológico , Mesoporfirinas/administração & dosagem , Micelas , Fármacos Fotossensibilizantes/administração & dosagem , Animais , Doenças Cardiovasculares/metabolismo , Sobrevivência Celular/efeitos dos fármacos , Sobrevivência Celular/efeitos da radiação , Células Endoteliais/efeitos dos fármacos , Células Endoteliais/metabolismo , Células Endoteliais/efeitos da radiação , Feminino , Luz , Mesoporfirinas/sangue , Mesoporfirinas/farmacocinética , Mesoporfirinas/uso terapêutico , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Knockout , Camundongos Nus , Fotoquimioterapia , Fármacos Fotossensibilizantes/sangue , Fármacos Fotossensibilizantes/farmacocinética , Fármacos Fotossensibilizantes/uso terapêutico , Poliésteres/administração & dosagem , Poliésteres/farmacocinética , Poliésteres/uso terapêutico , Células RAW 264.7 , Oxigênio Singlete/química , Distribuição Tecidual
7.
Int J Pharm ; 525(2): 388-396, 2017 Jun 20.
Artigo em Inglês | MEDLINE | ID: mdl-28336456

RESUMO

The success of siRNA gene therapy requires the availability of safe and efficient delivery systems. In the present study, we investigated poly(vinyl benzyl trimethylammonium chloride) (PVTC) and its block copolymer with poly(oligo(ethyleneglycol) methacrylate) (POEGMA) as delivery vector for siRNA. Small polyplexes ranging from 8 to 25nm in diameter were formed in aqueous solution by spontaneous self-assembly of both the homopolymer and block copolymer with siRNA and the formed particles were stable at physiological ionic strength. It was shown that when human ovarian adenocarcinoma cells were transfected, siRNA polyplexes based on PVTC (40kDa) and PVTC-POEGMA-4 (PP4, 34kDa) efficiently induced luciferase gene silencing to the same extent as the formulation based on a commercial lipid (Lipofectamine®) (∼80%), and showed higher gene silencing than the linear polyethylenimine formulation linear polyethylenimine (∼35%). Importantly, the POEGMA block polymers displayed a significantly lower cytotoxicity as compared to L-pEI. siRNA polyplexes based on the block polymers displayed high cellular uptake resulting in ∼50% silencing of luciferase expression also in the presence of serum. These results demonstrate that PVTC-based polymers are promising siRNA delivery vectors.


Assuntos
Inativação Gênica , Terapia Genética/métodos , Poliestirenos/química , Compostos de Amônio Quaternário/química , RNA Interferente Pequeno/administração & dosagem , Adenocarcinoma/genética , Adenocarcinoma/terapia , Linhagem Celular Tumoral , Humanos , Polímeros , Transfecção
8.
Nanoscale ; 8(47): 19592-19604, 2016 Dec 01.
Artigo em Inglês | MEDLINE | ID: mdl-27748778

RESUMO

Cancer vaccines are at present mostly based on tumor associated protein antigens but fail to elicit strong cell-mediated immunity in their free form. For protein-based vaccines, the main challenges to overcome are the delivery of sufficient proteins into the cytosol of dendritic cells (DCs) and processing by, and presentation through, the MHC class I pathway. Recently, we developed a cationic dextran nanogel in which a model antigen (ovalbumin, OVA) is reversibly conjugated via disulfide bonds to the nanogel network to enable redox-sensitive intracellular release. In the present study, it is demonstrated that these nanogels, with the bound OVA, were efficiently internalized by DCs and were capable of maturating them. On the other hand, when the antigen was just physically entrapped in the nanogels, OVA was prematurely released before the particles were taken up by cells. When combined with an adjuvant (polyinosinic-polycytidylic acid, poly(I:C)), nanogels with conjugated OVA induced a strong protective and curative effect against melanoma in vivo. In a prophylactic vaccination setting, 90% of the mice vaccinated with nanogels with conjugated OVA + poly(I:C) did not develop a tumor. Moreover, in a therapeutic model, 40% of the mice showed clearance of established tumors and survived for the duration of the experiment (80 days) while the remaining mice showed substantial delay in tumor progression. In conclusion, our results demonstrate that conjugation of antigens to nanogels via reducible covalent bonds for intracellular delivery is a promising strategy to induce effective antigen-specific immune responses against cancer.


Assuntos
Apresentação de Antígeno , Vacinas Anticâncer/imunologia , Células Dendríticas/imunologia , Nanopartículas , Animais , Linhagem Celular Tumoral , Feminino , Melanoma Experimental/terapia , Camundongos , Camundongos Endogâmicos C57BL , Células NIH 3T3 , Ovalbumina/imunologia
9.
Mol Pharm ; 12(1): 150-61, 2015 Jan 05.
Artigo em Inglês | MEDLINE | ID: mdl-25384057

RESUMO

The applicability of small interfering RNA (siRNA) in future therapies depends on the availability of safe and efficient carrier systems. Ideally, siRNA delivery requires a system that is stable in the circulation but upon specific uptake into target cells can rapidly release its cargo into the cytoplasm. Previously, we evaluated a novel generation of carrier systems ("decationized" polyplexes) for DNA delivery, and it was shown that folate targeted decationized polyplexes had an excellent safety profile and showed intracellular triggered release upon cell specific uptake. Targeted decationized polyplexes consist of a core of disulfide cross-linked poly(hydroxypropyl methacrylamide) (pHPMA) stably entrapping nucleic acids and a shell of poly(ethylene glycol) (PEG) decorated with folate molecules. In the present study, the applicability of folate targeted decationized polyplexes for siRNA delivery was investigated. This required optimization of the carrier system particularly regarding the cross-linking density of the core of the polyplexes. Stable and nanosized siRNA decationized polyplexes were successfully prepared by optimizing the cross-link density of their core. Upon incubation in human plasma, a significant portion of siRNA remained entrapped in the decationized polyplexes as determined by fluorescence correlation spectroscopy (FCS). When tested in a folate receptor overexpressing cell line stably expressing luciferase, Skov3-luc, sequence specific gene silencing was observed. As expected, neither interference on the intrinsic luciferase expression nor on the cell metabolic activity (determined by XTT) was induced by the free-polymer or the siRNA polyplexes. In conclusion, targeted decationized polyplexes are safe and stable carriers that interact with the targeted cells and rapidly disassemble upon cell entry making them promising siRNA delivery systems.


Assuntos
Técnicas de Transferência de Genes , Metacrilatos/química , Nanopartículas/química , RNA Interferente Pequeno/administração & dosagem , Materiais Biocompatíveis/química , Linhagem Celular Tumoral , Ácido Fólico/química , Inativação Gênica , Humanos , Espectroscopia de Ressonância Magnética , Microscopia Eletrônica de Transmissão , Nanotecnologia , Ácidos Nucleicos/química , Polietilenoglicóis/química , Polímeros/química , Espectrofotometria Ultravioleta
10.
Pharm Res ; 31(10): 2593-604, 2014 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-24627415

RESUMO

PURPOSE: To investigate the effect of polyethylene glycol (PEG) in nanoparticles based on blends of hydroxylated aliphatic polyester, poly(D,L-lactic-co-glycolic-co-hydroxymethyl glycolic acid) (PLGHMGA) and PEG-PLGHMGA block copolymers on their degradation and release behavior. METHODS: Protein-loaded nanoparticles were prepared with blends of varying ratios of PEG-PLGHMGA (molecular weight of PEG 2,000 and 5,000 Da) and PLGHMGA, by a double emulsion method with or without using poly(vinyl alcohol) (PVA) as surfactant. Bovine serum albumin and lysozyme were used as model proteins. RESULTS: PEGylated particles prepared without PVA had a zeta potential ranging from ~ -3 to ~-35 mV and size ranging from ~200 to ~600 nm that were significantly dependent on the content and type of PEG-block copolymer. The encapsulation efficiency of the two proteins however was very low (<30%) and the particles rapidly released their content in a few days. In contrast, all formulations prepared with PVA showed almost similar particle properties (size: ~250 nm, zeta potential: ~-1 mV), while loading efficiency for both model proteins was rather high (80-90%). Unexpectedly, independent of the type of formulation, the nanoparticles had nearly the same release and degradation characteristics. NMR analysis showed almost a complete removal of PEG in 5 days which explains these marginal differences. CONCLUSIONS: Protein release and particle degradation are not substantially influenced by the content of PEG, likely because of the fast shedding of the PEG blocks. These PEG shedding particles are interesting system for intracellular delivery of drugs.


Assuntos
Portadores de Fármacos/química , Muramidase/administração & dosagem , Nanopartículas/química , Poliésteres/química , Polietilenoglicóis/química , Soroalbumina Bovina/administração & dosagem , Portadores de Fármacos/síntese química , Composição de Medicamentos , Liberação Controlada de Fármacos , Estabilidade de Medicamentos , Interações Hidrofóbicas e Hidrofílicas , Microscopia Eletrônica de Transmissão , Tamanho da Partícula , Poliésteres/síntese química , Polietilenoglicóis/síntese química , Propriedades de Superfície
SELEÇÃO DE REFERÊNCIAS
DETALHE DA PESQUISA