RESUMEN
Human mesenchymal stem cells (hMSC) are multipotent cells that display the unique ability to home and engraft in tumor stroma. This remarkable tumor tropic property has generated a great deal of interest in many clinical settings. Recently, we showed that hMSC represent an excellent base for cell-mediated anticancer therapy since they are able to internalize paclitaxel (PTX) and to release it in an amount sufficient to inhibit tumor cell proliferation. In order to shed light on the dynamics of drug uptake and release, in the present paper we describe the synthesis of two novel thiophene-based fluorophore-paclitaxel conjugates, namely PTX-F32 and PTX-F35, as tools for in vitro drug tracking. We aimed to study the ability of these novel derivatives to be efficiently internalized by hMSC and, in a properly engineered coculture assay, to be released in the medium and taken up by tumor cells. In order to ensure better stability of the conjugates toward enzymatic hydrolysis, the selected oligothiophenes were connected to the taxol core at the C7 position through a carbamate linkage between PTX and the diamino linker. Antiproliferative experiments on both tumor cells and stromal cells clearly indicate that, in good correlation with the parent compound, cells are sensitive to nanomolar concentrations of the fluorescent conjugates. Moreover, in the coculture assay we were able to monitor, by fluorescence microscopy, PTX-F32 trafficking from hMSC toward glioblastoma U87 tumor cells. Our work paves the way for novel possibilities to perform extensive and high quality fluorescence-based analysis in order to better understand the cellular mechanisms involved in drug trafficking, such as microvescicle/exosome mediated release, in hMSC vehicle cells.
Asunto(s)
Sistemas de Liberación de Medicamentos , Colorantes Fluorescentes/análisis , Células Madre Mesenquimatosas/citología , Células Madre Mesenquimatosas/metabolismo , Paclitaxel/análisis , Paclitaxel/metabolismo , Tiofenos/química , Transporte Biológico , Línea Celular Tumoral , Exosomas/metabolismo , Colorantes Fluorescentes/química , Humanos , Conformación Molecular , Espectrometría de FluorescenciaRESUMEN
Mesenchymal stem cells (MSC) have the unique ability to home and engraft in tumor stroma. These features render them potentially a very useful tool as targeted delivery vehicles which can deliver therapeutic drugs to the tumor stroma. In the present study, we investigate whether fluorescent core-shell PMMA nanoparticles (FNPs) post-loaded with a photosensitizer, namely meso-tetrakis (4-sulfonatophenyl) porphyrin (TPPS) and uploaded by MSC could trigger osteosarcoma (OS) cell death in vitro upon specific photoactivation. In co-culture studies we demonstrate using laser confocal microscopy and time lapse imaging, that only after laser irradiation MSC loaded with photosensitizer-coated fluorescent NPs (TPPS@FNPs) undergo cell death and release reactive oxygen species (ROS) which are sufficient to trigger cell death of all OS cells in the culture. These results encourage further studies aimed at proving the efficacy of this novel tri-component system for PDT applications.
Asunto(s)
Neoplasias Óseas/tratamiento farmacológico , Células Madre Mesenquimatosas , Osteosarcoma/tratamiento farmacológico , Fotoquimioterapia , Fármacos Fotosensibilizantes/administración & dosificación , Porfirinas/administración & dosificación , Apoptosis/efectos de los fármacos , Neoplasias Óseas/metabolismo , Línea Celular Tumoral , Supervivencia Celular/efectos de los fármacos , Sistemas de Liberación de Medicamentos , Humanos , Nanopartículas/administración & dosificación , Osteosarcoma/metabolismo , Especies Reactivas de Oxígeno/metabolismoRESUMEN
A new aryltetralin lignan derivative, 1, was obtained by reacting dimethyl succinate and piperonal, furnishing the lactone 4-(3',4'-methylenedioxybenzyl)-4,5-dihydro-2(3H)-furanone, which was reacted once again with piperonal and LDA to give the dibenzylbutirolactone 7-hydroxyhinokinin. The cyclization of 7-hydroxyhinokinin into polygamain occurred in the presence of trifluoroacetic acid. The reduction of the furanic ring of polygamain was done by its reaction with DIBAL in THF, furnishing the diol functionalized lignin derivative 1 as single diastereomer. The enantiomeric fractions of 1 were obtained by preparative enantioselective HPLC. The absolute stereochemistry was assigned by electronic circular dichroism (ECD) and nuclear magnetic resonance (NMR) spectroscopy. An all-trans relative configuration was determined by NMR on the bases of ¹H coupling constants and nuclear Overhauser effect (n.O.e.) experiments. The absolute configuration at C1 was assigned on the basis of the ECD sign at 296 nm by comparison to the ECD spectra of structural analogues with defined stereochemistry. The assignment of the absolute configuration was confirmed by applying the exciton chirality method to the well-defined ECD couplets at 285 and 200 nm allied to the two electronic transitions L(b) and B(b) of the aromatic moieties, respectively. Rac-1 and its enantiomeric isomers were evaluated against important bacteria responsible for dental caries. The best results obtained for the (1R,2S,3S) isomer were against Streptococcus mutans (250 µM), Streptococcus salivarius (250 µM), Streptococcus sobrinus (280 µM) and Streptococcus mitis (280 µM). The (1S,2R,3R) isomer was active only against Streptococcus sanguinis (280 µM). The enantiomeric mixture was less active than the (1R,2S,3S) isomer.
Asunto(s)
Antibacterianos/química , Cariostáticos/química , Diseño de Fármacos , Lactonas/química , Lignanos/química , Tetrahidronaftalenos/química , Antibacterianos/análisis , Antibacterianos/farmacología , Cariostáticos/análisis , Cariostáticos/farmacología , Cromatografía Líquida de Alta Presión , Dicroismo Circular , Lactonas/análisis , Lactonas/farmacología , Lignanos/análisis , Lignanos/farmacología , Pruebas de Sensibilidad Microbiana , Conformación Molecular , Estructura Molecular , Antisépticos Bucales/análisis , Antisépticos Bucales/química , Antisépticos Bucales/farmacología , Resonancia Magnética Nuclear Biomolecular , Estereoisomerismo , Streptococcus/efectos de los fármacos , Streptococcus/crecimiento & desarrollo , Tetrahidronaftalenos/análisis , Tetrahidronaftalenos/farmacologíaRESUMEN
The iodine-magnesium exchange reaction allows the preparation of polyfunctional aryl, heteroaryl, or alkenyl magnesium reagents at low temperature. These reagents display the typical reactivity of Grignard compounds and undergo various copper-catalyzed reactions such as allylation or 1,4-addition. Using this halogen-metal exchange reaction, it was possible to generate polyfunctional magnesium reagents on the solid phase.