RESUMO
This work is devoted to studying the effects of non-magnetic shell coating on nanoparticles in a low frequency alternating magnetic field (LF AMF) on tumor cells in vitro. Two types of iron oxide nanoparticles with the same magnetic core with and without silica shells were synthesized. Nanoparticles with silica shells significantly decreased the viability of PC3 cancer cells in a low frequency alternating magnetic field according to the cytotoxicity test, unlike uncoated nanoparticles. We showed that cell death results from the intracellular membrane integrity failure, and the calcium ions concentration increase with the subsequent necrosis. Transmission electron microscopy images showed that the uncoated silica nanoparticles are primarily found in an aggregated form in cells. We believe that uncoated nanoparticles lose their colloidal stability in an acidic endosomal environment after internalization into the cell due to surface etching and the formation of aggregates. As a result, they encounter high endosomal macromolecular viscosity and become unable to rotate efficiently. We assume that effective rotation of nanoparticles causes cell death. In turn, silica shell coating increases nanoparticles stability, preventing aggregation in endosomes. Thus, we propose that the colloidal stability of magnetic nanoparticles inside cells is one of the key factors for effective magneto-mechanical actuation.
Assuntos
Nanopartículas de Magnetita , Neoplasias , Campos Magnéticos , Magnetismo , Nanopartículas de Magnetita/toxicidade , Dióxido de SilícioRESUMO
Intravital microscopy is widely used for in vivo studies of the mechanisms of carcinogenesis and response to antitumor therapy. For visualization of tumor cells in vivo, cell lines expressing fluorescent proteins are needed. Expression of exogenous proteins can affect cell growth rate and their tumorigenic potential. Therefore, comprehensive analysis of the morphofunctional properties of transduced cells is required for creating appropriate models of tumor microenvironment. In the present study, six lines of mouse tumor cells expressing green and red fluorescent proteins were derived. Analysis of cells morphology, growth kinetics, and response to chemotherapy in vitro revealed no significant differences between wild-type and transduced cell lines. Introduction of fluorescent proteins into the genome of 4T1 (murine breast cancer) and B16-F10 (murine melanoma) cells did not affect tumor growth rate after subcutaneous implantation to mice, while both CT26-GFP and CT26-RFP cells (murine colon cancer) were rejected starting from day 8 after implantation. Elucidation of the mechanisms underlying CT26-GFP/RFP rejection is required to modify transduction technique for creating the models of tumor microenvironment accessible for in vivo visualization. Transduced 4T1 and B16-F10 cell lines can be used for intravital microscopic imaging of tumor cells, neoplastic vasculature, and leukocyte subpopulations.
Assuntos
Microscopia Intravital/métodos , Proteínas Luminescentes/análise , Microambiente Tumoral/fisiologia , Animais , Linhagem Celular Tumoral , Neoplasias do Colo/diagnóstico por imagem , Neoplasias do Colo/metabolismo , Proteínas de Fluorescência Verde/análise , Humanos , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Microambiente Tumoral/genética , Proteína Vermelha FluorescenteRESUMO
Correction for 'New ferrocene-based 2-thio-imidazol-4-ones and their copper complexes. Synthesis and cytotoxicity' by D. A. Guk et al., Dalton Trans., 2018, DOI: 10.1039/c8dt03164a.
RESUMO
HYPOTHESIS: Hydrophobic bacteriochlorin based photosensitizer (PS) can be effectively immobilized on MNP covered by human serum albumin (HSA). PS loading into MNP protein shell allows solubilizing PS in water solution without altering its photodynamic activity. MNP@PS can serve as diagnostic tool for tracking PS delivery to tumor tissues by MRI. EXPERIMENTS: Immobilization on MNP-HSA-PEG was performed by adding PS solution in organic solvents with further purification. MNP@PS were characterized by DLS, HAADF STEM and AFM. Absorbance and fluorescence measurements were used to assess PS photophysical properties before and after immobilization. MNP@PS internalization into CT26 cells was investigated by confocal microscopy in vitro and MRI/IVIS were used for tracking MNP@PS delivery to tumors in vivo. FINDINGS: MNP@PS complexes were stable in water solution and retained PS photophysical activity. The length of side chain affected MNP@PS size, loading capacity and cell internalization. In vitro testing demonstrated MNP@PS delivery to cancer cells followed by photoinduced toxicity. In vivo studies confirmed that as-synthetized complexes can be used for MRI tracking over drug accumulation in tumors.
Assuntos
Antibióticos Antineoplásicos/administração & dosagem , Neoplasias do Colo/tratamento farmacológico , Doxorrubicina/administração & dosagem , Sistemas de Liberação de Medicamentos , Imageamento por Ressonância Magnética , Nanopartículas de Magnetita/química , Fármacos Fotossensibilizantes/administração & dosagem , Porfirinas/administração & dosagem , Animais , Antibióticos Antineoplásicos/farmacologia , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Doxorrubicina/farmacologia , Ensaios de Seleção de Medicamentos Antitumorais , Feminino , Humanos , Interações Hidrofóbicas e Hidrofílicas , Nanopartículas de Magnetita/administração & dosagem , Camundongos , Camundongos Endogâmicos BALB C , Tamanho da Partícula , Fármacos Fotossensibilizantes/farmacologia , Porfirinas/farmacologia , Albumina Sérica Humana/química , Propriedades de SuperfícieRESUMO
Synthesis, characterization (HRMS, NMR, EPR, XANES, UV-Vis spectroscopy, and electrochemistry), DNA and BSA binding and in vitro biological screening of two new ferrocene-incorporated thiohydantoin derivatives (5 and 6) and their copper coordination compounds are reported. The ferrocene-based thiohydantoin derivatives were prepared by copper-catalyzed azide alkyne cycloaddition reactions between alkynyl ferrocenes and 5-(Z)-3-(2-azidoethyl)-2-(methylthio)-5-(pyridin-2-ylmethylene)-1H-imidazol-4H-one. Alkynyl ferrocenes necessary for these syntheses were prepared by new procedures. Intermolecular redox reactions between the ferrocene fragment and copper(+2) coordinated ions were studied by different methods to determine the mechanism and kinetic constants of redox processes. Ferrocene-containing imidazolones (5 and 6) and their copper complexes were also tested for their in vitro cytotoxic activity against MCF-7 and A-549 carcinoma cells, and also against the noncancerous cell line Hek-293. The results showed modest cytotoxicity against the subjected cancer cell line compared with cisplatin. The ability of the obtained compounds to cause DNA degradation and cell apoptosis was investigated, and the distribution of cytosol/pellets was studied by AAS.
Assuntos
Antineoplásicos/farmacologia , Complexos de Coordenação/farmacologia , Cobre/farmacologia , Inibidores Enzimáticos/farmacologia , Imidazóis/farmacologia , Metalocenos/farmacologia , Telomerase/antagonistas & inibidores , Animais , Antineoplásicos/síntese química , Antineoplásicos/química , Apoptose/efeitos dos fármacos , Bovinos , Linhagem Celular , Proliferação de Células/efeitos dos fármacos , Complexos de Coordenação/síntese química , Complexos de Coordenação/química , Cobre/química , Clivagem do DNA , Relação Dose-Resposta a Droga , Ensaios de Seleção de Medicamentos Antitumorais , Inibidores Enzimáticos/síntese química , Inibidores Enzimáticos/química , Células HEK293 , Humanos , Imidazóis/síntese química , Imidazóis/química , Metalocenos/química , Estrutura Molecular , Soroalbumina Bovina/química , Relação Estrutura-Atividade , Telomerase/metabolismoRESUMO
The microbial resistance to antibiotics is a genuine global threat. Consequently, a search of new inhibitors remains of acute importance due to the increasing spread of multidrug resistance. Here we present a new type of non-ß-lactam ß-lactamase inhibitor PA-34 based on natural phenoxyaniline, identified using computer-assisted screening of scaffolds related to those of known low-affinity inhibitors. The compound displays reversible competitive inhibition of bacterial ß-lactamase TEM-171, with a Ki of 88 µM. Using enzyme kinetics, infra-red spectroscopy, fluorescence quenching and computer docking, we propose that the inhibitor binds at the entrance to the enzyme active site. This is a novel inhibition mechanism compared to binding covalently to the catalytic serine in the active site or non-covalently to the allosteric site. The residues involved in binding the inhibitor are conserved among molecular class A ß-lactamases. The identified compound and its proposed binding mode may have a potential for a regulation of the catalytic activity of a wide range of class A ß-lactamases. We also hypothesise that the presented route for finding non-ß-lactam compounds may be an effective and durable approach for combating bacterial antibiotic resistance.
Assuntos
Compostos de Anilina/farmacologia , Proteínas de Bactérias/antagonistas & inibidores , Inibidores de beta-Lactamases/farmacologia , beta-Lactamases/metabolismo , Acilação , Compostos de Anilina/química , Proteínas de Bactérias/química , Proteínas de Bactérias/metabolismo , Sítios de Ligação , Domínio Catalítico , Simulação por Computador , Descoberta de Drogas/métodos , Eletroforese em Gel de Poliacrilamida , Fluorescência , Cinética , Simulação de Acoplamento Molecular , Estrutura Molecular , Espectroscopia de Infravermelho com Transformada de Fourier/métodos , Inibidores de beta-Lactamases/química , beta-Lactamases/químicaRESUMO
A variety of structurally unrelated organic compounds has been reported to have antibacterial activity. Among these, certain small-molecule translation inhibitors have attracted a great deal of attention, due to their relatively high selectivity against prokaryotes, and an appropriate therapeutic index with minor "off target" effects. However, ribosomes are being considered as poorly druggable biological targets, thereby making some routine computational-based approaches to rational drug design and its development rather ineffective. Taking this into account, diversity-oriented biological screening can reasonably be considered as the most advantageous strategy. Thus, using a high-throughput screening (HTS) platform, we applied a unique biological assay for in vitro evaluation of thousands of organic molecules, especially targeted against bacterial ribosomes and translation. As a result, we have identified a series of structurally diverse small-molecule compounds that induce a reporter strain sensitive to translation and DNA biosynthesis inhibitors. In a cell free system, several molecules were found to strongly inhibit protein biosynthesis. Among them, compounds bearing a 2-guanidino-quinazoline core demonstrated the most promising antibacterial activity. With regard to the preliminary structure-activity relationship (SAR) study, we revealed that relatively small substituents at positions 4, 6 and 8 of the quinazoline ring significantly enhance the target activity whereas modification of the guanidine group leads to decrease or loss of antibacterial potency. This novel class of translation inhibitors can properly be regarded as a promising starting point for the development of novel antibacterial therapeutic or screening tools.
Assuntos
Antibacterianos/química , Biossíntese de Proteínas/efeitos dos fármacos , Inibidores da Síntese de Proteínas/química , Quinazolinas/química , Antibacterianos/farmacologia , Bactérias/efeitos dos fármacos , Bactérias/patogenicidade , Desenho de Fármacos , Humanos , Infecções/tratamento farmacológico , Infecções/microbiologia , Inibidores da Síntese de Proteínas/farmacologia , Quinazolinas/farmacologia , Bibliotecas de Moléculas Pequenas/química , Bibliotecas de Moléculas Pequenas/farmacologia , Relação Estrutura-AtividadeRESUMO
Superparamagnetic iron oxide magnetic nanoparticles (MNPs) are successfully used as contrast agents in magnetic-resonance imaging. They can be easily functionalized for drug delivery functions, demonstrating great potential for both imaging and therapeutic applications. Here we developed new pH-responsive theranostic core-shell-corona nanoparticles consisting of superparamagentic Fe3O4 core that displays high T2 relaxivity, bovine serum albumin (BSA) shell that binds anticancer drug, doxorubicin (Dox) and poly(ethylene glycol) (PEG) corona that increases stability and biocompatibility. The nanoparticles were produced by adsorption of the BSA shell onto the Fe3O4 core followed by crosslinking of the protein layer and subsequent grafting of the PEG corona using monoamino-terminated PEG via carbodiimide chemistry. The hydrodynamic diameter, zeta-potential, composition and T2 relaxivity of the resulting nanoparticles were characterized using transmission electron microscopy, dynamic light scattering, thermogravimetric analysis and T2-relaxometry. Nanoparticles were shown to absorb Dox molecules, possibly through a combination of electrostatic and hydrophobic interactions. The loading capacity (LC) of the nanoparticles was 8 wt.%. The Dox loaded nanoparticles release the drug at a higher rate at pH 5.5 compared to pH 7.4 and display similar cytotoxicity against C6 and HEK293 cells as the free Dox.
Assuntos
Antibióticos Antineoplásicos/administração & dosagem , Doxorrubicina/administração & dosagem , Óxido Ferroso-Férrico/química , Nanopartículas Metálicas , Neoplasias/diagnóstico , Neoplasias/terapia , Nanomedicina Teranóstica , Linhagem Celular Tumoral , Humanos , Microscopia Eletrônica de TransmissãoRESUMO
We report the synthesis and characterization of a new sulfur-containing derivative of bacteriochlorophyll a. The latter was isolated from biomass of the nonsulfur purple bacterium Rhodobacter capsulatus strain B10. The developed photosensitizer is N-aminobacteriopurpurinimide with an exocyclic amino group acylated with a lipoic acid moiety, which is a biogenic substance that acts as a cofactor of the pyruvate dehydrogenase and α-ketoglutarate dehydrogenase complexes in the body. The disulfide moiety of lipoic acid confers the compound aurophilicity, thus allowing its conjugation with gold nanoparticles (NP-Au) via S-Au bonds. The shape and the size of the resulting nanoconjugate with immobilized photosensitizer (PS-Au) were assessed by dynamic light scattering and transmission electron microscopy. The conjugated nanoparticles are spherical with hydrodynamic diameter of 100-110 nm. The PS-Au conjugate absorbs light at 824 nm and emits strong fluorescence at 830 nm, which allowed in vivo study of its dynamic biodistribution in rats bearing sarcoma M-1. Compared to the free photosensitizer, PS loaded on the gold nanoparticles (PS-Au) showed extended circulation time in the blood and enhanced tumor uptake due to nonspecific passive targeting when the drug accumulates in tumor sites through the leaky tumor neovasculature and does not return to the circulation.
Assuntos
Bacterioclorofila A/farmacologia , Ouro/farmacologia , Nanopartículas Metálicas/administração & dosagem , Neoplasias/tratamento farmacológico , Fotoquimioterapia/métodos , Fármacos Fotossensibilizantes/química , Fármacos Fotossensibilizantes/farmacologia , Animais , Bacterioclorofila A/química , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Ouro/química , Bicamadas Lipídicas/química , Nanopartículas Metálicas/química , Fármacos Fotossensibilizantes/síntese química , Ratos , Distribuição TecidualRESUMO
Toxic effects of cadmium chloride in concentration range from 1 to 300 µM on differentiated human intestinal epithelial Caco-2 cells after three hours of exposure were investigated. Processes of disorganization of the actin cytoskeleton associated with the toxic effects of cadmium were characterized by fluorescent microscopy. The cadmium-induced activation of cellular stress response processes (changes in the mRNA expression of caspase-3, heat-shock and oxidative stress genes) has been demonstrated. The study revealed dose-dependent changes in mRNA expression levels of proteins involved in the formation of adherens (E-Cadherin and p120 catenin) and tight intercellular junction contacts (Claudin 4 and ZO1). The time- and concentration-dependent trend of cell monolayer transepithelial resistance lowering, characterizing the loss of intercellular contacts density with prolongation of cell exposure cadmium chloride was estimated. Results indicates that proteins associated with tight and adhesion junctions are primary targets of cadmium. Amongst genes involved in cell junction formation, the genes encoding E-Cadherin and p120-catenin proved to be the most sensitive to cadmium influence.