RESUMO
The complexity of drug delivery mechanisms calls for the development of new transport system designs. Here, we report a robust synthetic procedure toward stable glycodendrimer (glyco-DDM) series bearing glucose, galactose, and oligo(ethylene glycol)-modified galactose peripheral units. In vitro cytotoxicity assays showed exceptional biocompatibility of the glyco-DDMs. To demonstrate applicability in drug delivery, the anticancer agent doxorubicin (DOX) was encapsulated in the glyco-DDM structure. The anticancer activity of the resulting glyco-DDM/DOX complexes was evaluated on the noncancerous (BJ) and cancerous (MCF-7 and A2780) cell lines, revealing their promising generation- and concentration-dependent effect. The glyco-DDM/DOX complexes show gradual and pH-dependent DOX release profiles. Fluorescence spectra elucidated the encapsulation process. Confocal fluorescence microscopy demonstrated preferential cancer cell internalization of the glyco-DDM/DOX complexes. The conclusions were supported by computer modeling. Overall, our results are consistent with the assumption that novel glyco-DDMs and their drug complexes are very promising in drug delivery and related applications.
Assuntos
Antineoplásicos , Neoplasias Ovarianas , Antineoplásicos/farmacologia , Linhagem Celular Tumoral , Doxorrubicina/química , Doxorrubicina/farmacologia , Portadores de Fármacos/química , Sistemas de Liberação de Medicamentos/métodos , Liberação Controlada de Fármacos , Feminino , Humanos , Polietilenoglicóis/química , SilanosRESUMO
Although, in general, nanogels present a good biocompatibility and are able to mimic biological tissues, their unstability and uncontrollable release properties still limit their biomedical applications. In this study, a simple approach was used to develop dual-cross-linked dendrimer/alginate nanogels (AG/G5), using CaCl2 as cross-linker and amine-terminated generation 5 dendrimer (G5) as a cocrosslinker, through an emulsion method. Via their strong electrostatic interactions with anionic AG, together with cross-linker Ca(2+), G5 dendrimers can be used to mediate the formation of more compact structural nanogels with smaller size (433 ± 17 nm) than that (873 ± 116 nm) of the Ca(2+)-cross-linked AG nanogels in the absence of G5. Under physiological (pH 7.4) and acidic (pH 5.5) conditions, the sizes of Ca(2+)-cross-linked AG nanogels gradually decrease probably because of their degradation, while dual-cross-linked AG/G5 nanogels maintain a relatively more stable structure. Furthermore, the AG/G5 nanogels effectively encapsulate the anticancer drug doxorubicin (Dox) with a loading capacity 3 times higher than that of AG nanogels. The AG/G5 nanogels were able to release Dox in a sustained way, avoiding the burst release observed for AG nanogels. In vitro studies show that the AG/G5-Dox NGs were effectively taken up by CAL-72 cells (a human osteosarcoma cell line) and maintain the anticancer cytotoxicity levels of free Dox. Interestingly, G5 labeled with a fluorescent marker can be integrated into the nanogels and be used to track the nanogels inside cells by fluorescence microscopy. These findings demonstrate that AG/G5 nanogels may serve as a general platform for therapeutic delivery and/or cell imaging.
Assuntos
Antineoplásicos/farmacologia , Dendrímeros/química , Doxorrubicina/farmacologia , Sistemas de Liberação de Medicamentos , Fluorescência , Polietilenoglicóis/química , Polietilenoimina/química , Animais , Antineoplásicos/química , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Relação Dose-Resposta a Droga , Doxorrubicina/química , Ensaios de Seleção de Medicamentos Antitumorais , Humanos , Camundongos , Células NIH 3T3 , Nanogéis , Relação Estrutura-AtividadeRESUMO
Although doxorubicin (Dox) has been widely used in the treatment of different types of cancer, its insufficient cellular uptake and intracellular release is still a limitation. Herein, we report an easy process for the preparation of redox-sensitive nanogels that were shown to be highly efficient in the intracellular delivery of Dox. The nanogels (AG/Cys) were obtained through in situ cross-linking of alginate (AG) using cystamine (Cys) as a cross-linker via a miniemulsion method. Dox was loaded into the AG/Cys nanogels by simply mixing it in aqueous solution with the nanogels, that is, by the establishment of electrostatic interactions between the anionic AG and the cationic Dox. The results demonstrated that the AG/Cys nanogels are cytocompatible, have a high drug encapsulation efficiency (95.2 ± 4.7%), show an in vitro accelerated release of Dox in conditions that mimic the intracellular reductive conditions, and can quickly be taken up by CAL-72 cells (an osteosarcoma cell line), resulting in higher Dox intracellular accumulation and a remarkable cell death extension when compared with free Dox. The developed nanogels can be used as a tool to overcome the problem of Dox resistance in anticancer treatments and possibly be used for the delivery of other cationic drugs in applications beyond cancer.
Assuntos
Alginatos/química , Antibióticos Antineoplásicos/farmacologia , Doxorrubicina/farmacologia , Portadores de Fármacos/farmacologia , Nanoestruturas/química , Absorção , Antibióticos Antineoplásicos/química , Antibióticos Antineoplásicos/metabolismo , Linhagem Celular Tumoral , Forma Celular/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Doxorrubicina/química , Doxorrubicina/metabolismo , Portadores de Fármacos/química , Portadores de Fármacos/metabolismo , Humanos , Cinética , Oxirredução , Espectroscopia de Infravermelho com Transformada de FourierRESUMO
Cisplatin (cis-diamminedichloroplatinum(II)) is a potent chemotherapeutic agent commonly used to treat cancer. However, its use also leads to serious side effects, such as nephrotoxicity, ototoxicity, and cardiotoxicity, which limit the dose that can be safely administered to patients. To minimize these problems, dendrimers may be used as carriers for cisplatin through the coordination of their terminal functional groups to platinum. Here, cisplatin was conjugated to half-generation anionic PAMAM dendrimers in mono- and bidentate forms, and their biological effects were assessed in vitro. After preparation and characterization of the metallodendrimers, their cytotoxicity was evaluated against several cancer cell lines (A2780, A2780cisR, MCF-7, and CACO-2 cells) and a non-cancer cell line (BJ cells). The results showed that all the metallodendrimers were cytotoxic and that the cytotoxicity level depended on the cell line and the type of coordination mode (mono- or bidentate). Although, in this study, a correlation between dendrimer generation (number of carried metallic fragments) and cytotoxicity could not be completely established, the monodentate coordination form of cisplatin resulted in lower IC50 values, thus revealing a more accessible cisplatin release from the dendritic scaffold. Moreover, most of the metallodendrimers were more potent than the cisplatin, especially for the A2780 and A2780cisR cell lines, which showed higher selectivity than for non-cancer cells (BJ cells). The monodentate G0.5COO(Pt(NH3)2Cl)8 and G2.5COO(Pt(NH3)2Cl)32 metallodendrimers, as well as the bidentate G2.5COO(Pt(NH3)2)16 metallodendrimer, were even more active towards the cisplatin-resistant cell line (A2780cisR cells) than the correspondent cisplatin-sensitive one (A2780 cells). Finally, the effect of the metallodendrimers on the hemolysis of human erythrocytes was neglectable, and metallodendrimers' interaction with calf thymus DNA seemed to be stronger than that of free cisplatin.
Assuntos
DNA/química , Portadores de Fármacos/química , Nanoestruturas/química , Polímeros/química , RNA/química , Animais , Portadores de Fármacos/síntese química , Portadores de Fármacos/metabolismo , Portadores de Fármacos/farmacocinética , Géis , Humanos , Polímeros/síntese química , Polímeros/metabolismo , Polímeros/farmacocinéticaRESUMO
Dendrimers have been studied as promising materials for the delivery of anticancer drugs. In this work, low-generation (0-2) nitrile poly(alkylidenamine)-based dendrimers were explored as nanocarriers for the organometallic complex [Ru(η5-C5H5)(PPh3)2]+ (RuCp+) and investigated for their anticancer action and involved mechanisms, which were evaluated both in vitro and in vivo. It was observed that their biological behaviour is generation dependent, where the highest generation metallodendrimer (G2Ru) was overall more effective than the other metallodendrimers. G2Ru was active against a set of six cancer cell lines, revealing its important selectivity for these cells (the IC50 values were about 4-fold lower than that for non-cancer cells). Importantly, the in vivo studies with G2Ru in an MCF-7 xenograft mouse model showed that it exhibited low systemic toxicity, low accumulation in the main organs of the mice, preferential accumulation in the tumour, and remarkable capacity to limit tumour growth. The in vitro and in vivo studies revealed that G2Ru caused high levels of cell necrosis and apoptosis. The in vitro cell death mechanism studies showed the capacity of G2Ru to induce mitochondrial depolarization and ROS production. Altogether, pre-clinical results indicated G2Ru as a promising anticancer drug and the potential of low-generation poly(alkylidenamine)-based dendrimers as drug nanocarriers.
Assuntos
Antineoplásicos , Dendrímeros , Rutênio , Humanos , Camundongos , Animais , Rutênio/farmacologia , Dendrímeros/farmacologia , Antineoplásicos/farmacologia , Apoptose , Morte CelularRESUMO
We studied the possibility of detection of [Ru(η5-C5H5)(PPh3)2Cl] (abbreviated by RuCp) complex as a model system for Ru-based metallodrugs in human urine by using matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) without previous purification or removal of inorganic salts. Inorganic salts might prevent the detection of RuCp by MALDI-TOF MS, most likely through the increased number and intensity of background/organic matrix signals. This problem might be overcome by the acquisition of matrix-free spectra and the addition of nanoparticles, such as carbon dots, to the urine solution. Our results suggest that RuCp is easily detectable by MALDI-TOF MS in all acquisition conditions, with the CHCA matrix being the best for acquisition in phosphate-containing solutions, whereas in urine, DHB and matrix-free approach demonstrated the highest sensitivity, precision, and reproducibility. The sensitivity of matrix-free MALDI detection of RuCp could be increased by the addition of carbon dots to the urine. Based on theoretical calculations for all matrix/analyte combinations, the model for the interaction of RuCp with carbon dots was established, and higher sensitivity explained.
Assuntos
Carbono , Humanos , Reprodutibilidade dos Testes , Espectrometria de Massas por Ionização e Dessorção a Laser Assistida por MatrizRESUMO
Acquired immune deficiency syndrome (AIDS) due to human immunodeficiency virus type-1 (HIV-1) represents one of the most important sexually transmitted infections (STI) worldwide. Great international efforts have been made to stop new infections but, to date, several compounds failed as microbicides at different stages of clinical trials. The quest to design new molecules that could prevent these infections is essential. In this work, we synthesized the first, second and third generations of anionic dendrimers having carboxylate and sulfonate terminal groups, respectively named G1C, G2C, G3C and G1S, G2S, and G3S, starting from a family of poly(alkylideneamine) dendrimers with nitrile termini. The anionic terminal groups of these dendrimers were expected to prompt them to act against HIV-1 infection. All dendrimers were fully characterized by 1H- and 13C-NMR, FTIR, MS and zeta potential techniques. Importantly, they were able to remain stable in the solid state and aqueous solutions at least for one and a half years. Screening of these six new dendrimers was then performed to shed light on their potential anti-HIV-1 activity and their mechanism of action. Results showed that the dendrimers were cytocompatible and that G1C and G1S dendrimers had important activity against R5-HIV-1NLAD8 and X4-HIV-1NL4.3 isolates by acting directly on viral particles and blocking their entry in host cells. Additionally, G1C and G1S dendrimers maintained their inhibitory effect at different pH values. Through a vaginal irritation assay carried out in BALB/c mice, the safety of these new dendrimers for topical application was also shown. Taken together, our results clearly show that G1C and G1S dendrimers are strong candidates for developing an effective microbicide to prevent HIV-1 new infections.
Assuntos
Anti-Infecciosos/química , Dendrímeros/química , Animais , Ânions/química , Anti-Infecciosos/farmacologia , Anti-Infecciosos/uso terapêutico , Ácidos Carboxílicos/química , Linhagem Celular , Sobrevivência Celular/efeitos dos fármacos , Dendrímeros/farmacologia , Dendrímeros/uso terapêutico , Feminino , Infecções por HIV/tratamento farmacológico , Infecções por HIV/patologia , HIV-1/efeitos dos fármacos , Humanos , Camundongos , Camundongos Endogâmicos BALB C , Ácidos Sulfônicos/química , Vagina/efeitos dos fármacos , Vagina/patologia , Internalização do Vírus/efeitos dos fármacosRESUMO
Despite the wide research done in the field, the development of advanced drug delivery systems with improved drug delivery properties and effective anticancer capability still remains a great challenge. Based on previous work that showed the potentialities of the nanoclay Laponite as a pH-sensitive doxorubicin (Dox) delivery vehicle, herein we report a simple method to modulate its extent of drug release at different pH values. This was achieved by alternate deposition of cationic poly(allylamine) hydrochloride and anionic poly(sodium styrene sulfonate) (PAH/PSS) polyelectrolytes over the surface of Dox-loaded Laponite nanoparticles using the electrostatic layer-by-layer (LbL) self-assembly approach. The successful formation of polyelectrolyte multilayer-coated Dox/Laponite systems was confirmed by Dynamic Light Scattering and zeta potential measurements. Systematic studies were performed to evaluate their drug release profiles and anticancer efficiency. Our results showed that the presence of the polyelectrolyte multilayers improved the sustained release properties of Laponite and allowed a fine tuning of the extension of drug release at neutral and acidic pH values. The cytotoxicity presented by polyelectrolyte multilayer-coated Dox/Laponite systems towards MCF-7 cells was in accordance with the drug delivery profiles. Furthermore, cellular uptake studies revealed that polyelectrolyte multilayer-coated Dox/Laponite nanoparticles can be effectively internalized by cells conducting to Dox accumulation in cell nucleus.
Assuntos
Doxorrubicina/química , Nanopartículas/química , Polímeros/química , Silicatos/química , Núcleo Celular/metabolismo , Humanos , Concentração de Íons de Hidrogênio , Células MCF-7 , Nanopartículas/metabolismoRESUMO
The efficacy of the anticancer drug doxorubicin (Dox) is limited by an insufficient cellular uptake and drug resistance, which is partially due to ion trapping in acidic environments such as the extracellular environment of solid tumors and the interior of endolysosome vesicles. Herein, we describe the preparation and in vitro evaluation of a new type of nanohybrid for anticancer drug delivery which is capable of carrying a high load of the cationic Dox through the cell membrane. In addition, the nanohybrids use the acidic environment of the endolysosomes to release the drug, simultaneously helping to disrupt the endolysosomes and diminishing endolysosome Dox trapping. Furthermore, as the nanohybrid carriers are capable of sustained drug delivery, those that remain in the cytoplasm and still contain Dox are expected to exert a prolonged anticancer activity. Briefly, Dox is loaded onto biocompatible anionic Laponite(®) (LP) nanodisks with a high aspect ratio (25 nm in diameter and 0.92 nm in thickness) through strong electrostatic interactions to get Dox-loaded LP disks. Alginate (AG), a biocompatible natural polymer, is then coated onto the Dox-loaded LP disks (LP/Dox/AG nanohybrids) to prevent the burst release of the drug. The results demonstrate that the nanohybrids have a high encapsulation efficiency (80.8 ± 10.6%), are sensitive to pH and display a sustained drug release behavior. Cell culture experiments indicate that the LP/Dox/AG nanohybrids can be effectively internalized by CAL-72 cells (an osteosarcoma cell line), and exhibit a remarkable higher cytotoxicity to cancer cells than the free Dox. The merits of Laponite(®)/alginate nanohybrids, such as biocompatibility, high loading capacity and stimulus responsive release of cationic chemotherapeutic drugs, render them as excellent platforms for drug delivery.
Assuntos
Alginatos/farmacologia , Antineoplásicos/farmacologia , Doxorrubicina/farmacologia , Nanopartículas/química , Silicatos/farmacologia , Alginatos/química , Soluções Tampão , Morte Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Forma Celular/efeitos dos fármacos , Doxorrubicina/química , Endocitose/efeitos dos fármacos , Ácido Glucurônico/química , Ácido Glucurônico/farmacologia , Ácidos Hexurônicos/química , Ácidos Hexurônicos/farmacologia , Humanos , Hidrodinâmica , Concentração de Íons de Hidrogênio/efeitos dos fármacos , Nanopartículas/ultraestrutura , Silicatos/química , TitulometriaRESUMO
Degradable hybrid hydrogels with improved stability are prepared by incorporating nanodisks of biocompatible laponite (LP) in alginate (AG) hydrogels using Ca(2+) as a crosslinker. The Dox-loaded hybrid hydrogels give a controlled Dox release at physiological environment in a sustained manner. Under conditions that mimic the tumor environment, both the sustainability in the Dox release (up to 17 d) and the release efficiency from LP/AG-Dox hydrogels are improved. The in situ degradation of these hybrid hydrogels gives rise to nanohybrids that might serve as vehicles for carrying Dox through the cell membrane and diminish the effect of Dox ion-trapping in the acidic extracellular environment of the tumor and/or in the endo-lysosomal cell compartments.
Assuntos
Antibióticos Antineoplásicos/farmacologia , Doxorrubicina/farmacologia , Sistemas de Liberação de Medicamentos/métodos , Hidrogéis/administração & dosagem , Alginatos/química , Antibióticos Antineoplásicos/administração & dosagem , Antibióticos Antineoplásicos/farmacocinética , Neoplasias Ósseas/tratamento farmacológico , Neoplasias Ósseas/patologia , Linhagem Celular Tumoral , Doxorrubicina/administração & dosagem , Doxorrubicina/farmacocinética , Estabilidade de Medicamentos , Ácido Glucurônico/química , Ácidos Hexurônicos/química , Humanos , Hidrogéis/química , Osteossarcoma/tratamento farmacológico , Osteossarcoma/patologia , Silicatos/químicaRESUMO
The development of pH-sensitive drug delivery nanosystems that present a low drug release at the physiological pH and are able to increase the extent of the release at a lower pH value (like those existent in the interstitial space of solid tumors (pH 6.5) and in the intracellular endolysosomal compartments (pH 5.0)) is very important for an efficient and safe cancer therapy. Laponite (LP) is a synthetic silicate nanoparticle with a nanodisk structure (25 nm in diameter and 0.92 nm in thickness) and negative-charged surface, which can be used for the encapsulation of doxorubicin (DOX, a cationic drug) through electrostatic interactions and exhibit good pH sensitivity in drug delivery. However, the colloidal instability of LP still limits its potential clinical applications. In this study, we demonstrate an elegant strategy to develop stable Laponite-based nanohybrids through the functionalization of its surface with an amphiphile PEG-PLA copolymer by a self-assembly process. The hydrophobic block of PEG-PLA acts as an anchor that binds to the surface of drug-loaded LP nanodisks, maintaining the core structure, whereas the hydrophilic PEG part serves as a protective stealth shell that improves the whole stability of the nanohybrids under physiological conditions. The resulting nanocarriers can effectively load the DOX drug (the encapsulation efficiency is 85%), and display a pH-enhanced drug release behavior in a sustained way. In vitro biological evaluation indicated that the DOX-loaded nanocarriers can be effectively internalized by CAL-72 cells (an osteosarcoma cell line), and exhibit a remarkable higher anticancer cytotoxicity than free DOX. The merits of Laponite/PEG-PLA nanohybrids, such as good cytocompatibility, excellent physiological stability, sustained pH-responsive release properties, and improved anticancer activity, make them a promising platform for the delivery of other therapeutic agents beyond DOX.