Nanocápsulas de poliaminoácidos para la liberación selectiva de fármacos antitumorales

El objetivo principal de esta tesis ha sido el diseño y desarrollo de nanocápsulas de poliaminoácidos para la liberación de fármacos antitumorales. Estos sistemas compuestos por un núcleo oleoso y una cubierta de ácido poliglutámico (PGA), y su modificación con polietilenglicol (PEG), emergen como una nueva plataforma de vehiculización selectiva para moléculas hidrofóbicas antitumorales como la plitidepsina y el docetaxel. Se ha prestado especial atención al efecto del PEG, evaluando diferentes grados de PEGilación (24 y 57% p/p de PEG), con la intención de conseguir sistemas con mayor permanencia en el torrente circulatorio y modificar así la cinética de eliminación en plasma de las moléculas asociadas. Las nanocápsulas de PGA-PEG con un elevado grado de PEGilación incrementaron al doble la vida media y el MRT de la plitidepsina, mientras el AUC resultó ser 5 veces mayor en comparación con la solución de plitidepsina en Cremophor® EL, formulación utilizada como referencia. Los experimentos in vivo mostraron que los sistemas altamente pegilados presentan una menor toxicidad, son mejor tolerados respecto a la formulación de referencia, y presentan una fuerte inhibición del crecimiento tumoral en el modelo xenograft de tumor renal (MRI-H-121). Adicionalmente, las nanocápsulas de PGA-PEG con elevado grado de PEGilación cargadas docetaxel mostraron una elevada actividad antitumoral en el modelo de glioma U87, reduciendo en un 60% el crecimiento del tumor y mostrando un aumento significativo de la supervivencia con respecto a los animales tradatos con el Taxotere®. En una segunda etapa, se evaluó el potencial de otro tipo de nanocápsulas, las nanocápsulas de poliarginina (PARG), como nuevos vehículos para la administración oral de fármacos antitumorales. Los estudios llevados a cabo en cultivos celulares Caco-2 demostraron que las nanocápsulas pueden a promover el transporte paracelular de fármacos modificando las uniones entre células epiteliales, internalizándose en la monocapa celular de manera eficiente. Tras su administración oral, mediante estudios de imagen in vivo, se confirmó la permanencia de las nanocápsulas en el tracto gastrointestinal. Todos estos resultados ponen de manifiesto el potencial de las nanocápsulas de poliaminoácidos como sistemas de vehiculización para mejorar la absorción parenteral y oral de fármacos antitumorales

Alcian blue staining to study nanoparticle-cell interactions at transmission electron microscopy

Alcian blue staining has been used to visualise nanoparticles at transmission electron microscop

Alcian blue staining to track the intracellular fate of hyaluronic-acid-based nanoparticles at transmission electron microscopy

The main step in the assessment of nanomaterial safety and suitability for biomedical use is the location and the dynamic tracking of nanoparticles (NPs) inside cells or tissues. To precisely investigate the uptake mechanisms and intracellular fate of NPs, transmission electron microscopy is the technique of choice; however, the detection of NPs may sometimes be problematic. In fact, while NPs containing strongly electron dense (e.g. metal) components do not require specific detection methods at the ultrastructural level, organic NPs are hardly detectable in the intracellular enviromnent due to their intrinsic moderate electron density. In this study, the critical-electrolyte-concentration Alcian Blue method set up by Schofield et at in 1975 was applied to track hyaluronic-acid-based NPs in muscle cells in vitro. This long-established histochemical method proved to be a powerful tool allowing to identify not only whole NPs while entering cells and moving into the cytoplasm, but also their remnants following lysosomal degradation and extrusion

Hyaluronic acid-based nanocomplexes as an innovative therapeutic tool to treat myotonic dystrophy

Hyaluronic acid-based nanocomplexes have been synthesized and tested as an innovative therapeutic tool to treat myotonic dystroph

Sulphated locust bean gum-coated lipid nanocapsules as potential lung delivery carriers

Drugs pertaining to Biopharmaceutics Classification System (BCS) classes II and IV have limitations in their delivery, including in the lung. Therefore, drug delivery carriers have been proposed to improve the therapeutic effectiveness of such drugs. This work proposes lipid nanocapsules (LNC) as a potential platform for lung drug delivery. Locust bean gum (LBG), which is a galactomannan, was used as polymeric shell, protecting the oily core of the nanocapsules and providing their surface with hydrophilic character. Due to the neutral character of LBG, in order to enable nanocapsule formation, a sulphate derivative (LBGS) was prepared, which was confirmed by Fourier-transformed infrared (FTIR) spectroscopy. The electrostatic interaction between the negatively charged sulphate groups of LBGS and the positively charged groups of the used cationic lipid (1,2-dioleoyloxy-3- trimethylammoniumpropanchloride, DOTAP), allowed the formation of monodisperse nanocapsules, with sizes around 200 nm and strongly negative zeta potentials, between -70 and -85 mV. Envisaging potential lung drug delivery, the LBGS-coated LNC were co-formulated with mannitol using spray-drying, producing microencapsulated nanocapsules. Feret’s diameter was determined to be 2.6 ± 1.8 µm and 3.1 ± 1.9 µm for Man (control) and Man/LNC microparticles, respectively. Further studies are underway in order to optimise both the nanoplatform and the dry powder formulation

Fucoidan from Fucus Vesiculosus: Evaluation of the impact of the Sulphate content on Nanoparticle production and Cell toxicity

The composition of seaweeds is complex, with vitamins, phenolic compounds, minerals, and polysaccharides being some of the factions comprising their structure. The main polysaccharide in brown seaweeds is fucoidan, and several biological activities have been associated with its structure. Chitosan is another marine biopolymer that is very popular in the biomedical field, owing to its suitable features for formulating drug delivery systems and, particularly, particulate systems. In this work, the ability of fucoidan to produce nanoparticles was evaluated, testing different amounts of a polymer and using chitosan as a counterion. Nanoparticles of 200–300 nm were obtained when fucoidan prevailed in the formulation, which also resulted in negatively charged nanoparticles. Adjusting the pH of the reaction media to 4 did not affect the physicochemical characteristics of the nanoparticles. The IC50 of fucoidan was determined, in both HCT−116 and A549 cells, to be around 160 µg/mL, whereas it raised to 675–100 µg/mL when nanoparticles (fucoidan/chitosan = 2/1, w/w) were tested. These marine materials (fucoidan and chitosan) provided features suitable to formulate polymeric nanoparticles to use in biomedical applications.info:eu-repo/semantics/publishedVersio

A new potential nano-oncological therapy based on polyamino acid nanocapsules

A critical objective in cancer therapy is to reduce the systemic toxicity through the modification of the biodistribution of anticancer drugs. Herein, we disclose a new biodegradable nanocarrier, polyglutamic acid (PGA) nanocapsules, and present the in vivo pharmacokinetics/toxicity proof-of-concept for the anticancer drug plitidepsin. These novel nanocapsules were prepared using a modified solvent displacement technique where the polyamino acid was electrostatically deposited onto the lipid core. The nanocapsules exhibited an average size of 200 nm, a negative zeta potential and a great capacity for the encapsulation of plitidepsin (encapsulation efficiency above 90%). In addition, the nanocapsules could be freeze-dried and showed an adequate stability profile upon storage. Finally, the in vivo proof-of-concept studies performed in mice indicated that the encapsulation provided the drug with a prolonged blood circulation and a significantly reduced toxicity. In fact, the maximum tolerated dose of the nanoencapsulated drug was more than 3 times that of the reference formulation (Cremophor® EL plitidepsin solution). Overall, beyond the value of this specific formulation, the work reported here represents the evidence of the potential of polyamino acid nanocapsules in nano-oncological therapyThe authors would like to acknowledge financial support from CENIT-NANOFAR XS53 project, PharmaMar, Spain, the Ministry of Sciences and Innovation ((CTQ2009-10963), the Xunta de Galicia (Competitive Reference Groups-FEDER funds Ref. 2010/18, and CN2011/037) and the European Commission FP7 EraNet — EuroNanoMed Program-Instituto Carlos III (Lymphotarg proyect, Ref. PS09/02670). Giovanna Lollo has a fellowship from the Ministry of Education of Spain. Marcos Garcia Fuentes acknowledges an Isidro Parga Pondal Fellowship from Xunta de GaliciaS

Synergy between intraperitoneal aerosolization (PIPAC) and cancer nanomedicine : cisplatin-loaded polyarginine-hyaluronic acid nanocarriers efficiently eradicate peritoneal metastasis of advanced human ovarian cancer

Intra-abdominal dissemination of peritoneal nodules, a condition known as peritoneal carcinomatosis (PC), is typically diagnosed in ovarian cancer patients at the advanced stages. The current treatment of PC consists of perioperative systemic chemotherapy and cytoreductive surgery, followed by intra-abdominal flushing with solutions of chemotherapeutics such as cisplatin and oxaliplatin. In this study, we developed cisplatin-loaded polyarginine-hyaluronic acid nanoscale particles (Cis-pARG-HA NPs) with high colloidal stability, marked drug loading efficiency, unimpaired biological activity, and tumor-targeting ability. Injected Cis-pARG-HA NPs showed enhanced antitumor activity in a rat model of PC, compared to injection of the free cisplatin drug. The activity of Cis-pARG-HA NPs could even be further improved when administered by an intra-abdominal aerosol therapy, referred to as pressurized intraperitoneal aerosol chemotherapy (PIPAC). PIPAC is hypothesized to ensure a more homogeneous drug distribution together with a deeper drug penetration into peritoneal tumor nodules within the abdominal cavity. Using fluorescent pARG-HA NPs, this enhanced nanoparticle deposit on tumors could indeed be observed in regions opposite the aerosolization nozzle. Therefore, this study demonstrates that nanoparticles carrying chemotherapeutics can be synergistically combined with the PIPAC technique for IP therapy of disseminated advanced ovarian tumors, while this synergistic effect was not observed for the administration of free cisplatin

Drug delivery system for platinum-based drugs

The present invention concerns the nanomedicine field for anticancer therapy. The treatment of cancer using plat¬ inum-based compounds comprises certain drawbacks such as biocompatibility, loading efficacy, leakage of drugs during storage and in the bloodstream, more particularly due to the nature of the nanocarriers for platinum delivery. The inventors found that a novel nanosystem allows improving platinum-based drug in vivo performance, kinetics and efficacy. In particular, the present invention re¬ lates to nanoparticles useful as drug delivery system, said nanoparticles being formed from at least: (a) platinum-based drug, (b) poly-L-arginine, and (c) hyaluronic acid. Particularly, the inventors tested these nanoparticles in terms of entrapment efficiency and also carried out in vitro experiments in 2D cell culture (viability studies o B6KPC3, A549 and HT-29 cells) and 3D cell model (spheroids made of HTC-1 16) and in vivo experiments (by injecting intravenously to mice said nanoparticles or comparative oxaliplatin solution) to prove their efficiency

Alcian blue staining to track the intracellular fate of hyaluronic-acid-based nanoparticles at transmission electron microscopy.

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