Effect of the Protein Corona Formation on Antibody Functionalized Liquid Lipid Nanocarriers.

The main aim of this study is to report basic knowledge on how a protein corona (PC) could affect or modify the way in which multifunctionalized nanoparticles interact with cells. With this purpose, we have firstly optimized the development of a target-specific nanocarrier by coupling a specific fluorescent antibody on the surface of functionalized lipid liquid nanocapsules (LLNCs). Thus, an anti-HER2-FITC antibody (αHER2) has been used, HER2 being a surface receptor that is overexpressed in several tumor cells. Subsequently, the in vitro formation of a PC has been developed using fetal bovine serum supplemented with human fibrinogen. Dynamic Light Scattering (DLS), Nanoparticle Tracking Analysis (NTA), Laser Doppler Electrophoresis (LDE), and Gel Chromatography techniques have been used to assure a complete physico-chemical characterization of the nano-complexes with (LLNCs-αHER2-PC) and without (LLNCs-αHER2) the surrounding PC. In addition, cellular assays were performed to study the cellular uptake and the specific cellular-nanocarrier interactions using the SKBR3 (high expression of HER2) breast cancer cell line and human dermal fibroblasts (HDFa) (healthy cell line without expression of HER2 receptors as control), showing that the SKBR3 cell line had a higher transport rate (50-fold) than HDFa at 60 min with LLNCs-αHER2. Moreover, the SKBR3 cell line incubated with LLNCs-αHER2-PC suffered a significant reduction (40%) in the uptake. These results suggest that the formation of a PC onto LLNCs does not prevent specific cell targeting, although it does have an important influence on cell uptake.

Progress and Hurdles of Therapeutic Nanosystems against Cancer.

Nanomedicine against cancer, including diagnosis, prevention and treatment, has increased expectations for the solution of many biomedical challenges in the fight against this disease. In recent decades, an exhaustive design of nanosystems with high specificity, sensitivity and selectivity has been achieved due to a rigorous control over their physicochemical properties and an understanding of the nano-bio interface. However, despite the considerable progress that has been reached in this field, there are still different hurdles that limit the clinical application of these nanosystems, which, along with their possible solutions, have been reviewed in this work. Specifically, physiological processes as biological barriers and protein corona formation related to the administration routes, designing strategies to overcome these obstacles, promising new multifunctional nanotherapeutics, and recent clinical trials are presented in this review.

Anti-CD44-Conjugated Olive Oil Liquid Nanocapsules for Targeting Pancreatic Cancer Stem Cells.

The latest trends in cancer research and nanomedicine focus on using nanocarriers to target cancer stem cells (CSCs). Specifically, lipid liquid nanocapsules are usually developed as nanocarriers for lipophilic drug delivery. Here, we developed olive oil liquid NCs (O2LNCs) functionalized by covalent coupling of an anti-CD44-fluorescein isothiocyanate antibody (αCD44). First, O2LNCs are formed by a core of olive oil surrounded by a shell containing phospholipids, a nonionic surfactant, and deoxycholic acid molecules. Then, O2LNCs were coated with an αCD44 antibody (αCD44-O2LNC). The optimization of an αCD44 coating procedure, a complete physicochemical characterization, as well as clear evidence of their efficacy in vitro and in vivo were demonstrated. Our results indicate the high targeted uptake of these αCD44-O2LNCs, and the increased antitumor efficacy (up to four times) of paclitaxel-loaded-αCD44-O2LNC compared to free paclitaxel in pancreatic CSCs (PCSCs). Also, αCD44-O2LNCs were able to selectively target PCSCs in an orthotopic xenotransplant in vivo model.

Smart Drug-Delivery Systems for Cancer Nanotherapy.

BACKGROUND: Despite all the advances achieved in the field of tumor-biology research, in most cases conventional therapies including chemotherapy are still the leading choices. The main disadvantage of these treatments, in addition to the low solubility of many antitumor drugs, is their lack of specificity, which leads to the occurrence of severe side effects due to nonspecific drug uptake by healthy cells. OBJECTIVE: The purpose of this manuscript is to review and analyze the recent progress made in cancer nanotherapy. RESULTS: Progress in nanotechnology and its application in medicine have provided new opportunities and different smart systems. Such systems can improve the intracellular delivery of the drugs due to their multifunctionality and targeting potential. First, we provide a global overview of cancer and different smart nanoparticles currently used in oncology. Then, we analyze in detail the development of drug-delivery strategies in cancer therapy, focusing mainly on the intravenously administered smart nanoparticles. Finally, we discuss the challenges, clinical trials, marketed nanomedicines and future directions of the nanotherapy applied to cancer treatment. CONCLUSION: In this review, we have evidenced the tremendous potential that smart drug-delivery systems have to enhance the therapeutic effect of current standard treatment modalities, including chemotherapies and radiotherapies.

Dual delivery nanosystem for biomolecules. Formulation, characterization, and in vitro release.

Because of the biocompatible and biodegradable properties of poly (lactic-co-glycolic acid) (PLGA), nanoparticles (NPs) based on this polymer have been widely studied for drug/biomolecule delivery and long-term sustained-release. In this work, two different formulation methods for lysozyme-loaded PLGA NPs have been developed and optimized based on the double-emulsion (water/oil/water, W/O/W) solvent evaporation technique. They differ mainly in the phase in which the surfactant (Pluronic® F68) is added: water (W-F68) and oil (O-F68). The colloidal properties of these systems (morphology by SEM and STEM, hydrodynamic size by DLS and NTA, electrophoretic mobility, temporal stability in different media, protein encapsulation, release, and bioactivity) have been analyzed. The interaction surfactant-protein depending on the formulation procedure has been characterized by surface tension and dilatational rheology. Finally, cellular uptake by human mesenchymal stromal cells and cytotoxicity for both systems have been analyzed. Spherical hard NPs are made by the two methods However, in one case, they are monodisperse with diameters of around 120nm (O-F68), and in the other case, a polydisperse system of NPs with diameters between 100 and 500nm is found (W-F68). Protein encapsulation efficiency, release and bioactivity are maintained better by the W-F68 formulation method. This multimodal system is found to be a promising "dual delivery" system for encapsulating hydrophilic proteins with strong biological activity at the cell-surface and cytoplasmic levels.

Synthesis and characterization of lipid immuno-nanocapsules for directed drug delivery: selective antitumor activity against HER2 positive breast-cancer cells.

Lipid nanocapsules (LNC) are usually developed as nanocarriers for lipophilic drug delivery. The surface characteristics of these colloidal particles are determinant for a controlled and directed delivery to target tissues with specific markers. We report the development of immuno-nanocapsules, in which some antibody molecules with different immuno-specificity are conjugated to the nanocapsule surface, offering the standardization of a simple method to obtain vectorized nanosystems with specific recognition properties. Nanocapsules were prepared by a solvent-displacement technique, producing an oily core coated by a functional shell of different biocompatible molecules and surface carboxylic groups. Three different antibodies (one a specific HER2 oncoprotein antibody) were conjugated with these nanoparticles by the carbodiimide method, which allows the covalent immobilization of protein molecules through carboxylic surface groups. The immuno-nanocapsules were completely characterized physico-chemically via electrokinetic and colloidal stability experiments, confirming the correct immobilization of these antibody molecules on the colloidal nanoparticles. Also, additional immunological analyses verified that these IgG-LNC complexes showed the expected specific immuno-response. Finally, different healthy and tumoral breast-cell lines were cultured in vitro with Nile-Red-loaded and docetaxel-loaded HER2 immuno-nanocapsules. The results indicate that our immuno-nanocapsules can increase their uptake in HER2 overexpressing tumoral cell lines.

Novel drug delivery system based on docetaxel-loaded nanocapsules as a therapeutic strategy against breast cancer cells.

In the field of cancer therapy, lipid nanocapsules based on a core-shell structure are promising vehicles for the delivery of hydrophobic drugs such as docetaxel. The main aim of this work was to evaluate whether docetaxel-loaded lipid nanocapsules improved the anti-tumor effect of free docetaxel in breast cancer cells. Three docetaxel-loaded lipid nanocapsules were synthesized by solvent displacement method. Cytotoxic assays were evaluated in breast carcinoma (MCF-7) cells treated by the sulforhodamine B colorimetric method. Cell cycle was studied by flow cytometry and Annexin V-FITC, and apoptosis was evaluated by using propidium iodide assays. The anti-proliferative effect of docetaxel appeared much earlier when the drug was encapsulated in lipid nanoparticles than when it was free. Docetaxel-loaded lipid nanocapsules significantly enhanced the decrease in IC(50) rate, and the treated cells evidenced apoptosis and a premature progression of the cell cycle from G(1) to G(2)-M phase. The chemotherapeutic effect of free docetaxel on breast cancer cells is improved by its encapsulation in lipid nanocapsules. This approach has the potential to overcome some major limitations of conventional chemotherapy and may be a promising strategy for future applications in breast cancer therapy.

Characterization of different functionalized lipidic nanocapsules as potential drug carriers.

Lipid nanocapsules (LNC) based on a core-shell structure consisting of an oil-filled core with a surrounding polymer layer are known to be promising vehicles for the delivery of hydrophobic drugs in the new therapeutic strategies in anti-cancer treatments. The present work has been designed as basic research about different LNC systems. We have synthesized-and physico-chemically characterized-three different LNC systems in which the core was constituted by olive oil and the shell by different phospholipids (phosphatidyl-serine or lecithin) and other biocompatible molecules such as Pluronic(®) F68 or chitosan. It is notable that the olive-oil-phosphatidyl-serine LCN is a novel formulation presented in this work and was designed to generate an enriched carboxylic surface. This carboxylic layer is meant to link specific antibodies, which could facilitate the specific nanocapsule uptake by cancer cells. This is why nanoparticles with phosphatidyl-serine in their shell have also been used in this work to form immuno-nanocapsules containing a polyclonal IgG against a model antigen (C-reactive protein) covalently bounded by means of a simple and reproducible carbodiimide method. An immunological study was made to verify that these IgG-LNC complexes showed the expected specific immune response. Finally, a preliminary in vitro study was performed by culturing a breast-carcinoma cell line (MCF-7) with Nile-Red-loaded LNC. We found that these cancer cells take up the fluorescent Nile- Red molecule in a process dependent on the surface properties of the nanocarriers.