Receptor-targeted nanocarriers for therapeutic delivery to cancer

Abstract

Efficient and site-specific delivery of therapeutic drugs is a critical challenge in clinical treatment of cancer. Nano-sized carriers such as liposomes, micelles, and polymeric nanoparticles have been investigated for improving bioavailability and pharmacokinetic properties of therapeutics via various mechanisms, for example, the enhanced permeability and retention (EPR) effect. Further improvement can potentially be achieved by conjugation of targeting ligands onto nanocarriers to achieve selective delivery to the tumour cell or the tumour vasculature. Indeed, receptor-targeted nanocarrier delivery has been shown to improve therapeutic responses both in vitro and in vivo. A variety of ligands have been investigated including folate, transferrin, antibodies, peptides and aptamers. Multiple functionalities can be incorporated into the design of nanoparticles, e.g., to enable imaging and triggered intracellular drug release. In this review, we mainly focus on recent advances on the development of targeted nanocarriers and will introduce novel concepts such as multi-targeting and multi-functional nanoparticles.     

Monoolein-based nanocarriers for enhanced folate receptor-mediated RNA delivery to cancer cells

We report the development and characterization of a novel nanometric system for specific delivery of therapeutic siRNA for cancer treatment. This vector is based on a binary mixture of the cationic surfactant dioctadecyldimethylammonium chloride (DODAC) and the helper lipid monoolein (MO). These liposomes were previously validated by our research group as promising non-viral vectors for nucleic acid delivery. In this work, the DODAC:MO vesicles were for the first time functionalized with polyethylene glycol and PEG-folate conjugates to achieve both maximal stability in biological fluids and increase selectivity toward folate receptor α expressing cells. The produced DODAC:MO:PEG liposomes were highly effective in RNA complexation (close to 100%), and the resulting lipoplexes also demonstrated high stability in conditions simulating their administration by intravenous injection (physiological pH, high NaCl, heparin and fetal bovine serum concentrations). In addition, cell uptake of the PEG-folate-coated lipoplexes was significantly greater in folate receptor α positive breast cancer cells (39% for 25?µg/mL of lipid and 31% for 40?µg/mL) when compared with folate receptor α negative cells (31% for 25?µg/mL of lipid and 23% for 40?µg/mL) and to systems without PEG-folate (≈13% to 16% for all tested conditions), supporting their selectivity towards the receptor. Overall, the results support these systems as appealing vectors for selective delivery of siRNA to cancer cells by folate receptor α-mediated internalization, aiming at future therapeutic applications of interest.     

pH-responsive polysaccharide-based polyelectrolyte complexes as nanocarriers for lysosomal delivery of therapeutic proteins

Nanopharmaceutics composed of a carrier and a protein have the potential to improve the activity of therapeutical proteins. Therapy for lysosomal diseases is limited by the lack of effective protein delivery systems that allow the controlled release of specific proteins to the lysosomes. Here we address this problem by developing functional polyelectrolyte-based nanoparticles able to promote acidic pH-triggered release of the loaded protein. Trimethyl chitosan (TMC) was synthesized and allowed to form polyelectrolyte complexes (PECs) with the lysosomal enzyme α-GAL through self-assembly and ionotropic gelation, with average particle size <200 nm, polydispersity index (PDI) <0.2, ζ potential of ～ 20 mV, and a protein loading efficiency close to 65%. These polyelectrolyte nanoparticles were stable and active under physiological conditions and able to release the enzyme at acidic pH, as demonstrated by in situ atomic force microscopy (AFM). These nanoparticles were further functionalized with Atto 647N for single-particle characterization and tracking their cellular uptake and fate using high-resolution fluorescence microscopy. In contrast with their precursor, TMC, PECs were efficiently internalized by human endothelial cells and mostly accumulated in lysosomal compartments. The superior physicochemical characteristics of the TMC/α-GAL PECs together with their excellent cellular uptake properties indicate their enormous potential as advanced protein delivery systems for the treatment of lysosomal storage diseases.     

Receptor-targeted cancer therapy

Insight into the molecular mechanisms of malignant transformation is changing the way cancer is being treated. Conventional treatment strategies target the DNA of all dividing cells, resulting in a significantly increased risk of collateral toxicity. In addition, the accumulation of multiple mutations leads to drug resistance in many cancer cells. Targeted strategies have now been developed that specifically disrupt oncogenically active cell surface receptors and endogenous signaling molecules. These agents have a much greater selectivity for tumor tissue and decreased risk of side effects. Increased signaling through ErbB receptors via gene amplification, overexpression, and mutation has been implicated in many human cancers and associated with poor prognosis. Interruption of this process has been shown to cause antitumor effects. Downregulation of the ErbB receptors, HER-2/neu, and later EGFR, with monoclonal antibodies was the first demonstration of targeted therapy. Subsequently, the ErbB tyrosine kinase domain has been successfully targeted with small molecule inhibitors. The development of novel ErbB-directed entities is ongoing, with particular promise being shown by strategies targeting receptor interaction in oligomeric complexes.     

Stimuli-responsive nanocarriers for intracellular delivery

The emergence of different nanoparticles (NPs) has made a significant revolution in the field of medicine. Different NPs in the form of metallic NPs, dendrimers, polymeric NPs, carbon quantum dots and liposomes have been functionalized and used as platforms for intracellular delivery of biomolecules, drugs, imaging agents and nucleic acids. These NPs are designed to improve the pharmacokinetic properties of the drug, improve their bioavailability and successfully surpass physiological or pathological obstacles in the biological system so that therapeutic efficacy is achieved. In this review I present some of the current approaches used in intracellular delivery systems, with a focus on various stimuli-responsive nanocarriers, including cell-penetrating peptides, to highlight their various biomedical applications.     

Lectin functionalized nanocarriers for gene delivery

Gene therapy has emerged as one of the most promising therapeutic methods to treat various diseases. However, inadequate gene transfection efficacy during gene therapy demands further development of more efficient gene delivery strategies. Targeting genetic material to specific sites of action endows numerous advantages over non-targeted delivery. An ample variety of non-viral gene delivery vectors have been developed in recent years owing to the safety issues raised by viral vectors. Non-viral gene delivery vectors containing specific targeting ligands on their surfaces have been reported to enhance the gene transfection efficiency via receptor-mediated endocytosis for gene delivery. Among various targeting moieties investigated, carbohydrates and lectins (carbohydrate-binding proteins) played an essential role in gene delivery via either direct or reverse lectin targeting strategies. Lectins have a specific carbohydrate binding domain that can bind specifically to the carbohydrates. This review sheds light on various gene delivery nanovectors conjugated with either lectins or carbohydrates for enhanced gene transfection.     

Exploiting BBB disruption for the delivery of nanocarriers to the diseased CNS

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Gene delivery by functional inorganic nanocarriers

Gene delivery into cells to elicit cellular response has received a great attention recently. Viruses, lipids, peptides, cationic polymers and certain inorganic nanomaterials have been reported as gene delivery vectors. In this review, we focus on the recent literature on gene delivery using inorganic nanoparticles. This emerging field of study is concisely summarized and illustrated by selected examples and recent patents. New approaches and directions towards the practical use of multifunctional nanocarriers are highlighted.     

Factorial design based curcumin ethosomal nanocarriers for the skin cancer delivery: in vitro evaluation

Abstract

Melanoma is the most deadly and life-threatening form of skin cancer with progressively higher rates of incidence worldwide. The objective of the present investigation is to develop and to statistically optimize and characterize curcumin (CUR) loaded ethosomes for treatment of melanoma. A two factor, three level (3²) factorial design approach was employed for the optimization of ethosomes. The prepared ethosomes were evaluated for size, zeta potential, entrapment efficiency, in vitro skin permeation and deposition ability. The optimized ethosomal formulation was evaluated for in vitro cytotoxicity and cellular uptake studies using A375 human melanoma cells. The optimized formulation has imperfect round shaped unilamellar structures with a mean vesicle size of 247?±?5.25?nm and an entrapment efficiency of 92.24?±?0.20%. The in vitro skin permeation studies proved the superiority of ethosomes over the traditional liposomes in terms of the amount of drug permeated and deposited in skin layers. Fluorescence microscopy showed the enhanced penetration of ethosomes into the deeper layers of the skin. In vitro cytotoxicity and cellular uptake studies revealed that curcumin ethosomes have significantly improved cytotoxicity and cellular uptake in A375 human melanoma cell lines. The colony formation assay results showed that curcumin ethosomes have a superior antiproliferative effect as they effectively inhibit the clonogenic ability of A375 cells. The flow cytometry results indicate that curcumin ethosomes induce cell death in A375 cells by apoptosis mechanism. The present study provides a strong rationale and motivation for further investigation of newly developed curcumin ethosomes as a potential therapeutic strategy for melanoma treatment.     

Receptor-targeted liposomal delivery of boron-containing cholesterol mimics for boron neutron capture therapy (BNCT)

Liposomes have been a main focus of tumor-selective boron delivery strategies in boron neutron capture therapy (BNCT), a binary method for the treatment of cancer that is based on the nuclear reaction between boron atoms and low-energy thermal neutrons. Three novel carboranyl cholesterol derivatives were prepared as lipid bilayer components for the construction of nontargeted and receptor-targeted boronated liposomes for BNCT. A major structural feature of these novel boronated cholesterol mimics is the replacement of the B and the C ring of cholesterol with a carborane cluster. Computational analyses indicated that all three boronated compounds have structural features and physicochemical properties that are very similar to those of cholesterol. One of the synthesized boronated cholesterol mimics was stably incorporated into non-, folate receptor (FR)-, and vascular endothelial growth factor receptor-2 (VEGFR-2)-targeted liposomes. No major differences were found in appearance, size distribution, and lamellarity between conventional dipalmitoylphosphatidylcholine (DPPC)/cholesterol liposomes, nontargeted, and FR-targeted liposomal formulations of this carboranyl cholesterol derivative. FR-targeted boronated liposomes were taken up extensively in FR overexpressing KB cells in vitro, and the uptake was effectively blocked in the presence of free folate. In contrast, a boronated cholesterol mimic incorporated into nontargeted liposomes showed significantly lower cellular uptake. There was no apparent in vitro cytotoxicity in FR overexpressing KB cells and VEGFR-2 overexpressing 293/KDR cells when these were incubated with boronated FR- and (VEGFR-2)-targeted liposomes, respectively, although the former accumulated extensively in KB cells and the latter effectively interacted with VEGFR-2 by causing autophosphorylation and protecting 293/KDR cells from SLT (Shiga-like toxin)-VEGF cytotoxicity.     

A comprehensive overview of exosomes as drug delivery vehicles — Endogenous nanocarriers for targeted cancer therapy

Exosomes denote a class of secreted nanoparticles defined by size, surface protein and lipid composition, and the ability to carry RNA and proteins. They are important mediators of intercellular communication and regulators of the cellular niche, and their altered characteristics in many diseases, such as cancer, suggest them to be important both for diagnostic and therapeutic purposes, prompting the idea of using exosomes as drug delivery vehicles, especially for gene therapy. This review covers the current status of evidence presented in the field of exosome-based drug delivery systems. Components for successful exosome-based drug delivery, such as choice of donor cell, therapeutic cargo, use of targeting peptide, loading method and administration route are highlighted and discussed with a general focus pertaining to the results obtained in models of different cancer types. In addition, completed and on-going clinical trials are described, evaluating exosome-based therapies for the treatment of different cancer types. Due to their endogenous origin, exosome-based drug delivery systems may have advantages in the treatment of cancer, but their design needs further refinement to justify their usage on the clinical scale.     

Cell penetrating peptide-modified pharmaceutical nanocarriers for intracellular drug and gene delivery

Cell-penetrating peptides (CPPs) including TAT peptide (TATp) have been successfully used for intracellular delivery of a broad variety of cargos including various nanoparticulate pharmaceutical carriers (liposomes, micelles, nanoparticles). Here, we will consider the main results in this area, with a special emphasis on TATp-mediated delivery of liposomes and DNA. We will also address the development of "smart" stimuli-sensitive nanocarriers, where cell-penetrating function can be activated by the decreased pH only inside the biological target minimizing thus the interaction of drug-loaded nanocarriers with nontarget cells.     

Design and evaluation of multifunctional nanocarriers for selective delivery of coenzyme Q10 to mitochondria

Impairments of mitochondrial functions have been associated with failure of cellular functions in different tissues, leading to various pathologies. We report here a mitochondria-targeted nanodelivery system for coenzyme Q10 (CoQ10) that can reach mitochondria and deliver CoQ10 in adequate quantities. Multifunctional nanocarriers based on ABC miktoarm polymers (A = poly(ethylene glycol (PEG), B = polycaprolactone (PCL), and C = triphenylphosphonium bromide (TPPBr)) were synthesized using a combination of click chemistry with ring-opening polymerization, self-assembled into nanosized micelles, and were employed for CoQ10 loading. Drug loading capacity (60 wt %), micelle size (25-60 nm), and stability were determined using a variety of techniques. The micelles had a small critical association concentration and were colloidally stable in solution for more than 3 months. The extraordinarily high CoQ10 loading capacity in the micelles is attributed to good compatibility between CoQ10 and PCL, as indicated by the low Flory-Huggins interaction parameter. Confocal microscopy studies of the fluorescently labeled polymer analog together with the mitochondria-specific vital dye label indicated that the carrier did indeed reach mitochondria. The high CoQ10 loading efficiency allowed testing of micelles within a broad concentration range and provided evidence for CoQ10 effectiveness in two different experimental paradigms: oxidative stress and inflammation. Combined results from chemical, analytical, and biological experiments suggest that the new miktoarm-based carrier provides a suitable means of CoQ10 delivery to mitochondria without loss of drug effectiveness. The versatility of the click chemistry used to prepare this new mitochondria-targeting nanocarrier offers a widely applicable, simple, and easily reproducible procedure to deliver drugs to mitochondria or other intracellular organelles.     

Strategies for ocular siRNA delivery: Potential and limitations of non-viral nanocarriers

ABSTRACT: Controlling gene expression via small interfering RNA (siRNA) has opened the doors to a plethora of therapeutic possibilities, with many currently in the pipelines of drug development for various ocular diseases. Despite the potential of siRNA technologies, barriers to intracellular delivery significantly limit their clinical efficacy. However, recent progress in the field of drug delivery strongly suggests that targeted manipulation of gene expression via siRNA delivered through nanocarriers can have an enormous impact on improving therapeutic outcomes for ophthalmic applications. Particularly, synthetic nanocarriers have demonstrated their suitability as a customizable multifunctional platform for the targeted intracellular delivery of siRNA and other hydrophilic and hydrophobic drugs in ocular applications. We predict that synthetic nanocarriers will simultaneously increase drug bioavailability, while reducing side effects and the need for repeated intraocular injections. This review will discuss the recent advances in ocular siRNA delivery via non-viral nanocarriers and the potential and limitations of various strategies for the development of a 'universal' siRNA delivery system for clinical applications.     

Diatomite biosilica nanocarriers for siRNA transport inside cancer cells

BackgroundDiatomite is a natural porous biomaterial of sedimentary origin, formed by fragments of diatom siliceous skeletons, called “frustules”. Due to large availability in many areas of the world, chemical stability, and non-toxicity, these fossil structures have been widespread used in lot of industrial applications, such as food production, water extracting agent, production of cosmetics and pharmaceutics. However, diatomite is surprisingly still rarely used in biomedical applications. In this work, we exploit diatomite nanoparticles for small interfering ribonucleic acid (siRNA) transport inside human epidermoid cancer cells (H1355).MethodsMorphology and composition of diatomite microfrustules (average size lower than 40 μm) are investigated by scanning electron microscopy equipped by energy dispersive X-ray spectroscopy, Fourier transform infrared analysis, and photoluminescence measurements. Nanometric porous particles (average size lower than 450 nm) are obtained by mechanical crushing, sonication, and filtering of micrometric frustules. siRNA bioconjugation is performed on both micrometric and nanometric fragments by silanization.ResultsIn-vitro experiments show very low toxicity on exposure of the cells to diatomite nanoparticle concentration up to 300 μg/ml for 72 h. Confocal microscopy imaging performed on cancer cells incubated with siRNA conjugated nanoparticles demonstrates a cytoplasmatic localization of vectors. Gene silencing by delivered siRNA is also demonstrated.ConclusionOur studies endorse diatomite nanoparticles as non-toxic nanocarriers for siRNA transport in cancer cells.General significancesiRNA is a powerful molecular tool for cancer treatment but its delivery is inefficient due to the difficulty to penetrate the cell membrane. siRNA-diatomite nanoconjugate may be well suited for delivery of therapeutic to cancer cells.     

Rational design for multifunctional non-liposomal lipid-based nanocarriers for cancer management: theory to practice

Nanomedicines have gained more and more attention in cancer therapy thanks to their ability to enhance the tumour accumulation and the intracellular uptake of drugs while reducing their inactivation and toxicity. In parallel, nanocarriers have been successfully employed as diagnostic tools increasing imaging resolution holding great promises both in preclinical research and in clinical settings. Lipid-based nanocarriers are a class of biocompatible and biodegradable vehicles that provide advanced delivery of therapeutic and imaging agents, improving pharmacokinetic profile and safety. One of most promising engineering challenges is the design of innovative and versatile multifunctional targeted nanotechnologies for cancer treatment and diagnosis. This review aims to highlight rational approaches to design multifunctional non liposomal lipid-based nanocarriers providing an update of literature in this field.     

Formulations for delivery of therapeutic proteins

Advances in biotechnology have now created a capacity to produce therapeutically active proteins on a commercial scale, opening the potential for their application in an array of disease conditions. The process of translation of the variety of different therapeutic proteins into the medicines used in clinics is now occurring. To assist in this translation, new formulations to deliver proteins could play an important role. These new formulations need to more adequately address the pharmacological and therapeutic requirement for each particular protein/peptide and, in that way, either improve present therapies or extend with new entries the current list of protein based medicines used in clinic.

"SMART" drug delivery systems: double-targeted pH-responsive pharmaceutical nanocarriers

To develop targeted pharmaceutical carriers additionally capable of responding to certain local stimuli, such as decreased pH values in tumors or infarcts, targeted long-circulating PEGylated liposomes and PEG-phosphatidylethanolamine (PEG-PE)-based micelles have been prepared with several functions. First, they are capable of targeting a specific cell or organ by attaching the monoclonal antimyosin antibody 2G4 to their surface via pNP-PEG-PE moieties. Second, these liposomes and micelles were additionally modified with biotin or TAT peptide (TATp) moieties attached to the surface of the nanocarrier by using biotin-PE or TATp-PE or TATp-short PEG-PE derivatives. PEG-PE used for liposome surface modification or for micelle preparation was made degradable by inserting the pH-sensitive hydrazone bond between PEG and PE (PEG-Hz-PE). Under normal pH values, biotin and TATp functions on the surface of nanocarriers were "shielded" by long protecting PEG chains (pH-degradable PEG(2000)-PE or PEG(5000)-PE) or by even longer pNP-PEG-PE moieties used to attach antibodies to the nanocarrier (non-pH-degradable PEG(3400)-PE or PEG(5000)-PE). At pH 7.4-8.0, both liposomes and micelles demonstrated high specific binding with 2G4 antibody substrate, myosin, but very limited binding on an avidin column (biotin-containing nanocarriers) or internalization by NIH/3T3 or U-87 cells (TATp-containing nanocarriers). However, upon brief incubation (15-30 min) at lower pH values (pH 5.0-6.0), nanocarriers lost their protective PEG shell because of acidic hydrolysis of PEG-Hz-PE and acquired the ability to become strongly retained on an avidin column (biotin-containing nanocarriers) or effectively internalized by cells via TATp moieties (TATp-containing nanocarriers). We consider this result as the first step in the development of multifunctional stimuli-sensitive pharmaceutical nanocarriers.