Application of heparinized cellulose affinity membranes in recombinant adeno-associated virus serotype 2 binding and delivery

The microporous affinity membrane based on cellulose matrices offers minimal mass-transfer effects in membrane chromatography with low nonspecific adsorption. In this research, we tested a novel application of the microporous, heparinized cellulose membranes for their affinity toward recombinant adeno-associated virus serotype 2 (rAAV2, which uses heparan sulfate proteoglycans as the primary cellular receptor) to develop a controlled, substrate-mediated viral vector delivery. We conjugated rAAV2 to an epoxy-crosslinked heparin cellulose membrane, which led to vector transduction upon cellular adhesion. When adhered, human fibroblasts exhibited proliferation kinetics similar to those on the standard polystyrene tissue-culture surface. Using fluorescent proteins as the reporter, we showed that the heparin-bound rAAV2 particles remained active and that the rAAV2-heparin binding was reversible and capable of mediating transgene delivery in cell culture. In addition, we applied the affinity membrane to adsorb unpurified rAAV2 from the crude lysate of packaging cells via the ligand–receptor binding, avoiding the use of conventional ultracentrifugation or chromatography in preparation of infectious rAAV2 for transduction. Our work explores a new application of affinity cellulose membranes in substrate-mediated viral vector delivery, which can be a useful tool in developing protocols for localized gene transfer.     

Application of cellulose acetate butyrate-based membrane for osmotic drug delivery

The release rate of drugs from an OROS^® is controlled by semipermeable membranes composed typically of cellulose acetate (CA) with various flux enhancers. Cellulose acetate butyrate (CAB) was identified as a viable alternative. The CAB membrane matched the CA membrane in robustness but had superior drying properties, offering particular advantages for thermolabile formulations. Studies were conducted to characterize CAB membrane properties with respect to performance of OROS^® systems. Four different membrane formulations with varying plasticizer type and concentration were investigated. The CAB based membranes exhibited superior drying characteristics and similar functionality to the CA:polyethylene glycol (PEG) membranes used as a control. A linear relationship was observed between the level of flux enhancer and release rate. The stability of the membrane was evaluated based on release profiles after system storage at various conditions. The CAB membranes appeared to have stability comparable to the standard CA membrane. A linear relationship between membrane weight and release rate as well as the time required to release 90% of a drug from the system [T₉₀] for a model formulation was observed. In conclusion, the newly identified alternative membrane composition allows for the use of thinner membranes, thereby reducing cost of goods, coating time and, most importantly, membrane drying time.     

Application of solid phase peptide synthesis to engineering PEO–peptide block copolymers for drug delivery

This work describes a simple, versatile solid-phase peptide-synthesis (SPPS) method for preparing micelle-forming poly(ethylene oxide)-block-peptide block copolymers for drug delivery. To demonstrate its utility, this SPPS method was used to construct two series of micelle-forming block copolymers (one of constant core-composition and variable length; the other of constant core length and variable composition). The block copolymers were then used to study in detail the effect of size and composition on micellization. The various block copolymers were prepared by a combination of SPPS for the peptide block, followed by solution–phase conjugation of the peptide block with a proprionic acid derivative of poly(ethylene oxide) (PEO) to form the PEO-b-peptide block copolymer. The composition of each block component was characterized by mass spectrometry (MALDI and ES-MS). Block copolymer compositions were characterized by ¹H NMR. All the block copolymers were found to form micelles as judged by transmission electron microscopy (TEM) and light scattering analysis. To demonstrate their potential as drug delivery systems, micelles prepared from one member of the PEO-b-peptide block copolymer series were physically loaded with the anticancer drug doxorubicin (DOX). Micelle static and dynamic stability were found to correlate strongly with micelle core length. In contrast, these same micellization properties appear to be a complex function of core composition, and no clear trends could be identified from among the set of compositionally varying, fixed length block copolymer micelles. We conclude that SPPS can be used to construct biocompatible block copolymers with well-defined core lengths and compositions, which in turn can be used to study and to tailor the behavior of block copolymer micelles.     

Functionalized silica matrices for controlled delivery of cephalexin

The surface of SBA-15 ordered mesoporous silica was functionalized with sulphonic and amine functional groups to determine the effect of changes in surface acidity on cephalexin adsorption and subsequent release. Cephalexin (CPX), which belongs to the group of cephalosporins or β-lactam antibiotics, was impregnated on functionalized SBA-15. The functionalized silica materials were characterized by SEM, TGA, FTIR and N₂ adsorption, and an in vitro drug delivery test was performed. SEM micrographs showed the packed cylinders correspond to SBA 15 materials. Likewise, N₂ adsorption–desorption isotherms demonstrated that the SBA-15 structure was obtained when IV-type isotherms were displayed. The inalterable stabilization of the drug was confirmed by FTIR spectroscopy. For all the samples studied, the delivery profiles exhibit two steps. A fast initial stage was obtained over the first 5 h, followed by a slower one. Regarding this second stage, the time needed to attain a plateau was undoubtedly altered by the surface functionalization.     

Preparation of mannan modified anionic PCL-PEG-PCL nanoparticles at one-step for bFGF antigen delivery to improve humoral immunity

In this article, blank anionic poly(-caprolactone)-poly(ethylene glycol)-poly(-caprolactone) (PCEC) and anionic mannan modified PCEC (MPCEC) nanoparticles with nearly the same particle size and zeta potential were prepared by emulsion solvent evaporation method. Human basic fibroblast growth factor (bFGF) was absorbed onto anionic nanoparticles surface due to electrostatic interaction. The obtained bFGF-nanoparticles complexes were injected subcutaneously into C57BL/6 mice at 20 μg of bFGF/dose on week 0, 1, 2 and 3. The mice serum was collected on week 4, and bFGF-specific autoantibody total IgG, IgG1 and IgG2a titer in serum was determined by ELISA. The results indicated that the autoantibody IgG, IgG1 and IgG2a titer of the mice immunized by bFGF–MPCEC complexes were higher than that immunized by either bFGF–PCEC or bFGF–Alum. This phenomenon might be due to that mannan functionalized MPCEC nanoparticles could be targeted to dendritic cells (DCs) to improve humoral immunity. The prepared anionic mannan modified PCEC nanoparticles (MPCEC) might have great potential application in vaccine delivery systems.     

Superabsorbent hydrogels based on cellulose for smart swelling and controllable delivery

Novel superabsorbent hydrogels were prepared successfully from carboxymethylcellulose sodium (CMC) and cellulose in the NaOH/urea aqueous system by using epichlorohydrin (ECH) as cross-linker. The structure and morphology of the hydrogels were characterized by FT-IR spectroscope, thermogravimetric analysis and scanning electron microscope. The results revealed that the CMC contributed to the enhanced size of pore, whereas cellulose as a strong backbone in the hydrogel to support it for keeping its appearance. Their equilibrium swelling ratio in distilled water and different physiological fluids were evaluated, indicating the maximum swelling ratio in water reached an exciting level of 1000 as the hydrogels still keeping a steady appearance. Moreover, the hydrogels exhibited smart swelling and shrinking in NaCl or CaCl₂ aqueous solution, as well as the release behavior of bovine serum albumin (BSA) that could be controlled by changing CMC content. The cellulose-based hydrogels are promising for the applications in the biomaterials area.     

The potential of TEMPO-oxidized nanofibrillar cellulose beads for cell delivery applications

The advanced development of cell carriers for regenerative medicine and cell therapy demands materials able to sustain cell viability prior to their delivery to the target tissue, an ability that can be controlled by the shape, size and degradability of the matrix. TEMPO-oxidized nanofibrillar cellulose (ToNFC) macromolecules are negatively charged and therefore can be easily formulated by ionotropic gelation into beads of varying sizes that can release their payload through an erosion-controlled process. We report here for the first time on the preparation of ToNFC beads via ionic gelation using CaCl₂ and on their loading with OSTEO-1 rat bone cells, with a view to examine their capacity of sustaining the cell viability and of releasing the bone cells in a controlled manner. The initial results obtained demonstrate that ToNFC is able to protect the OSTEO-1 cells and to maintain their viability for at least 2 weeks. Following gradual disintegration of the beads, a significant cell release and subsequent proliferation was observed after 7 days. These results indicate the considerable potential of nanofibrillar cellulose (ToNFC) for applications in cell therapy and regenerative medicine.     

Application of silicon dioxide nanoparticles modified with tumor-targeting ligands for cellular delivery of nucleoside triphosphate analogues

A key advantage of amino-modified SiO₂ nanoparticles for delivery of phosphorylated nucleosides is a broad possibility for functionalization. It can be modified with ligands currently investigated in targeted drug delivery. To improve the efficacy for intracellular delivery, SiO₂ nanoparticles were functionalized with tumor-targeting ligands folic acid, biotin or 5-fluorouracil. Studies of accumulation of these conjugates in HCT116 colon carcinoma cells revealed that the uptake of modified conjugates was significantly bigger compared to unmodified nanoparticles, with the biotinylated conjugate as the preferred compound. The nanocomposites of biotin modified SiO₂ and 2′,3′-dideoxyuridine triphosphate showed a pronounced antiproliferative potency relative to the unmodified nanocomposites. Thus, multi-functionalization of SiO₂ nanoparticle based conjugates has a major potential for delivery of nucleoside triphosphate analogues, therefore tentatively enhancing their bioactivity.     

Modification of cellulose by graft polymerization for use in drug delivery systems

Cellulose-based biodegradable polymers—as microspheres or hydrogels—are suitable for drug delivery systems. In this work, cellulose microfibers were converted to cellulose esters for subsequent graft copolymerization either by free radical or atom transfer radical polymerization (ATRP). For the former, carboxymethyl cellulose (CMC) was prepared and then modified through grafting of poly(hydroxyethyl acrylate) or polyacrylamide. ATRP was achieved by chloroacetylation of cellulose followed by graft copolymerization of hydroxyethyl acrylate or acrylamide monomers. The degree of substitution for CMC and chloroacetylated cellulose (CAC) was determined by the method described in US Pharmacopeia NF24 and by titration method, respectively. CMC, CAC, and the grafted copolymers were characterized by Fourier transform infrared spectroscopy, differential scanning calorimetry, thermal gravimetric analysis, X-ray diffraction, and atomic force microscopy; the latter technique clearly shows the chain growth of the synthetic polymers on the backbone surface. Furthermore, cephalexin antibiotic was loaded on the copolymers, and the resultant in vitro drug release studied in three different media (buffer solutions with pH equal to 3, 6.1, and 8).     

Application of dynamic 2D FTIR to cellulose

Cellulose, the dominant polymer in the biosphere, is a homopolysaccharide composed of (1,4)-β-d-glucopyranose. Interactions between and within the cellulose polymer chains are mainly determined by inter- and intramolecular hydrogen bonds, which are therefore mainly responsible for mechanical properties of cellulosic materials. The coupling of dynamic mechanical analysis (DMA) and 2D step-scan Fourier transform infrared (FTIR) spectroscopy, is shown to be a very promising way of investigating these submolecular interactions in cellulosic materials. The broad and unstructured band in the OH-stretching vibration region (3100 and 3700 cm⁻¹) of the cellulose vibrational spectra, which contains information about the intra- and intermolecular hydrogen bonds, can be unraveled by this new technique. In the experiments reported here, cellulose sheets have been stretched sinusoidally at low strains while being irradiated with polarized infrared light. For the obtained dynamic IR signals (the in-phase and the out-of-phase responses of the sample), the dynamic IR cross-correlation was defined. It consists of two terms which are referred to as the synchronous and the asynchronous 2D infrared correlation intensities. In the 2D spectra, obtained by DMA–FTIR, several distinct peaks are observed in the OH-range between 3700 and 3100 cm⁻¹ which may be related to specific interactions.     

Surface modification of cellulose nanocrystal with chitosan oligosaccharide for drug delivery applications

A novel drug delivery system based on two of the most abundant natural biopolymers was developed by modifying the surface of oxidized cellulose nanocrystal (CNC) with chitosan oligosaccharide (CS_OS). First, the primary alcohol moieties of CNC were selectively oxidized to carboxyl groups using the 2,2,6,6-tetramethylpiperidine-1-oxyl radical catalyst. The amino groups of CS_OS were then reacted with carboxylic acid groups on oxidized CNC (CNC-OX) via the carbodiimide reaction using N-hydroxysuccinimide and 1-ethyl-3-(3-dimethylaminopropyl)-carbodiimide as coupling agents. Successful grafting of CS_OS to CNC-OX was confirmed by infrared spectroscopy, thermogravimetry, potentiometric titration, and zeta potential measurements. The grafting resulted in a conversion of ~90 % carboxyl groups on CNC-OX and the degree of substitution was 0.26. CNC–CS_OS nanoparticles showed a binding efficiency of 21.5 % and a drug loading of 14 % w/w. A drug selective electrode was used to directly measure the concentration of procaine hydrochloride released from CNC–CS_OS particles. The in vitro drug release was studied at pH 8 and the nanoparticles revealed a fast release of up to 1 h, which can be used as biocompatible and biodegradable drug carriers for transdermal delivery applications.     

A peptide aldehyde microarray for high-throughput profiling of cellular events

Microarrays provide exciting opportunities in the field of large-scale proteomics. With the aim to elucidate enzymatic activity and profiles within native biological samples, we developed a microarray comprising a focused positional-scanning library of enzyme inhibitors. The library was diversified across P(1)-P(4) positions, creating 270 different inhibitor sublibraries which were immobilized onto avidin slides. The peptide aldehyde-based small-molecule microarray (SMM) specifically targeted cysteine proteases, thereby enabling large-scale functional assessment of this subgroup of proteases, within fluorescently labeled samples, including pure proteins, cellular lysates, and infected samples. The arrays were shown to elicit binding fingerprints consistent with those of model proteins, specifically caspases and purified cysteine proteases from parasites (rhodesein and cruzain). When tested against lysates from apoptotic Hela and red blood cells infected with Plasmodium falciparum, clear signatures were obtained that were readily attributable to the activity of constituent proteases within these samples. Characteristic binding profiles were further able to distinguish various stages of the parasite infection in erythrocyte lysates. By converting one of our brightest microarray hits into a probe, putative protein markers were identified and pulled down from within apoptotic Hela lysates, demonstrating the potential of target validation and discovery. Taken together, these results demonstrate the utility of targeted SMMs in dissecting cellular biology in complex proteomic samples.     

Analytical characterization of chitosan nanoparticles for peptide drug delivery applications

Chitosan-cyclodextrin hybrid nanoparticles (NPs) were obtained by the ionic gelation process in the presence of glutathione (GSH), chosen as a model drug. NPs were characterized by means of transmission electron microscopy and zeta-potential measurements. Furthermore, a detailed X-ray photoelectron spectroscopy study was carried out in both conventional and depth-profile modes. The combination of controlled ion-erosion experiments and a scrupulous curve-fitting approach allowed for the first time the quantitative study of the GSH in-depth distribution in the NPs. NPs were proven to efficiently encapsulate GSH in their inner cores, thus showing promising perspectives as drug carriers.     

Invert and double emulsions as a base for microheterogeneous matrices for transdermal delivery of lipophilic drugs

Stable invert emulsions water/oil and double emulsions oil₁/water/oil₂ containing polyisobuthylene as pressure-sensitive polymeric adhesive are prepared. The dispersed phase of the invert emulsions contained micelles of the surfactant Tween 80 (Tw) with solubilized drug (Felodipine) and a skin permeation enhancer (glycerol monooleate, GMO). The active components (Felodipine and GMO) of the double emulsions were incorporated into the internal dispersed phase, while the intermediate water layer contained Tw and hydroxypropylcellulose. Ultradispersed polymer films with good adhesion to skin were prepared from both types of emulsions. The films based on double emulsions demonstrate the ability to release Felodipine at therapeutically effective levels and maintain these levels during the first 24 h to attain a therapeutically required dose. The invert and double emulsions were for the first time utilized as bases for microheterogeneous matrices for delivery of a lipophilic drug in bioavailable form.     

Nanotechnologies for drug delivery: Application to cancer and autoimmune diseases

Polymer-based nanotechnologies are now proposed as an alternative to classical formulations for drug administration, delivery and targeting. Therapeutic applications of the first generation of nanotechnologies include the treatment of cancer liver diseases. Avoiding the recognition by the liver is also possible by developing long circulating polymeric colloidal carriers (“stealth” systems) able to avoid the opsonization process and the recognition by the macrophages. The design of such carriers of second generation is based on the physico-chemical concept of the “steric repulsion”: by grafting polyethyleneglycol chains at the surface of nanoparticles, the adsorption of seric proteins may be dramatically reduced due to steric hindrance. Such an approach allows maintaining the drug carrier for a longer time into the circulation and the resulting extravasation towards non reticuloendothelial-located cancers may become possible. Now, new applications and exciting perspectives are proposed for the delivery of drugs to previously non accessible diseased sanctuaries, like the brain (treatment of glioma and autoimmune diseases of the brain) or the ocular tissues (treatment of the autoimmune uveitis). Finally, the use of nanotechnologies for the delivery of nucleic acids (oligonucleotides) is also discussed in this review.     

Applications of bacterial cellulose in food,cosmetics and drug delivery

Bacterial cellulose (BC) is a versatile biopolymer with better material properties, such as purity, high degree of porosity, relative high permeability to liquid and gases, high water-uptake capacity, tensile strength and ultrafine network. This review explores the applications of BC and its hydrogels in the fields of food, cosmetics and drug delivery. Applications of BC in foods are ranging from traditional dessert, low cholesterol diet, vegetarian meat, and as food additive and dietary aid to novel applications, such as immobilization of enzymes and cells. Applications in cosmetics include facial mask, facial scrub, personal cleansing formulations and contact lenses. BC for controlled drug delivery, transdermal drug delivery, dental drug delivery, protein delivery, tissue engineering drug delivery, macromolecular prodrug delivery and molecularly imprinted polymer based enantioselective drug delivery are also discussed in this review. The applications of BC in food and cosmetics provide the basis for BC-based functional foods, nutraceuticals, cosmeceuticals and medicated cosmetics. On the basis of current studies, the BC-based drug delivery could be further fine-tuned to get more sophisticated control on stimuli-responsive drug release. Along with the currently available literature, further experiments are required to obtain a blueprint of drug in vivo performance, bioavailability and in vitro–in vivo correlation.     

Estrogen-derived steroidal metal complexes: agents for cellular delivery of metal centers to estrogen receptor-positive cells

Targeted cellular delivery of drugs to specific tissues is an important goal in biomedical chemistry. Achieving this requires harnessing and applying molecular-level recognition events prevalent in (or specific to) the desired tissue type. Tissues rich in estrogen receptors (ERs), which include many types of breast cancer, accumulate molecules that have high binding affinities for these receptors. Therefore, molecules that (i) bind to the ER, (ii) have favorable cellular transport properties, and (iii) contain a second functionality (such as a center that may be used for diagnostic imaging or medical therapy) are exciting synthetic targets in the field of drug delivery. To this end, we have prepared a range of metallo-estrogens based on 17alpha-ethynylestradiol and examined their binding to the ER both as isolated receptor and in whole cell assays (ER positive MCF-7 cells). Estrogens functionalized with metal binding units are prepared by palladium-catalyzed cross-coupling reactions and a wide range of metal centers introduced readily. All the compounds prepared and tested exhibit effective binding to the estrogen receptor and are delivered across the cell membrane into MCF-7 cells. In the whole cell assays, despite their monocationic nature, the palladium and platinum complexes prepared exhibit similar (and even enhanced) receptor binding affinities compared to their corresponding neutral free ligands. It is unprecedented for a higher ER binding affinity to be observed for a cationic complex than for its metal-free ligand.     

Degradable self-assembling dendrons for gene delivery: experimental and theoretical insights into the barriers to cellular uptake

This paper uses a combined experimental and theoretical approach to gain unique insight into gene delivery. We report the synthesis and investigation of a new family of second-generation dendrons with four triamine surface ligands capable of binding to DNA, degradable aliphatic-ester dendritic scaffolds, and hydrophobic units at their focal points. Dendron self-assembly significantly enhances DNA binding as monitored by a range of experimental methods and confirmed by multiscale modeling. Cellular uptake studies indicate that some of these dendrons are highly effective at transporting DNA into cells (ca. 10 times better than poly(ethyleneimine), PEI). However, levels of transgene expression are relatively low (ca. 10% of PEI). This indicates that these dendrons cannot navigate all of the intracellular barriers to gene delivery. The addition of chloroquine indicates that endosomal escape is not the limiting factor in this case, and it is shown, both experimentally and theoretically, that gene delivery can be correlated with the ability of the dendron assemblies to release DNA. Mass spectrometric assays demonstrate that the dendrons, as intended, do degrade under biologically relevant conditions over a period of hours. Multiscale modeling of degraded dendron structures suggests that complete dendron degradation would be required for DNA release. Importantly, in the presence of the lower pH associated with endosomes, or when bound to DNA, complete degradation of these dendrons becomes ineffective on the transfection time scale-we propose this explains the poor transfection performance of these dendrons. As such, this paper demonstrates that taking this kind of multidisciplinary approach can yield a fundamental insight into the way in which dendrons can navigate barriers to cellular uptake. Lessons learned from this work will inform future dendron design for enhanced gene delivery.     

Stability and solution concentration enhancement of resveratrol by solid dispersion in cellulose derivative matrices

Resveratrol is a highly biologically active phytoalexin, found in many plant materials that are common elements of the human diet, such as grapes, nuts, and red wine. The therapeutic or disease preventative potential of this natural polyphenolic antioxidant has been limited in part due to its poor aqueous solubility and low oral bioavailability. We hypothesized that solid dispersion of resveratrol (Res) in cellulose derivative matrices might afford amorphous dispersions, from which supersaturated Res solutions would be produced in the human gastrointestinal (GI) tract, resulting in higher Res bioavailability. We carried out structure–property studies employing cellulose esters with a range of physical characteristics but possessing features suitable for use in amorphous solid dispersions: carboxymethylcellulose acetate butyrate (CMCAB), hydroxypropylmethylcellulose acetate succinate (HPMCAS) and cellulose acetate adipate propionate (CAAdP). The cellulose derivative results were compared with those of a negative control, pure crystalline Res, and a positive control, Res/poly(vinylpyrrolidinone) (PVP). Solid dispersions were characterized by powder X-ray diffraction (XRPD), modulated differential scanning calorimetry (MDSC), nuclear magnetic resonance (NMR) and Fourier transform infrared spectroscopy (FT-IR) of solid dispersions. HPMCAS and PVP solid dispersions afforded faster and more complete Res release at pH 6.8; however Res is also released from PVP matrices at pH 1.2. The carboxyl-containing cellulose derivatives release Res to only a small extent at pH 1.2. This combination of solution and solid phase stabilization against crystallization, and pH-triggered drug release makes these cellulose esters attractive candidates for Res bioavailability enhancement.