Jean-Michel Rabanel

Assessment of PEG on polymeric particles surface, a key step in drug carrier translation.

Injectable drug nanocarriers have greatly benefited in their clinical development from the addition of a superficial hydrophilic corona to improve their cargo pharmacokinetics. The most studied and used polymer for this purpose is poly(ethylene glycol), PEG. However, in spite of its wide use for over two decades now, there is no general consensus on the optimum PEG chain coverage-density and size required to escape from the mononuclear phagocyte system and to extend the circulation time. Moreover, cellular uptake and active targeting may have conflicting requirements in terms of surface properties of the nanocarriers which complicate even more the optimization process. These persistent issues can be largely attributed to the lack of straightforward characterization techniques to assess the coverage-density, the conformation or the thickness of a PEG layer grafted or adsorbed on a particulate drug carrier and is certainly one of the main reasons why so few clinical applications involving PEG coated particle-based drug delivery systems are under clinical trial so far. The objective of this review is to provide the reader with a brief description of the most relevant techniques used to assess qualitatively or quantitatively PEG chain coverage-density, conformation and layer thickness on polymeric nanoparticles. Emphasis has been made on polymeric particle (solid core) either made of copolymers containing PEG chains or modified after particle formation. Advantages and limitations of each technique are presented as well as methods to calculate PEG coverage-density and to investigate PEG chains conformation on the NP surface.

Effect of mechanical properties of hydrogel nanoparticles on macrophage cell uptake

Uptake and intracellular trafficking of hydrogel nanoparticles (NPs) of N,N-diethyl acrylamide and 2-hydroxyethyl methacrylate crosslinked with N,N′-methylene-bis-acrylamide were studied with a RAW 264.7 murine macrophage cell line. Results show that the uptake rate, the mechanism of internalization and the concentration of internalized NPs are correlated to the NP Young modulus. Soft NPs are found to be internalized preferentially via macropinocytosis while the uptake of stiff NPs is mediated by a clathrin-dependent mechanism. NPs with an intermediate Young modulus exhibit multiple uptake mechanisms. The accumulation rate of the NPs into lysosomal compartments of the cell is also dependent on the NP elasticity. Our results suggest that control over the mechanical properties of hydrogel NPs can be used to tailor the cellular uptake mechanism and kinetics of drug delivery.

Progress Technology in Microencapsulation Methods for Cell Therapy

Cell encapsulation in microcapsules allows the in situ delivery of secreted proteins to treat different pathological conditions. Spherical microcapsules offer optimal surface‐to‐volume ratio for protein and nutrient diffusion, and thus, cell viability. This technology permits cell survival along with protein secretion activity upon appropriate host stimuli without the deleterious effects of immunosuppressant drugs. Microcapsules can be classified in 3 categories: matrix‐core/shell microcapsules, liquid‐core/shell microcapsules, and cells‐core/shell microcapsules (or conformal coating). Many preparation techniques using natural or synthetic polymers as well as inorganic compounds have been reported. Matrix‐core/shell microcapsules in which cells are hydrogel‐embedded, exemplified by alginates capsule, is by far the most studied method. Numerous refinement of the technique have been proposed over the years such as better material characterization and purification, improvements in microbead generation methods, and new microbeads coating techniques. Other approaches, based on liquid‐core capsules showed improved protein production and increased cell survival. But aside those more traditional techniques, new techniques are emerging in response to shortcomings of existing methods. More recently, direct cell aggregate coating have been proposed to minimize membrane thickness and implants size. Microcapsule performances are largely dictated by the physicochemical properties of the materials and the preparation techniques employed. Despite numerous promising pre‐clinical results, at the present time each methods proposed need further improvements before reaching the clinical phase.

Drug-Loaded Nanocarriers: Passive Targeting and Crossing of Biological Barriers

Poor bioavailability and poor pharmacokinetic characteristics are some of the leading causes of drug development failure. Therefore, poorly-soluble drugs, fragile proteins or nucleic acid products may benefit from their encapsulation in nanosized vehicles, providing enhanced solubilization, protection against degradation, and increased access to pathological compartments. A key element for the success of drug-loaded nanocarriers is their ability to either cross biological barriers themselves, or allow loaded drugs to traverse them to achieve optimal pharmacological action at pathological sites. Depending on the mode of administration, nanocarriers may have to cross different physiological barriers in their journey towards their target. In this review, the crossing of biological barriers by passive targeting strategies will be presented for intravenous delivery (vascular endothelial lining, particularly for tumor vasculature and blood brain barrier targeting), oral administration (gastrointestinal lining), and upper airway administration (pulmonary epithelium). For each specific barrier, background information will be provided on the structure and biology of the tissues involved as well as available pathways for nano-objects or loaded drugs (diffusion and convection through fenestration, transcytosis, tight junction crossing, etc.). The determinants of passive targeting - size, shape, surface chemistry, surface patterning of nanovectors - will be discussed in light of current results. Perspectives on each mode of administration will be presented. The focus will be on polymeric nanoparticles and dendrimers, although advances in liposome technology will be also reported as they represent the largest body in the drug delivery literature.

Selectins Ligand Decorated Drug Carriers for Activated Endothelial Cell Targeting

New active particulate polymeric vectors based on branched polyester copolymers of hydroxy-acid and allyl glycidyl ether were developed to target drugs to the inflammatory endothelial cell surface. The hydroxyl and carboxyl derivatives of these polymers allow grafting of ligand molecules on the polyester backbones at different densities. A known potent nonselective selectin ligand was selected and synthesized using a new scheme. This synthesis allowed the grafting of the ligand to the polyester polymers, preserving its binding activity as assessed by docking simulations. Selectin expression on human umbilical cord vascular endothelial cells (HUVEC) was induced with the pro-inflammatory bacterial lipopolysaccharide (LPS) or with the nonselective inhibitor of nitric oxide synthase L-NAME. Strong adhesion of the ligand decorated nanoparticles was evidenced in vitro on activated HUVEC. Binding of nanoparticles bearing ligand molecules could be efficiently inhibited by prior incubation of cells with free ligand, demonstrating that adhesion of the nanoparticles is mediated by specific interaction between the ligand and the selectin receptors. These nanoparticles could be used for specific drug delivery to the activated vascular endothelium, suggesting their application in the treatment of diseases with an inflammatory component such as rheumatoid arthritis and cancer.

Functionalized nanospheres loaded with anti-angiogenic drugs: Cellular uptake and angiosuppressive efficacy

The objective of this study was to develop polymeric nanospheres (NPs) that are able to selectively target the activated vascular endothelium and to deliver co-encapsulated anti-angiogenic agents for improved treatment efficacy in inflammatory diseases with an angiogenic component. We evaluated a novel poly(d,l)-lactide (PLA)-based polymer, grafted with a synthetic ligand specific for selectin (PLA-g-SEL), for the preparation of functionalized NPs. The NPs were produced according to a double emulsion-solvent diffusion/evaporation method, allowing the co-encapsulation of hydrophilic and lipophilic drugs. Incorporation of the functionalized polymer enhanced the internalization of fluorescein-labeled NPs by lipopolysaccharide-activated vascular endothelial cells relative to control NPs, as evidenced by confocal laser scanning microscopy and quantitative fluorescence measurements. Two anti-angiogenic agents, endostatin and paclitaxel, were co-loaded in the functionalized NPs. Respective drug loadings were optimized by adjusting polymer composition, as well as by the microemulsion technique. NPs loaded with either of the chosen drugs or with a combination of them were tested for their anti-angiogenic efficacy in human umbilical vascular endothelial cell (HUVEC) culture in vitro and rat aorta tissue culture ex vivo models. An enhanced anti-proliferative effect on HUVECs and heightened anti-angiogenic action on rat aorta ring cultures was observed for the loaded drugs compared to the free molecules. Moreover, combined loaded treatments were found to be more potent, evoking additive and even synergetic outcomes (at lower doses) greater than the corresponding single-loaded treatments in inhibiting new vessels sprouting in rat aortic rings.

Direct Measurement of Mechanical and Adhesive Properties of Living Cells Using Surface Forces Apparatus

The adhesive and mechanical properties of living cells assembled into a monolayer on two different substrates were investigated using the surface forces apparatus (SFA) technique. The force measurements allowed elastic and bending moduli of the cells plated on substrates to be determined. The moduli are in good agreement with data reported in the literature for single cells determined using atomic force microscopy. Results confirm that the nature of the cell–substrate interactions can mediate cell mechanical and adhesive properties.

Unified Scaling of the Structure and Loading of Nanoparticles Formed by Diffusion-Limited Coalescence

The present study establishes the scaling laws describing the structure of spherical nanoparticles formed by diffusion-limited coalescence. We produced drug-loaded nanoparticles from a poly(ethylene glycol)-poly(d,l-lactic acid) diblock polymer (PEG- b-PLA) by the nanoprecipitation method using different types of micromixing chambers to explore multiple mixing regimes and characteristic times. We first show that the drug loading of the nanoparticles is not controlled by the mixing time but solely by the drug-to-polymer ratio (D:P) in the feed and the hydrophobicity of the drug scaled via the partition coefficient P. We then procure compelling evidence that particles formed via diffusion/coalescence exhibit a relative distribution of PEG blocks between the particle core and its shell that depends only on mixing conditions (not on D:P). Scaling laws of PEG relative distribution and chain surface density were derived in different mixing regimes and showed excellent agreement with experimental data. In particular, results made evident that PEG blocks entrapment in the core of the particles occurs in the slow-mixing regime and favors the overloading (above the thermodynamic limit) of the particles with hydrophilic drugs. The present analysis compiles effective guidelines for the scale up of nanoparticles structure and properties with mixing conditions, which should facilitate their future translation to medical and industrial settings.

Effect of surface chemistry of polymeric nanoparticles on cutaneous penetration of cholecalciferol

&NA; We investigated the influence of nanoparticle (NP) surface composition on different aspects of skin delivery of a lipophilic drug: chemical stability, release and skin penetration. Cholecalciferol was chosen as a labile model drug. Poly(lactic acid) (PLA)‐based NPs without surface coating, with a non‐ionic poly(ethylene glycol) (PEG) coating, or with a zwitterionic poly(2‐methacryloyloxyethyl phosphorylcholine) (PMPC) coating were prepared using flash nanoprecipitation. Process was optimized to obtain similar hydrodynamic diameters. Polymeric NPs were compared to non‐polymeric cholecalciferol formulations. Cholecalciferol stability in aqueous medium was improved by polymeric encapsulation with a valuable effect of a hydrophilic coating. However, the in vitro release of the drug was found independent of the presence of any polymer, as for the drug penetration in an intact skin model. Such tendency was not observed in impaired skin since, when stratum corneum was removed, we found that a neutral hydrophilic coating around NPs reduced drug penetration compared to pure drug NPs and bare PLA NPs. The nature of the hydrophilic block (PEG or PMPC) had however no impact. We hypothesized that NPs surface influenced drug penetration in impaired skin due to different electrostatic interactions between NPs and charged skin components of viable skin layers.