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Blood-neural barriers regulate nutrient supply to neuronal tissues and prevent neurotoxicity. In particular, the inner blood-retinal barrier (iBRB) and blood–brain barrier (BBB) share common origins in development, and similar morphology and function in adult tissue, while barrier breakdown and leakage of neurotoxic molecules can be accompanied by neurodegeneration. Therefore, pre-clinical research requires human in vitro models that elucidate pathophysiological mechanisms and support drug discovery, to add to animal in vivo modeling that poorly predict patient responses. Advanced cellular models such as microphysiological systems (MPS) recapitulate tissue organization and function in many organ-specific contexts, providing physiological relevance, potential for customization to different population groups, and scalability for drug screening purposes. While human-based MPS have been developed for tissues such as lung, gut, brain and tumors, few comprehensive models exist for ocular tissues and iBRB modeling. Recent BBB in vitro models using human cells of the neurovascular unit (NVU) showed physiological morphology and permeability values, and reproduced brain neurological disorder phenotypes that could be applicable to modeling the iBRB. Here, we describe similarities between iBRB and BBB properties, compare existing neurovascular barrier models, propose leverage of MPS-based strategies to develop new iBRB models, and explore potentials to personalize cellular inputs and improve pre-clinical testing.

Nanomedicine has emerged as a major field of academic research with direct impact on human health. While a first generation of products has been successfully commercialized and has significantly contributed to enhance patient's life, recent advances in material design and the emergence of new therapeutics are contributing to the development of more sophisticated systems. As the field matures, it is important to comprehend the challenges related to nanoparticle commercial development for a more efficient and predictable path to the clinic. Areas covered: The review provides an overview of nanoparticle-based delivery systems currently on the market and in clinical trials, and discuss the principal challenges for their commercial development, both from a manufacturing and regulatory perspective, to help gain understanding of the translational path for these systems. Expert Opinion: Clinical translation of nanoparticle-based delivery systems remains challenging on account of their 3D nanostructure and requires robust nano-manufacturing process along with adequate analytical tools and methodologies. By identifying early enough in the development the product critical attributes and understanding their impact on the therapeutic performance, the developers of nanopharmaceuticals will be better equipped to develop efficient product pipelines. Second-generation products are expected to broaden nanopharmaceutical global market in the upcoming years.

PEGylated chitosan-based nanoparticles offer attractive platforms for siRNA cocktail delivery into tumors. Still, therapeutic efficacy requires us to select a rational combination of siRNAs and an efficient tumor delivery after systemic administration. Here, we showed that non-covalent PEGylation of chitosan-based nanoparticles loaded with siRNA targeting two key transporters of energy fuels for cancer cells, namely the lactate transporter MCT1 and the glutamine transporter ASCT2, could lead to significant antitumor effects. As a ligand, we tested variations of the prototypical RGD peptidomimetic (RGDp). A higher siRNA delivery was obtained with naphthyridine-containing RGDp randomly conjugated on the PEG chain by clip photochemistry and the use of a lipophilic linker than when using traditional chain-end grafting and RGDp with a hydrophilic linker. The antiproliferative effects resulting from ASCT2 and MCT1 silencing were validated separately in vitro in conditions mimicking specific metabolic profiles of cancer cells and in vivo upon concomitant delivery. The combination of those siRNA and the selected components of targeted RGDp nanoparticles led to a dramatic tumor growth inhibition upon peri-tumoral but also systemic administration in mice. Altogether these data emphasize the convenience of using non-covalent PEGylated chitosan particles to produce sheddable stealth protection compatible with an efficient siRNA delivery in tumors.

RNA interference, the process in which small interfering RNAs (SiRNAs) silence a specific gene and thus inhibit the associated protein, has opened new doors for the treatment of a wide range of diseases. However, efficient delivery of SiRNAs remains a challenge, especially due to their instability in biological environments and their inability to cross cell membranes. To protect and deliver SiRNAs to mammalian cells, a variety of polymeric nanocarriers have been developed. Among them, the polysaccharide chitosan has generated great interests. This derivative of natural chitin is biodegradable and biocompatible, and can complex SiRNAs into nanoparticles on account of its positive charges. However, chitosan presents some limitations that need to be taken into account when designing chitosan/SiRNA nanoparticles. Here, we describe a method to prepare SiRNA/chitosan nanoparticles with high gene silencing efficiency and low cytotoxicity by using the ionic gelation technique.

This study aims at developing chitosan-based nanoparticles suitable for an intravenous administration of small interfering RNA (siRNA) able to achieve (i) high gene silencingwithout cytotoxicity and (ii) stability in biological media including blood. Therefore, the influence of chitosan/tripolyphosphate ratio, chitosan physicochemical properties, PEGylation of chitosan as well as the addition of an endosomal disrupting agent and a negatively charged polymer was assessed. The gene silencing activity and cytotoxicity were evaluated on B16 melanoma cells expressing luciferase. We monitored the integrity and the size behavior of siRNA nanoparticles in human plasma using fluorescence fluctuation spectroscopy and single particle tracking respectively. The presence of PEGylated chitosan and poly(ethylene imine) was essential for high levels of gene silencing in vitro. Chitosan nanoparticles immediately released siRNA in plasma while the inclusion of hyaluronic acid and high amount of poly(ethylene glycol) in the formulation improved the stability of the particles. The developed formulations of PEGylated chitosan-based nanoparticles that achieve high gene silencing in vitro, low cytotoxicity and high stability in plasma could be promising for intravenous delivery of siRNA.

INTRODUCTION: Nucleic acids have witnessed a dramatic acceleration in their therapeutic exploitation and currently represent a growing number of applications in drug development pipelines. However, a more wide-spread development of therapeutics based on nucleic acids is restricted by their poor chemical and enzymatic stability in vivo, lack of cellular uptake and insufficient capability to reach intracellular targets.AREAS COVERED: Advanced formulation of nucleic acids in nano-sized carriers may help unlocking their potential as therapeutic agents. Nano-sized matters own specific features responsible for inducing characteristic interactions with biological molecules and tissues. These properties enable for the enhancement of the nano-formulation's therapeutic efficacy, but on the other hand, the nanomatters interactions in biological fluids are also responsible for adverse effects. The purpose of this review is to reflect on the complexity in the therapeutic delivery of RNA interference-based drugs emerging from the recent clinical experiences and report the actual technological and analytical advances introduced to solve it.EXPERT OPINION: The complexity in the therapeutic delivery of nucleic acids and the heterogeneity of side effects make the interpretation of the therapeutic outcome difficult. Hence the development of analytical approaches applicable in the field of nucleic acid delivery is becoming a major challenge.

Recently, chitosan has attracted significant attention in the formulation of small interfering RNA (siRNA). Because of its cationic nature, chitosan can easily complex siRNA, thus readily forming nanoparticles. Moreover, chitosan is biocompatible and biodegradable, which make it a good candidate for siRNA delivery in vivo. However, chitosan requires further development to achieve high efficiency. This review will describe the major barriers that impair the efficiency of the chitosan-based siRNA delivery systems, including the stability of the delivery system in biological fluids and endosomal escape. Several solutions to counteract these barriers have been developed and will be discussed. The parameters to consider for designing powerful delivery systems will be described, particularly the possibilities for grafting targeting ligands. Finally, optimized systems that allow in vivo therapeutic applications for both local and systemic delivery will be reviewed. This review will present recent improvements in chitosan-based siRNA delivery systems that overcome many of these system's previous pitfalls and pave the way to a new generation of siRNA delivery systems.

The mechanism of RNA silencing has first been evidenced in plants. Then, studies on the nematode Caenorhabditis elegans identified a similar phenomenon, called RNA interference (RNAi). RNAi was described as a naturally occurring mechanism that induced sequence-specific post-transcriptional gene silencing and that was mediated by long double stranded RNAs. Few years later, the administration of exogenous small interfering RNA (siRNA) of 21-22 nucleotides in mammalian cells successfully induced the silencing of endogenous and heterologous genes. This discovery has created new perspectives for the development of gene medicines, especially for the treatment of diseases that are characterized by an overexpression of specific proteins. The RNAi technology promised the rapid development of highly specific therapeutics that could theoretically silence any gene, thus overcoming the issue of undruggable targets. However, after the initial enthusiasm, it clearly appeared that the delivery of small RNAs molecules required precisely engineered nanosystems. Although recent progresses have been made in the understanding of the siRNA delivery modalities, as well as in the design of siRNA delivery systems, the translational development of RNAi-based drugs remains challenging and researchers are still looking for powerful delivery approaches. This work investigated the biopolymer chitosan as the basis material for the elaboration of a siRNA delivery system for tumor targeting. Chitosan exhibits attractive properties, such as its biocompatibility and biodegradability, its cationic nature and its chemical versatility that allows the grafting of targeting ligand. These characteristics make chitosan an ideal basis for nanoparticles (NPs) construction. The research was conducted in three steps. First, the formulation parameters for the design of optimal siRNA NPs using chitosan were established. The principle requirements for ideal NPs were suitable physico-chemical characteristics, espec, (BIFA - Sciences biomédicales et pharmaceutiques) -- UCL, 2013