Chitosan-based nanoparticles for in vivo delivery of interfering agents including siRNA

https://doi.org/10.1016/j.cocis.2013.06.005Get rights and content

Highlights

  • siRNA were successfully associated with 4 types of chitosan-based nanoparticles.

  • Association of interfering agents with chitosan-based nanoparticles is mediated by chitosan.

  • In vivo delivery of biologically active siRNA was successful with chitosan-based nanoparticles.

  • Activity of siRNA delivered in vivo is enhanced with targeted chitosan-based nanoparticles.

Abstract

The potential of siRNA to knock down expression of genes has been identified as an exciting strategy for specific treatments of disease-associated genes. However, their clinical development is pended to the achievement of their effective intracellular delivery in the target cells in vivo. So far, this was a bottleneck for fast development of siRNA in clinics because of their high enzymatic susceptibility in biological media and their poor intracellular uptake. The realization of therapeutic potential of the RNA interference approach strongly depended on the rational design of safe and effective carriers. This review considers carriers made of chitosan-based nanoparticles. It reports the methods of synthesis and the interactions of siRNA with chitosan which is at the basis of the association, stability and delivery to cells of siRNA with these carriers. Results of evaluations of the interference activity produced in vitro and in vivo by the interfering molecule delivered with chitosan-based nanoparticle carriers are discussed. As pointed out from different examples, the remarkable efficacy of the chitosan-based nanoparticles to deliver active interfering agents in vivo and to achieve a successful systemic delivery including by oral administration are very encouraging. Although we are still in the early stage of developments, it can be expected that results reported so far paved the road to stimulate further developments and strengthen their clinical application perspectives.

Introduction

siRNA interference remains a recent discovery. While the RNA interference phenomenon was first described 23 years ago, the elucidation of the mechanism occurred at the dawn of the 21st century [1]. The power of small interfering RNA (siRNA) to knock down expression of genes was immediately identified as an exciting new potential approach for the development of very specific treatments of disease-associated genes. This covers a wide range of pathologies. Therapeutic strategies based on an interference effect have the potential to control viral infections, cancer development and degenerative diseases [2••], [3]. The development of clinically relevant siRNA to achieve a reliable and specific gene silencing activity has progressed extremely rapidly. Several efficient siRNA have reached advanced stages of clinical developments when local treatments can be applied [2••]. siRNA are RNA molecules that are active by themselves directly after being transfected into cells. Their clinical development is pending to the achievement of their effective intracellular delivery in target cells in in vivo conditions. Their chemical nature is a bottleneck for the developments of treatments especially those based on a systemic delivery. The high enzymatic degradation susceptibility of siRNA in biological media and their polyanionic nature which contribute to their poor cellular uptake constitute major technical hurdle that prevent so far their appearance in the arsenal of molecules for a wide range of antiviral and antitumoral therapies. The realization of the therapeutic potential of RNA interference strongly depends on the rational design of safe and efficient delivery systems making possible the delivery of the siRNA to target cells in vivo after a systemic administration. Several potential delivery systems consist of viral particles but it is generally admitted that effective non-viral delivery system for siRNA will be preferred for obvious safety reasons [2••], [4••], [5•]. Several types of synthetic non-viral materials have been found suitable to achieve effective delivery of siRNA into cells in vitro [5•], [6•], [7], [8]. Unfortunately, they cannot be further developed to achieve in vivo delivery of siRNA due to their toxicological profile despite many efforts made to improve their safety profiles [7], [8]. Nevertheless, a few of them including lipofectamine and polyethyleneimine were developed as transfection agents for in vitro use. With time, they became references for the evaluation of the transfection given by new compounds in in vitro experiments. Today, chitosan, which is a polysaccharide, is the component which seems to be unanimously adopted for the development of nanoparticle delivery systems for nucleic acids. It is used in more than 80% of nanoparticles designed for genes and interfering components [9••], [10], [11]. These systems are assumed to present a good compromise between safety and transfection efficiency. They have extensively been used for gene delivery [12], [13]. Based on this experience they were tested as delivery systems for siRNA. The intense activity justifies that there are already several reviews on the subject especially covering the delivery of DNA by chitosan-based delivery systems [13]. A few review papers report researches carried on the application of an interference strategy thanks to the delivery of siRNA with chitosan-based delivery systems [14], [15], [16]. The aim of the present review was to describe the nanoparticles designed with chitosan that were suggested as delivery systems for siRNA and to give an overview on the results obtained so far on the in vitro and in vivo evaluations of the interference effects. For this, the review was balanced between the physico-chemical aspects of chitosan-based nanoparticles as colloids loaded with siRNA and those of the use of biological models to evaluate the potential of the approach delivering the siRNA with different delivery systems. The methods for the preparation of the nanoparticles, the characteristics and the performance in terms of siRNA loading are presented in the first part of this review. As the association of the siRNA with chitosan-based nanoparticles mainly results from interactions with chitosan, a second part of this review discusses the different parameters that govern this interaction including on the thermodynamic point of view. The last parts of the review summarize interference activities reported using the nanoparticles as it was evaluated considering different levels in the delivery challenge. This will review in vitro data in which the potentials of the delivery systems are evaluated in conditions allowing direct contact with cells. For the in vivo evaluations, the results were presented considering the different routes of administrations that were tested in the view to produce a local or a systemic effect.

Section snippets

Obtaining chitosan-based nanoparticles for siRNA delivery

Different types of nanoparticles were designed with chitosan to achieve in vivo delivery of interfering agents including siRNA. Methods considering the association or entrapment of siRNA in the nanoparticles are exploiting the polyanionic character of these molecules. The negative charges come from the phosphate group included in each nucleotide composing the two strands of the interfering agent. This polyanionic character is exploited to form polyelectrolyte complexes with polycations.

Characteristics of the interactions between chitosan and siRNA

Interactions of siRNA with chitosan govern many properties of the drug delivery system [14], [15]. First they control the association of the siRNA with the chitosan-based nanoparticles. Second, they control the stability of the complex which in turn controls the protection of siRNA against enzymatic degradation since the siRNAs remain protected when it forms a complex with the nanoparticles. Finally, these interactions are also important to control the release of the siRNA.

Despite the fact that

In vitro evaluation of siRNA delivery to cells

A few studies have reported the use of chitosan-based delivery systems to explore their potential to achieve transfection of siRNA into cells considering in vitro experiments. In these experiments, the carrier loaded with the siRNA is incubated with cells expressing the target mRNA which means that there is almost no barrier between the delivery system and the cell membrane. Then, it is expected that these conditions are the most favorable to observe the interference activity of a given siRNA

In vivo evaluation of siRNA delivery with chitosan-based nanoparticles

Several examples of in vivo delivery of siRNA using chitosan-based delivery systems can be found in the literature. Interestingly, these systems were tested to deliver interfering agents by different routes of administration including both local and systemic routes. Here we present the results available in the literature considering the mode of administration. This defines the level of the delivery challenge that needs to be achieved to obtain a therapeutic activity of the siRNA.

Conclusion

This review focuses on chitosan-based nanoparticle carriers for in vivo delivery of interfering molecules including siRNA. As pointed out, a few types of chitosan-based nanoparticles that differ from their structures were developed as carriers for the delivery of interfering agents. In these systems, chitosan or its derivatives play central roles controlling the association of siRNA and the stability of the nanoparticles, the release of the siRNA and the stability of the siRNA against enzymatic

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