Nanoparticulate systems for the delivery of antisense oligonucleotides

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Abstract

Antisense oligonucleotides are molecules that are able to inhibit gene expression being therefore potentially active for the treatment of viral infections or cancer. However, because of their poor stability in biological medium and their weak intracellular penetration, colloidal drugs carriers such as nanoparticles were developed for the delivery of oligonucleotides (ODN). ODN associated to nanoparticles were shown to be protected against degradation and to penetrate more easily into different types of cells. As a consequence, nanoparticles were shown to improve the efficiency of ODNs for the inhibition of the proliferation of cells expressing the point mutated Ha-ras gene. In vivo, polyalkylcyanoacrylate (PACA) nanoparticles were able to efficiently distribute the ODNs to the liver whereas the alginate nanosponges could concentrate the ODNs in the lungs. Finally, ODN loaded to PACA nanoparticles were able to improve in mice, the treatment of RAS cells expressing the point mutated Ha-ras gene.

Section snippets

Introduction: antisense strategy and the need for particulate formulations

The aim of antisense strategy is to interfere with gene expression by preventing the translation of proteins from mRNA. Theoretically, an antisense oligonucleotide is a short fragment (from 15 to 20 sequence bases) of deoxynucleotides that have a sequence complementary to a portion of the targeted mRNA. The antisense oligonucleotides then hybridize with the mRNA by Watson–Crick base pairing and blocks sterically the translation of this transcript into a protein [1] (Fig. 1). This mechanism is

Nanospheres

Several methods have been developed for preparing nanospheres. They can be classified in two main categories according to whether the formation of nanospheres requires a polymerization reaction, or whether it is achieved directly from a macromolecule or a preformed polymer (Fig. 4).

In vitro stability of ODNs adsorbed onto nanospheres

When adsorbed onto PIHCA nanospheres through binding to CTAB, ODNs were efficiently protected against enzymatic degradation even after 5 h incubation with phosphodiesterase or in cell culture media [25]. In addition, about 90% of the oligonucleotide still remained intact after overnight incubation with the enzyme (0.1 mg/ml). Similar results were obtained by Zobel et al. [57] with ODNs adsorbed onto DEAE containing nanospheres incubated with DNAase.

ODNs-DEAE-Nanospheres complexes were found to

Cell interactions with ODNs loaded nanospheres

Cell uptake studies of a 15mer oligothymidilate adsorbed onto PIHCA nanospheres was performed in non toxic conditions using U937 cells. It was shown that the uptake of the ODNs was dramatically increased when associated with nanospheres. After 24 h incubation, uptake of oligonucleotide was 8-times higher when adsorbed to nanospheres than when incubated as an ODN free solution [26], and it was markedly reduced (95%) at 4°C as compared to 37°C. These results clearly show that ODNs adsorbed onto

In vitro pharmacological activity of oligonucleotides-loaded nanospheres

The in vitro activity of oligonucleotide loaded PIHCA nanospheres was demonstrated in a few studies. Schwab et al. [66], [67] have shown that nanosphere-adsorbed antisense ODNs directed to a point mutation (G→U) in codon 12 of the Ha-ras mRNA selectively inhibited the proliferation of cells expressing the point mutated Ha-ras gene. With nanospheres, the efficient concentration was 100-times lower than with ODN free. A sequence specific inhibition of the profileration of T24 human bladder

In vivo studies with oligonucleotide nanospheres

The pharmacokinetic studies carried out with PIBCA nanospheres [65] have shown that although nanospheres did not markedly increase the blood half-life of a 33P-16 mer oligothymidylate, its tissue distribution was significantly modified. Indeed, when transported by PIBCA nanospheres, oligothymidylate was importantly taken up by the liver whereas a subsequent reduced distribution in the other organs was observed, especially in the kidneys. These data suggest that with the aid of nanospheres, the

Conclusion

The results presented in this review show that the association of ODNs with biodegradable nanospheres is possible if using a hydrophobic counter-ion or in the case of modified ODNs to become more hydrophobic or finally when oligonucleotides are entrapped in an hydrophilic system such as alginate nanosponges. These systems have been proved to be efficient not only for protecting the oligonucleotide from the degradation by 3′-exonucleases but also for increasing the intracellular capture of these

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