Smart PEG-Block-PLA/PLA Nanosystems: Impact of the Characteristics of the Polymer Blend on the Redox Responsiveness.

Nanocarriers (NCs) were designed from three polymer blends (B1, B2 and B3) and investigated as smart drug delivery systems (SDDS). The blends are composed of a "smart" copolymer, where methoxy poly(ethylene glycol) and poly(lactic acid) are connected via a redox-responsive disulfide bond (mPEG-SS-PLA), and of a "conventional" polymer, poly(lactic acid) (PLA). They differ by mPEG-SS-PLA/PLA ratio and PLA molecular weight. Nanoprecipitation was used to prepare NCs. Three concentrations were tested, and fluorescent dye Nile red (NR) was used as a model payload. The results show that the characteristics of the NCs, such as size and drug release kinetics, are influenced by the type of blend and the concentration used during the nanoprecipitation process. The more redox-responsive blend was B2 (ratio 1:3, PLA 5 kDa) at 16 mg/mL: the quantity of NR released was tripled upon 24 h of incubation in a reducing medium. This study reveals that the amount of disulfide bonds present in a NC is not the only parameter to be considered to design an SDDS. The stability of the SDDS in a presumably non-stimulating environment is also important to limit uncontrolled release during storage or in the body before the biological target is reached.

Three-Step Synthesis of a Redox-Responsive Blend of PEG-block-PLA and PLA and Application to the Nanoencapsulation of Retinol.

Smart polymeric nanocarriers have been developed to deliver therapeutic agents directly to the intended site of action, with superior efficacy. Herein, a mixture of poly(lactide) (PLA) and redox-responsive poly(ethylene glycol)-block-poly(lactide) (PEG-block-PLA) containing a disulfide bond was synthesized in three steps. The nanoprecipitation method was used to prepare an aqueous suspension of polymeric nanocarriers with a hydrodynamic diameter close to 100 nm. Retinol, an anti-aging agent very common in cosmetics, was loaded into these smart nanocarriers as a model to measure their capacity to encapsulate and to protect a lipophilic active molecule. Retinol was encapsulated with a high efficiency with final loading close to 10% w/w. The stimuli-responsive behavior of these nanocarriers was demonstrated in vitro, in the presence of l-Glutathione, susceptible to break of disulfide bond. The toxicity was low on human keratinocytes in vitro and was mainly related to the active molecule. Those results show that it is not necessary to use 100% of smart copolymer in a nanosystem to obtain a triggered release of their content.

New Formulation for the Delivery of Oligonucleotides Using "Clickable" siRNA-Polyisoprenoid-Conjugated Nanoparticles: Application to Cancers Harboring Fusion Oncogenes.

The aim of the present study is to take advantage of the unique property of polyisoprenoid chains to adopt a compact molecular conformation and to use these natural and biocompatible lipids as nanocarriers of drugs to deliver siRNA. A new chemical strategy is applied here to conjugate squalene (SQ) and solanesol (SOLA) to siRNA consisting of an activated variant of the azide-alkyne Huisgen cycloaddition also known as copper-free (Cu-free) click chemistry. We conjugated siRNA against TMPRSS2-ERG, a fusion oncogene found in more than 50% of prostate cancers to SQ or SOLA. First, several parameters such as molar ratio, solvents, temperature, incubation time, and the annealing schedule between both siRNA strands were investigated to bioconjugate the SQ or SOLA via Cu-free click chemistry. The best parameters of the new bioconjugation approach allowed us to (i) increase the synthesis yield up to 95%, (ii) avoid the formation of byproducts during the synthesis, and (iii) improve the reproducibility of the bioconjugation. Then, the biological activity of the resulting nanoparticles was assessed. In vitro, all four formulations were able to decrease the corresponding oncogene and oncoprotein expression. In vivo, only two of the four nanoformulations showed anti-neoplastic activity that seems to be tightly related to their dissimilar biodistribution behavior. In conclusion, we performed a new approach easily transposable for pharmaceutical development to synthesize siRNA-SQ and siRNA-SOLA and to obtain efficient siRNA-nanoparticles. The robustness of the process could be extended to several other polyterpenes and likely applied to other siRNA targeting genes whose overexpression results in the development of cancers or other genetic diseases.

Circulating Lipoproteins: A Trojan Horse Guiding Squalenoylated Drugs to LDL-Accumulating Cancer Cells.

Selective delivery of anticancer drugs to rapidly growing cancer cells can be achieved by taking advantage of their high receptor-mediated uptake of low-density lipoproteins (LDLs). Indeed, we have recently discovered that nanoparticles made of the squalene derivative of the anticancer agent gemcitabine (SQGem) strongly interacted with the LDLs in the human blood. In the present study, we showed both in vitro and in vivo that such interaction led to the preferential accumulation of SQGem in cancer cells (MDA-MB-231) with high LDL receptor expression. As a result, an improved pharmacological activity has been observed in MDA-MB-231 tumor-bearing mice, an experimental model with a low sensitivity to gemcitabine. Accordingly, we proved that the use of squalene moieties not only induced the gemcitabine insertion into lipoproteins, but that it could also be exploited to indirectly target cancer cells in vivo.

Synthesis of a deuterated probe for the confocal Raman microscopy imaging of squalenoyl nanomedicines.

The synthesis of ω-di-(trideuteromethyl)-trisnorsqualenic acid has been achieved from natural squalene. The synthesis features the use of a Shapiro reaction of acetone-d 6 trisylhydrazone as a key step to implement the terminal isopropylidene-d 6 moiety. The obtained squalenic acid-d 6 has been coupled to gemcitabine to provide the deuterated analogue of squalenoyl gemcitabine, a powerful anticancer agent endowed with self-assembling properties. The Raman spectra of both deuterated and non-deuterated squalenoyl gemcitabine nanoparticles displayed significant Raman scattering signals. They revealed no differences except from the deuterium peak patterns in the silent spectral region of cells. This paves the way for label-free intracellular trafficking studies of squalenoyl nanomedicines.

In vitro investigation of multidrug nanoparticles for combined therapy with gemcitabine and a tyrosine kinase inhibitor: Together is not better.

Combined therapy with gemcitabine and tyrosine-kinase inhibitors (i.e., sunitinib) has already demonstrated important benefits in pancreatic cancer treatment. Further therapeutic advantage could be achieved by their co-loading in a single nanoscale system, which enables (i) the co-existence of drugs with different mechanisms of action and pharmacokinetic profiles and (ii) the fine tuning of their release rate overcoming the rapid clearance often observed with free drugs. In this context, the already validated squalenoylation approach has been applied to the design of a multidrug nanoparticle (NP) made by co-self-assembly of the squalene-based prodrugs of gemcitabine (SQGem) and sunitinib (SQSun). We hypothesized that co-delivering of SQGem and SQSun in a single nanoparticle was capable to increase their cytotoxicity on MIA PaCa-2 pancreatic cancer cells compared to the monodrug NPs. Nevertheless, multidrug NPs (i.e., SQGem/SQSun NPs) were as efficient as the physical mixture of the individual monodrug NPs (SQGem NPs + SQSun NPs) thus suggesting that the cytotoxicity raised from the exposure of the cells simultaneously to the two bioconjugates rather than to their original loading into a single or two different nanoparticles. To be noted that the lack of differences in static 2D cultures does not exclude a different behavior in dynamic conditions in vivo.