Poly(N-methyl-N-vinylacetamide): A Strong Alternative to PEG for Lipid-Based Nanocarriers Delivering siRNA.

Lipid-based nanocarriers have demonstrated high interest in delivering genetic material, exemplified by the success of Onpattro and COVID-19 vaccines. While PEGylation imparts stealth properties, it hampers cellular uptake and endosomal escape, and may trigger adverse reactions like accelerated blood clearance (ABC) and hypersensitivity reactions (HSR). This work highlights the great potential of amphiphilic poly(N-methyl-N-vinylacetamide) (PNMVA) derivatives as alternatives to lipid-PEG for siRNA delivery. PNMVA compounds with different degrees of polymerization and hydrophobic segments, are synthesized. Among them, DSPE (1,2-distearoyl-sn-glycero-3-phosphoethanolamine)-PNMVA efficiently integrates into lipoplexes and LNP membranes and prevents protein corona formation around these lipid carriers, exhibiting stealth properties comparable to DSPE-PEG. However, unlike DSPE-PEG, DSPE-PNMVA24 shows no adverse impact on lipoplexes cell uptake and endosomal escape. In in vivo study with mice, DSPE-PNMVA24 lipoplexes demonstrate no liver accumulation, indicating good stealth properties, extended circulation time after a second dose, reduced immunological reaction, and no systemic pro-inflammatory response. Safety of DSPE-PNMVA24 is confirmed at the cellular level and in animal models of zebrafish and mice. Overall, DSPE-PNMVA is an advantageous substitute to DSPE-PEG for siRNA delivery, offering comparable stealth and toxicity properties while improving efficacy of the lipid-based carriers by minimizing the dilemma effect and reducing immunological reactions, meaning no ABC or HSR effects.

gH625 Cell-Penetrating Peptide Promotes the Endosomal Escape of Nanovectorized siRNA in a Triple-Negative Breast Cancer Cell Line.

The use of small interfering RNA (siRNA) to regulate oncogenes appears as a promising strategy in the context of cancer therapy, especially if they are vectorized by a smart delivery system. In this study, we investigated the cellular trafficking of a siRNA nanovector (called CS-MSN) functionalized with the cell-penetrating peptide gH625 in a triple-negative breast cancer model. With complementary techniques, we showed that siRNA nanovectors were internalized by both clathrin- and caveolae-mediated endocytosis. The presence of gH625 at the surface of the siRNA nanovector did not modify the entry pathway of CS-MSN, but it increased the amount of siRNA found inside the cells. Results suggested an escape of siRNA from endosomes, which is enhanced by the presence of the peptide gH625, whereas nanoparticles continued their trafficking into lysosomes. The efficiency of CS-MSN to inhibit the GFP in MDA-MB-231 cells was 1.7-fold higher than that of the nanovectors without gH625.

Formulation and in vitro evaluation of a siRNA delivery nanosystem decorated with gH625 peptide for triple negative breast cancer theranosis.

The development of an efficient small interfering RNA (siRNA) delivery system has held scientists interest since the discovery of the RNA interference mechanism (RNAi). This strategy gives hope for the treatment of many severe diseases. Herein, we developed hybrid nanovectors able to deliver siRNA to triple negative breast cancer cells. The nanovectors are based on PEGylated superparamagnetic iron oxide nanoparticles (SPION) functionalized with gH625 peptide, chitosan and poly-l-arginine. Every component has a key role and specific function: SPION is the core scaffolding the nanovector; PEG participates in the colloidal stability and the immune stealthiness; gH625 peptide promotes the nanovector internalization into cancer cells; cationic polymers provide the siRNA protection and favor siRNA endosomal escape and delivery to cytosol. The formulation was optimized by varying the amount of each compound. The efficacy of the siRNA retention and protection were investigated in the presence of high concentration of serum. Optimized nanovectors show a high uptake by MDA-MB-231 cells. The resulting down regulation of GFP expression was 73 ±â€¯3% with our nanovector compared to 59 ±â€¯8% obtained with the siRNA-Oligofectamine™ complex in the same conditions.

siRNA delivery system based on magnetic nanovectors: Characterization and stability evaluation.

Gene therapy and particularly small interfering RNA (siRNA) is a promising therapeutic method for treatment of various human diseases, especially cancer. However the lack of an ideal delivery system limits its clinical applications. Effective anticancer drug development represents the key for translation of research advances into medicines. Previously we reported, the optimization of magnetic siRNA nanovectors (MSN) formulation based on superparamagnetic iron oxide nanoparticles (SPION) and chitosan for systemic administration. This work aimed at using rational design to further optimize and develop MSN. Therefore, formulated MSN were first purified, then their physical and chemical properties were studied mainly through capillary electrophoresis. 95% of siRNA was found enclosed within the purified MSN (pMSN). pMSN showed colloidal stability at pH 7.4, effective protection of siRNA against ribonuclease degradation up to 24 hours and few siRNA release (less than 10%) at pH 7.4. These findings push toward further evaluation studies in vitro and/or in vivo, indicating the appropriateness of pMSN for cancer theranostics.