Synthesis and characterization of polymeric nanoparticles to target muscle cells

Duchenne muscular dystrophy (DMD) is a genetic disorder characterized by progressive degeneration of muscular tissue, caused by mutations in the dystrophin gene. It primarily affects males and leads to an average life expectancy of twenty years. Currently, there is no cure for DMD, but there are different treatments that attempt to improve quality of life and delay the disease’s progression. The research is focusing on other strategies involving gene therapy as a potential cure for the disease. While these approaches are giving encouraging results, in the clinical trials they are not as effective as expected: the inflammation and associated defects in the muscles complicate their regeneration. This is connected to the fact that stem cells are exhausted. Previous studies have demonstrated that miR-106b negatively regulates myogenic factor 5, impairing the production of muscle progenitors. It has been observed that inhibiting miR-106b can promote muscle regeneration. A possible solution could be vectorizing anti-miR-106b sponges specifically to muscle stem cells. To accomplish this, polymers developed by Grup d’Enginyeria de Materials (GEMAT) could be used to synthesize nanoparticles (NPs) functionalized with targeting peptides. This study focuses on polymeric nanoparticles (NPs) functionalized with the AAVMYO peptide, a targeting ligand known for its high efficiency and specificity for muscle tissue. AAVMYO interacts with α7β1 integrin, a laminin receptor overexpressed in DMD. We first corroborated its expression in C2C12 cell model using confocal immunofluorescence. Next, we focused on the uptake capacity of the NPs in different timing qualitatively and quantitatively. However, uptake levels for NPs with targeting peptides were not significantly higher than those without, suggesting limited targeting efficiency. To investigate further, we decided to conduct competition experiments between the peptide alone and our NPs and to verify, through confocal immunofluorescence, if the polymer with the peptides and the genetic material carried by the NPs colocalize. Our results indicate that the peptide’s targeting ability is not functioning as expected probably because of poor surface exposure of the peptide. A potential strategy to improve the results could be to synthesize a OM-pBAE with Dibenzocyclooctyne (DBCO), in which the peptide will be attached to the lateral chains thanks to the click chemistry. While the findings of this study are promising, future research should focus on optimizing peptide presentation to enhance targeting efficiency.