Protamine-Based Nanotherapeutics for Gene Delivery to Glioblastoma Cells

Isocitrate dehydrogenase wild-type glioblastoma is the most aggressive primary brain tumor classified as grade 4 of malignancy. Standard treatment, combining surgical resection, radiotherapy, and chemotherapy, often leads to severe side effects, with the emergence of tumor recurrence in all cases. Nucleic acid-based therapy has emerged as a promising strategy for cancer treatment. Non-viral nanosystems have become the vehicles of choice for gene delivery, due to their efficient nucleic acid encapsulation, protection, and intracellular transport. This work explores the potential of a formulation of low molecular weight protamine (LMWP) and dextran sulfate for gene delivery. The nanoparticles (NPs) were evaluated in terms of particle size, surface charge, morphology, and capacity to condense different nucleic acids. NPs formed by ionic complexation resulted in a homogeneous population of spherical particles with a low polydispersity index (PDI), small size, and positive surface charge. Competitive displacement assay demonstrated that the NPs could condense nucleic acids without alterations in their morphology and physicochemical characteristics, even after long-term storage. The efficacy of this formulation as a gene delivery system was evaluated <i>in vitro</i> in different glioblastoma cell lines and three-dimensional (3D) spheroids and <i>in vivo</i> using zebrafish models, showing negligible toxicity, efficient internalization, and consistent expression of fluorescent/luminescent proteins. Overall, these cationic polymeric NPs show promising features for their use as non-viral gene delivery vehicles for glioblastoma treatments.