Harnessing Argonaute 2 as an innovative delivery system for siRNA in glioblastoma treatment

Abstract

RNA interference via small interfering RNA (siRNA) offers promising potential for treating glioblastoma (GB), the most common and aggressive primary brain tumor. While local siRNA delivery could bypass the blood-brain barrier and achieve high drug concentrations, challenges remain in developing effective delivery systems that ensure siRNA stability, efficient cellular uptake, endolysosomal escape, and long-term brain availability. The Argonaute-2 (Ago2) protein, a core component of the RNA interference machinery, naturally binds siRNA and protects it from degradation. Ago2 is also found in biological fluids, such as blood, and in extracellular vesicles, including exosomes and microvesicles, making it a potential natural vector for siRNA delivery.

This study explores Ago2 as a natural vector for siRNA in nanoparticle-based delivery systems to enhance siRNA transport and gene silencing in GB. Neutral polymeric siRNA/Ago2-loaded nanoparticles (NPs), around 300 nm in size, were formulated with high encapsulation efficiencies (85.1 % for protein and 57.5 % for siRNA). In vitro testing in U87MG GB cells stably expressing luciferase (U87MG Bml1) showed that, in contrast to free siRNA/Ago2 complexes, siRNA/Ago2-loaded NPs effectively reduced luciferase expression compared to scramble siRNA formulations, with no significant cytotoxicity at 48 h. Interestingly, the introduction of exogenous Ago2 into cells unexpectedly upregulated luciferase expression, suggesting that Ago2 may also influence global gene expression beyond its role in siRNA-mediated silencing.

These findings highlight the potential of siRNA/Ago2-loaded NPs for local siRNA delivery in GB and suggest that Ago2 could play a broader regulatory role in gene expression, in addition to enhancing siRNA activity. Further investigation is needed to explore Ago2’s impact on gene regulation and its potential as a therapeutic tool in GB treatment, particularly in combination with biocompatible hydrogels for sustained delivery.

Materials

Ester-capped PLGA (PLGA-COOR) (Mn = 5.5 kDa, PDI 2.0) and PEG-PLGA copolymer (MnPEG = 5 kDa, MnPLGA = 25.7 kDa, PDI 2.1) were synthesized using a ring-opening polymerization method as previously described [37]. U87MG (Bmi1-3′UTR) luciferase-expressing human glioblastoma cell line (HTB-14TM) was obtained from the American Type Culture Collection (Manassas, United States) and modified at the Vector Production Center, Inserm U1089, Atlantic Gene Therapies facility in Nantes (FRANCE).

A. Rinaldi, E. Kluczka, M. Louis Fathy Nazir, M. Barbotin, C. Roy, L. Basset, S. Avril, S. Anthiya, G. Tosi, C. Jérôme, F. Boury, A. Rousseau, E. Garcion, F. Dumas, Harnessing Argonaute 2 as an innovative delivery system for siRNA in glioblastoma treatment, Journal of Drug Delivery Science and Technology, Volume 114, Part A, 2025, 107459, ISSN 1773-2247, https://doi.org/10.1016/j.jddst.2025.107459.