Bioengineered Bacteriophage-Like Nanoparticles as RNAi Therapeutics to Enhance Radiotherapy against Glioblastomas

Since glioblastomas (GBMs) are radioresistant malignancies and most GBM recurrences occur in radiotherapy, increasing the effectiveness of radiotherapy by gene-silencing has recently attracted attention. However, the difficulty in precisely tuning the composition and RNA loading in nanoparticles leads to batch-to-batch variations of the RNA therapeutics, thus significantly restricting their clinical translation. Here, we bioengineer bacteriophage Qβ particles with a designed broccoli light-up three-way junction (b-3WJ) RNA scaffold (contains two siRNA/miRNA sequences and one light-up aptamer) packaging for the silencing of genes in radioresistant GBM cells. The in vitro results demonstrate that the cleavage of de novo designed b-3WJ RNA by Dicer enzyme can be easily monitored in real-time using fluorescence microscopy, and the TrQβ@b-3WJLet-7_g^siEGFRsuccessfully knocks down EGFR and IKKα simultaneously and thereby inactivates NF-κB signaling to inhibit DNA repair. Delivery of TrQβ@b-3WJLet-7_g^siEGFRthrough convection-enhanced delivery (CED) infusion followed by 2Gy X-ray irradiation demonstrated that the median survival was prolonged to over 60 days compared with the 2Gy X-ray irradiated group (median survival: 31 days). Altogether, the results of this study could be critical for the design of RNAi-based genetic therapeutics, and CED infusion serves as a powerful delivery system for promoting radiotherapy against GBMs without evidence of systemic toxicity.