Fig. 6: Systemic delivery of SANPs-siPraja2 inhibits GBM growth.

a Schematic view of the experimental procedures. U87MG cells were stereotaxically implanted into the brain of nude mice (time 0). One week post-implantation, SANPs-siRNAs were i.v. injected into the caudal vein every 48 h, for a total of 14 days of treatment. At 3 weeks post-implantation, mice were sacrificed and tumor lesions isolated and further characterized. b Brain distribution of rhodamine-labeled SANPs-siPraja2 by fluorescence analysis at 9 h after i.v. injection. GBM lesions were identified by immunostaining the same brain sections with an anti-human vimentin antibody. Nuclei were stained with DAPI. Representative images are shown. Scale bar, 50 μm. c Tissue sections from tumor lesions were stained with hematoxylin/eosin. d Quantitative analysis of the tumor volume is expressed as a mean value ± SEM. Three independent experiments were performed. P value: ** = 0.0014. e Tumor sections were stained with hematoxylin/eosin or immunostained for Ki-67. Scale bar, 50 μm. f Quantitative analysis of Ki-67-positive cells in tumor lesions from control and SANPs-siPraja2 treated mice. The data were expressed as a mean value ± SEM from three independent experiments. P value: ** = 0.0055. g U87MG-Luc cells were injected into the brain of 6 weeks old CD1 mice. Three hours following implantation, bioluminescent intensity (BLI) was measured by intraperitoneal injection of 150 mg/kg d-Luciferin potassium salt. At 1-week post-injection, based on BLI measurement, mice were randomized into two experimental groups of 12 animals, and each group was treated by tail vein injection with SANPs-siPraja2 (GP1) and SANPs-siRNAc (GP2), respectively. Treatments were repeated twice a week for 4 weeks, then four mice for each group were sacrificed and organs collected. BLI analysis was performed every week and quantitative data were collected. A representative set of animals for each experimental group is shown. h Quantitative and cumulative analysis of BLI scores. *** <0.001. i Kaplan–Meier curve of treated animals. At 52 days from U87MG implantation, all the animals from SANPs-siRNAc group died. In contrast, about 40% of SANPs-siPraja2 mice were still healthy, but the experiment was terminated in accordance with Authorities guidelines. j Immunostaining analysis for praja2, KSR2, pThr172-AMPKα, and AMPKα1 in tumor sections from control and SANPs-siPraja2 treated mice. Scale bar, 50 μm.