Figure 9.

A large amount of functional siRNA is released and loaded into RISC following treatment with a GalNAc-conjugated endolytic peptide. An ESC GalNAc–siRNA (siTTR-1) targeting Ttr was dosed SC at 0.5 mg/kg. Half of the mice in the study were treated with 5 mg/kg of the GalNAc-INF7 endolytic peptide dosed SC 15 min after the siRNA dose. Cohorts of mice were sacrificed at several times post-dose (4 h through day 10). Ttr mRNA levels were quantified and normalized to Gapdh in control animals (A) or those that were treated with GalNAc-INF7 peptide (B). (C) Total siRNA sense strand liver levels were quantified by RT-qPCR in control (dark blue) or endolytic peptide-treated animals (light blue). (D) Total siRNA antisense strand liver levels were quantified by RT-qPCR in control (dark red) or endolytic peptide-treated animals (light red). (E) Sense strand RISC loading was quantified by RT-qPCR in control (dark blue) or endolytic peptide-treated animals (light blue). (F) Antisense strand RISC loading was quantified by RT-qPCR in control (dark red) or endolytic peptide-treated animals (light red). Additional control pull downs were performed with a mutant APP peptide (Supplementary Figure S3). Data is represented as mean ± SD, for N = 3. Welch's independent t-test was utilized to compare the mean value between the control and peptide-treated groups at each timepoint to identify significant differences in siRNA liver levels and RISC loading (*P ≤ 0.05, **P ≤ 0.01, ***P ≤ 0.001). Full results for the statistical analysis are included in the Supplemental Information.