Figure 6.

Multiparametric screen of ternary siRNA polyplexes reveals lead si-NP formulation DB4-PDB12. (A) si-NPs which are optimally tuned to overcome multiple siRNA delivery barriers such as size range, cell uptake, and endosomal escape achieve the highest target gene silencing in vitro (heat map parameters and thresholds are shown in Table S3; overlaid black line indicates level of residual luciferase activity for each si-NP formulation loaded with anti-luciferase siRNA). (B) Cell uptake trended inversely to si-NP surface PEG thickness (see PEG thickness calculations in Figure S8). (C) High cell uptake did not directly predict target gene silencing in vitro. (D) Incorporation of the hydrophobic and endosomolytic DB-core was the strongest predictor of gene silencing in vitro (p < 0.001). Lead si-NP formulations (DB4-PDB12 and DB4-PDB16; indicated in red) achieve > 80% gene silencing in (E) fibroblast (3T3s), (F) mesenchymal stem cells (MSCs), and (G) triple negative breast cancer (MDA-MB-231) cells. (H) The IC₅₀ for DB4-PDB12 in MDA-MB-231 cells is 15.4 nM. (Yellow: DB-PD, Red: DB-PDB, Orange: D-PDB)