Comparison of Cell-Penetrating\nand Fusogenic TAT-HA2\nPeptide Performance in Peptideplex, Multicomponent, and Conjugate\nsiRNA Delivery Systems

In this study, the performance of the cell-penetrating\nand fusogenic\npeptide, TAT-HA2, which consists of a cell-permeable HIV trans-activator\nof transcription (TAT) protein transduction domain and a pH-responsive\ninfluenza A virus hemagglutinin protein (HA2) domain, was comparatively\nevaluated for the first time in peptideplex, multicomponent, and conjugate\nsiRNA delivery systems. TAT-HA2 in all three systems protected siRNA\nfrom degradation, except in the conjugate system with a low Peptide/siRNA\nratio. The synergistic effect of different peptide domains enhanced\nthe transfection efficiency of multicomponent and conjugate systems\ncompared to that of peptideplexes, which was attributed to the surface\nconfiguration of TAT-HA2 peptides depending on the nature of attachment.\nParticularly, the multicomponent system showed better cellular uptake\nand endosomal escape than the peptideplexes, resulting in enhanced\nsiRNA delivery in the cytoplasm. In addition, the presence of cleavable\ndisulfide bonds in multicomponent and conjugate systems promoted the\neffective siRNA delivery in the cytoplasm, resulting in improved gene\nsilencing activity. The multicomponent system reduced the level of\nluciferase expression in SKOV3 cells to 45% (±4). In contrast,\nthe conjugate system and the commercially available siRNA transfection\nagent, Lipofectamine RNAiMax, caused luciferase suppression down to\n55% (±2) at a siRNA dose of 100 nM. For the same dose, the peptideplex\nsystem could only reduce the luciferase expression to 65% (±5).\nNone of the developed systems showed significant toxicity at any dose.\nOverall, the TAT-HA2 peptide is promising as a siRNA delivery vector;\nhowever, its performance depends on the nature of attachment and,\nas a result, its surface configuration on the developed delivery system.