Efficient cell delivery mediated by lipid‐specific endosomal escape of supercharged branched peptides

• Published in: Traffic (Copenhagen, Denmark) Vol. 19; no. 6; pp. 421 - 435
• Main Authors: Brock, Dakota J., Kustigian, Lauren, Jiang, Mengqiu, Graham, Kristin, Wang, Ting‐Yi, Erazo‐Oliveras, Alfredo, Najjar, Kristina, Zhang, Junjie, Rye, Hays, Pellois, Jean‐Philippe
• Format: Journal Article
• Language: English
• Published: Former Munksgaard John Wiley & Sons A/S 01.06.2018; Wiley Subscription Services, Inc
• Subjects: bis(monoacylglycero)phosphate; Bone morphogenetic proteins; Cell Line, Tumor; cellular delivery; cell‐penetrating peptides; Cytosol - metabolism; endosomal escape; Endosomes - metabolism; Flocculation; HeLa Cells; HIV; Human immunodeficiency virus; Humans; Intracellular Membranes - metabolism; Leakage; Lipid bilayers; Lipid Bilayers - metabolism; Lipids; Lipids - chemistry; Liposomes - metabolism; Macromolecules; membrane leakage; Peptides - metabolism; supercharged molecules; Tat protein; cellular delivery; bis(monoacylglycero)phosphate; membrane leakage; endosomal escape; supercharged molecules; cell-penetrating peptides
• ISSN: 1398-9219; 1600-0854; 1600-0854
• DOI: 10.1111/tra.12566

Various densely charged polycationic species, whether of biological or synthetic origin, can penetrate human cells, albeit with variable efficiencies. The molecular underpinnings involved in such transport remain unclear. Herein, we assemble 1, 2 or 3 copies of the HIV peptide TAT on a synthetic scaffold to generate branched cell‐permeable prototypes with increasing charge density. We establish that increasing TAT copies dramatically increases the cell penetration efficiency of the peptides while simultaneously enabling the efficient cytosolic delivery of macromolecular cargos. Cellular entry involves the leaky fusion of late endosomal membranes enriched with the anionic lipid BMP. Derivatives with multiple TAT branches induce the leakage of BMP‐containing lipid bilayers, liposomal flocculation, fusion and an increase in lamellarity. In contrast, while the monomeric counterpart 1TAT binds to the same extent and causes liposomal flocculation, 1TAT does not cause leakage, induce fusion or a significant increase in lamellarity. Overall, these results indicate that an increase in charge density of these branched structures leads to the emergence of lipid specific membrane‐disrupting and cell‐penetrating activities. Delivery agents presenting multiple branches of the TAT peptide unlock endosomal escape—a necessary step in the successful cytosolic penetration of cells. Above a threshold charge density, these supercharged species promote contact and restructuring of endosomal membranes enriched with the anionic lipid bis(monoacylglycerol)phosphate. In particular, peptide‐bound membranes undergo leaky fusion, an increase in lamellarity and flocculation. This multifaceted interplay revealed between polycationic peptides and anionic endosomal membranes provides a mechanistic basis for the development of efficient cellular delivery agents.