Cell-penetrating peptides (CPPs) currently represent one of the most promising tools for the cellular delivery of bioactive cargoes [1]. They generally correspond to short, purely cationic or amphipathic peptides. In the recent year, cyclic CPPs have attracted increasing attention because of their expected greater metabolic stability and internalization efficiency compared to the linear sequences [2]. We have evaluated in this study the impact of CPP cyclisation on their mechanism of entry.
A small library of cyclic CPPs with different ring sizes and functionalized with a fatty acid chain was prepared. Peptide head-to-tail cyclisation was performed by native chemical ligation [3]. For this, an N-terminal Cys residue and a C-terminal thioester (or a thioester precursor corresponding to an α-methylcysteine residue [4]) were introduced in the initial linear peptide sequences. CPPs were then conjugated to a bioactive peptide cargo by a disulfide bridge. The efficiency of cargo delivery inside cells was measured by a method based on MALDI-TOF MS [5] and the cargo intracellular distribution was analyzed by confocal microscopy. This study led to the identification of new structures that showed enhanced entry inside cells either by direct translocation or by glycosaminoglycan-dependent endocytosis.
[1] Fischer, R.; Fotin-Mleczek, M.; Hufnagel, H.; Brock, R. Chembiochem 2005, 6, 2126.
[2] Oh, D.; Nasrolahi Shirazi, A.; Northup, K.; Sullivan, B.; Tiwari, R. K.; Bisoffi, M.; Parang, K. Mol. Pharm. 2014, 11, 2845.
[3] Dawson, P.; Muir, T.; Clark-Lewis, I.; Kent, S. Science 1994, 266, 776.
[4] Burlina, F.; Papageorgiou, G.; Morris, C. ; White, P. D.; Offer, J. Chem. Sci. 2014, 5, 766.
[5] Burlina, F.; Sagan, S.; Bolbach, G.; Chassaing G. Angew Chem Int Ed 2005, 44, 4244.