The six amino acid antimicrobial peptide bLFcin6 penetrates cells and delivers siRNA

Abstract

Cell-penetrating peptides (CPPs) are a new class of vectors with high pharmaceutical potential to deliver bioactive cargos into cells. Here, we characterized bLFcin6, a six amino acid peptide derived from bovine lactoferricin, as a CPP. Uptake of bLFcin6 was measured by flow cytometry. The ability to delivery siRNA was analyzed in HeLa cells. bLFcin 6 exhibited concentration-dependent uptake and intracellular distribution. Below 7.5 μm, uptake of bLFcin6 was significantly lower than uptake of TAT (P < 0.05) because bLFcin6 has fewer cationic amino acids. Compared to CPP5 (RLRWR) and CPP6 (PFVYLI), bLFcin6 had a significantly higher internalization ratio above 2.5 μm because it has two tryptophan residues. Uptake of bLFcin 6 starts with an ionic cell-surface interaction. It is then rapidly internalized by lipid raft-dependent macropinocytosis, followed by release from macropinosomes into the cytosol and nucleus. Moreover, bLFcin6 formed stable electrostatic complexes with siRNA and delivered siRNA into cells, resulting in significant knockout activity at both the mRNA and protein levels. The knockout activity of siRNA delivered by bLFcin6 was similar to that mediated by TAT, although knockout by bLFcin6 required a higher molar ratio. In this study, bLFcin6 was tested for its ability to act as an siRNA-delivering CPP. © 2012 The Authors Journal compilation © 2012 FEBS.