Serum stability and physicochemical characterization of a novel amphipathic peptide C6M1 for siRNA delivery.
The efficient delivery of nucleic acids as therapeutic agents is a major challenge in gene therapy. Peptides have recently emerged as a novel carrier for delivery of drugs and genes. C6M1 is a designed amphipathic peptide with the ability to form stable complexes with short interfering RNA (siRNA). The peptide showed a combination of random coil and helical structure in water but mainly adopted a helical conformation in the presence of anions or siRNA. Revealed by dynamic light scattering (DLS) and microscopy techniques, the interaction of C6M1 and siRNA in water and HEPES led to complexes of ∼70 and ∼155 nm in size, respectively, but showed aggregates as large as ∼500 nm in PBS. The time-dependent aggregation of the complex in PBS was studied by DLS and fluorescence spectroscopy. At molar ratio of 15∶1, C6M1 was able to completely encapsulate siRNA; however, higher molar ratios were required to obtain stable complexes. Naked siRNA was completely degraded in 4 h in the solution of 50% serum; however C6M1 protected siRNA against serum RNase over the period of 24 h. Western blotting experiment showed ∼72% decrease in GAPDH protein level of the cells treated with C6M1-siRNA complexes while no significant knockdown was observed for the cells treated with naked siRNA.
Department of Chemical Engineering, University of Waterloo, Waterloo, Ontario, Canada; Waterloo Institute for Nanotechnology, University of Waterloo, Waterloo, Ontario, Canada.
Mousa
M
Jafari
Department of Chemical Engineering, University of Waterloo, Waterloo, Ontario, Canada; Waterloo Institute for Nanotechnology, University of Waterloo, Waterloo, Ontario, Canada.
Wen
W
Xu
Department of Chemical Engineering, University of Waterloo, Waterloo, Ontario, Canada; Waterloo Institute for Nanotechnology, University of Waterloo, Waterloo, Ontario, Canada.
Ran
R
Pan
Department of Chemical Engineering, University of Waterloo, Waterloo, Ontario, Canada.
Chad M
CM
Sweeting
Department of Chemical Engineering, University of Waterloo, Waterloo, Ontario, Canada; Department of Chemistry, University of Peradeniya, Peradeniya, Sri Lanka.
Desiree Nedra
DN
Karunaratne
Department of Chemical Engineering, University of Waterloo, Waterloo, Ontario, Canada; Waterloo Institute for Nanotechnology, University of Waterloo, Waterloo, Ontario, Canada.
Pu
P
Chen
D000818:Animals; D016466:CHO Cells; D057846:Cell-Penetrating Peptides / Q000737:chemistry*; D006224:Cricetinae; D003412:Cricetulus; D055785:Gene Knockdown Techniques; D005987:Glyceraldehyde-3-Phosphate Dehydrogenases / Q000378:metabolism; D055550:Protein Stability; D017433:Protein Structure, Secondary; D020871:RNA Stability; D034741:RNA, Small Interfering / Q000737:chemistry*; D012260:Ribonucleases / Q000737:chemistry; D044967:Serum / Q000737:chemistry / Q000201:enzymology; D014162:Transfection
C586100:C6M1 peptide; D057846:Cell-Penetrating Peptides; D034741:RNA, Small Interfering; D005987:Glyceraldehyde-3-Phosphate Dehydrogenases; D012260:Ribonucleases