A multifunctional nano device as non-viral vector for gene delivery: in vitro characteristics and transfection.
The aim of this work was to design a new multifunctional nano device (MND) for gene delivery. This MND was equipped with folic acid as ligand, which was conjugated to terminal amido of poly(aminopoly(ethylene glycol)cyanoacrylate-co-hexadecyl cyanoacrylate) (poly(H(2)NPEGCA-co-HDCA)) to synthesize poly(Folate-HNPEGCA-co-HDCA), protamine sulfate (PS) as DNA condenser and for nuclear transfer, PEG chain from poly(Folate-HNPEGCA-co-HDCA) for decreasing macrophages recognition and extending half-life, dioleoyl phosphatidylethanolamine (DOPE) for endosomal escape, and we supposed that the latent DOPE fusogenicity could be gently restored along with fast degradation of poly(Folate-HNPEGCA-co-HDCA) in MND membrane within endosome. Our experimental results showed that optimum complexation ( approximately 97%) of DNA was achieved at DNA:PS=1:3 (w/w). The MND showed different loading ratio by lipid film hydration technique with the highest loading ratio about 12%, the particle size range 200-400 nm, surface charge range 8 mV-15 mV. MND1 (poly(Folate-HNPEGCA-co-HDCA)/DOPE, 5:95, molar ratio) exhibited a high burst release effect with 60% of pDNA/PS released within 1 day at PBS (pH 4.5), but with 21.4% and 8.1% pDNA/PS release at PBS with pH 5.8 and 7.4 within 24 h, respectively. However, lesser pDNA/PS release occurred in MND2 (poly(Folate-HNPEGCA-co-HDCA)/DOPE, 10:90, molar ratio) with 46%, 16.9% and 7.8% of pDNA/PS released at PBS with pH 4.5, 5.8 and 7.4 within 24 h, respectively. After 1 day, pDNA/PS displayed a sustained release pattern. The amount of cumulated pDNA/PS release over 3 days was 75% and 51.2% at PBS with pH 4.5 for MND1 and MND2, respectively. The MND loading pDNA/PS showed that luciferase activity was over 0.5 ng luciferase/mg protein in KB cells, in particular, the MND1 showed the highest transfection efficiency (0.66 ng luciferase/mg protein) in KB cells, which was much higher compared with in A549 cells or other formulations such as LipofectAMINE, free pDNA/PS and control multifunctional nano device (CMND), whose lipid film was consisted of poly(H(2)NPEGCA-co-HDCA) and DOPE. In addition, MND also showed good protection during encapsulation and low cytotoxicity. As a result, MND could be a more potential non-viral vector for delivery of DNA.
Gao
Yu
Y
Institute of Materia Medica, Shanghai Institute for Biological Sciences, Chinese Academy of Sciences, Shanghai, China.
Gu
Wangwen
W
Chen
Lingli
L
Xu
Zhenghong
Z
Li
Yaping
Y
D045744:Cell Line, Tumor; D002470:Cell Survival / Q000235:genetics; D018014:Gene Transfer Techniques; D005822:Genetic Vectors / Q000008:administration & dosage* / Q000235:genetics* / Q000502:physiology; D006801:Humans; D053758:Nanoparticles / Q000008:administration & dosage*; D014162:Transfection / Q000379:methods*