- Research Article
Targeted cationic poly(D,L-lactic-co-glycolic acid) nanoparticles for gene delivery to cultured cells
Cellular & Molecular Biology Letters volume 14, Article number: 347 (2009)
We developed a new targeted cationic nanoparticulate system composed of poly(D,L-lactic-co-glycolic acid) (PLGA), 1,2-dioleoyl-3-(trimethylammonium) propane (DOTAP) and asialofetuin (AF), and found it to be a highly effective formulation for gene delivery to liver tumor cells. The nanoparticles (NP) were prepared by a modified solvent evaporation process that used two protocols in order to encapsulate (NP1 particles) or adsorb (NP2 particles) plasmid DNA. The final particles are in the nanoscale range. pDNA loaded in PLGA/DOTAP/AF particles with high loading efficiency showed a positive surface charge. Targeted asialofetuin-nanoparticles (AF-NP) carrying genes encoding for luciferase and interleukin-12 (IL-12) resulted in increased transfection efficiencies compared to free DNA and to plain (non-targeted) systems, even in the presence of 60% fetal bovine serum (FBS). The results of transfections performed on HeLa cells, defective in asialoglycoprotein receptors (ASGPr-), confirmed the receptor-mediated endocytosis mechanism. In summary, this is the first time that asialoglycoprotein receptor targeting by PLGA/DOTAP/DNA nanoparticles carrying the therapeutic gene IL-12 has been shown to be efficient in gene delivery to liver cancer cells in the presence of a very high concentration of serum, and this could be a potential system for in vivo application.
Dulbecco’s modified Eagle’s medium-high glucose
fetal bovine serum
scanning electron microscopy
Pouton, C.W. and Seymour, L.W. Key issues in non-viral gene delivery. Adv. Drug Deliv. Rev. 46 (2001) 187–203.
De Smedt, S.C., Demeester, J. and Hennink, W.E. Cationic polymer based gene delivery systems. Pharm. Res. 17 (2000) 113–126.
Singh, M., Briones, M., Ott, G. and O’Hagan, D. Cationic microparticles: A potent delivery system for DNA vaccines. Proc. Natl. Acad. Sci. USA. 97 (2000) 811–816.
Kim, I.S., Lee, S.K., Park, Y.M., Lee, Y.B., Shin, S.C., Lee, K.C. and Oh, I.J. Physicochemical characterization of poly(L-lactic acid) and poly(D,L-lactide-co-glycolide) nanoparticles with polyethylenimine as gene delivery carrier. Int. J. Pharm. 298 (2005) 255–262.
Oster, C.G., Kim, N., Grode, L., Barbu-Tudoran, L., Schaper, A.K., Kaufmann, S.H. and Kissel, T. Cationic microparticles consisting of poly(lactide-co-glycolide) and polyethylenimine as carriers systems for parental DNA vaccination. J. Control. Release 104 (2005) 359–377.
Moghimi, S.M., Hunter, A.C. and Murray, J.C. Long-circulating and target-specific nanoparticles: theory to practice. Pharmacol. Rev. 53 (2001) 283–318.
Vinogradov, S.V., Bronich, T.K. and Kabanov, A.V. Nanosized cationic hydrogels for drug delivery: preparation, properties and interactions with cells. Adv. Drug Deliv. Rev. 54 (2002) 135–147.
Leong, K.W., Mao, H.Q., Truong-Le, V.L., Roy, K., Walsh, S.M. and August, J.T. DNA-polycation nanospheres as non-viral gene delivery vehicles. J. Control. Release 53 (1998) 183–193.
Hirosue, S., Muller, B.G., Mulligan, R.C. and Langer, R. Plasmid DNA encapsulation and release from solvent diffusion nanospheres. J. Control. Release 70 (2001) 231–242.
Ravi Kumar, M. N., Bakowsky, U. and Lehr, C.M. Preparation and characterization of cationic PLGA nanospheres as DNA carriers. Biomaterials 25 (2004) 1771–1777.
Rhaese, S., von Briesen, H., Rubsamen-Waigmann, H., Kreuter, J. and Langer, K. Human serum albumin-polyethylenimine nanoparticles for gene delivery. J. Control. Release 92 (2003) 199–208.
Wang, D., Robinson, D.R., Kwon, G.S. and Samuel, J. Encapsulation of plasmid DNA in biodegradable poly(D, L-lactic-co-glycolic acid) microspheres as a novel approach for immunogene delivery. J. Control. Release 57 (1999) 9–18.
Hedley, M.L., Curley, J. and Urban, R. Microspheres containing plasmid-encoded antigens elicit cytotoxic T-cell responses. Nat. Med. 4 (1998) 365–368.
Diez, S. and Tros de Ilarduya, C. Versatility of biodegradable poly (D,L-lactic-co-glycolic acid) microspheres for plasmid DNA delivery. Eur. J. Pharm. Biopharm. 63 (2006) 188–197.
Arangoa, M.A., Düzgüneş, N. and Tros de ILarduya, C. Increased receptor-mediated gene delivery to the liver by protamine-enhanced-asialofetuin-lipoplexes. Gene Ther. 10 (2003) 5–14.
Wilson, J.M., Grossman, M., Cabrera, J.A., Wu, C.H. and Wu, G.Y. A novel mechanism for achieving transgene persistence in vivo after somatic gene transfer into hepatocytes. J. Biol. Chem. 267 (1992) 11483–11489.
Wu, G.Y. and Wu, C.H. Evidence for targeted gene delivery to HepG2 hepatoma cells in vitro. Biochemistry 27 (1988) 887–892.
Martinez-Fong, D., Mullersman, J.E., Purchio, A.F., Armendáriz-Borunda, J. and Martínez-Hernández, A. Nonenzymatic glycosylation of poly-L-lysine: a new tool for targeted gene delivery. Hepatology 20 (1994) 1602–1608.
Zanta, M.A., Boussif, O., Adib, A. and Behr, J.P. In vitro gene delivery to hepatocytes with galactosylated polyethylenimine. Bioconjug. Chem. 8 (1997) 839–844.
Ashwell, G. and Morell, A.G. The role of surface carbohydrates in the hepatic recognition and transport of circulating glycoproteins. Adv. Enzymol. Relat. Areas Mol. Biol. 41 (1974) 99–128.
Wu, J., Liu, P., Zhu, J.L., Maddukuri, S. and Zern, M.A. Increased liver uptake of liposomes and improved targeting efficacy by labeling with asialofetuin in rodents. Hepatology 27 (1998) 772–778.
Shive, M.S. and Anderson, J.M. Biodegradation and biocompatibility of PLA and PLGA microspheres. Adv. Drug Deliv. Rev. 28 (1997) 5–24.
Labhasetwar, V., Bonadio, J., Goldstein, S., Chen, W. and Levy, R.J. A DNA controlled-release coating for gene transfer: transfection in skeletal and cardiac muscle. J. Pharm. Sci. 87 (1998) 1347–1350.
Luo, D., Woodrow-Mumford, K., Belcheva, N. and Saltzman, W.M. Controlled DNA delivery systems. Pharm. Res. 16 (1999) 1300–1308.
Ando, S., Putnam, D., Pack, D.W. and Langer, R. PLGA microspheres containing plasmid DNA: preservation of supercoiled DNA via cryopreparation and carbohydrate stabilization. J. Pharm. Sci. 88 (1999) 126–130.
Hsu, Y.Y., Hao, T. and Hedley, M.L. Comparison of process parameters for microencapsulation of plasmid DNA in poly(D,L-lactic-co-glycolic) acid microspheres. J. Drug Target. 7 (1999) 313–323.
Panyam, J. and Labhasetwar, V. Biodegradable nanoparticles for drug and gene delivery to cells and tissue. Adv. Drug Deliv. Rev. 55 (2003) 329–347.
Prabha, S. and Labhasetwar, V. Critical determinants in PLGA/PLA nanoparticle-mediated gene expression. Pharm. Res. 21 (2004) 354–364.
Cohen, H., Levy, R.J., Gao, J., Fishbein, I., Kousaev, V., Sosnowski, S., Slomkowski, S. and Golomb, G. Sustained delivery and expression of DNA encapsulated in polymeric nanoparticles. Gene Ther. 7 (2000) 1896–1905.
Jeong, J.H. and Park, T.G. Poly(L-lysine)-g-poly(D,L-lactic-co-glycolic acid) micelles for low cytotoxic biodegradable gene delivery carriers. J. Control. Release 82 (2002) 159–166.
Walter, E. and Merkle, H.P. Microparticle-mediated transfection of nonphagocytic cells in vitro. J. Drug Target. 10 (2002) 11–21.
Chiou, H.C., Tangco, M.V., Levine, S.M., Robertson, D., Kormis, K., Wu, C.H., and Wu, G.Y. Enhanced resistance to nuclease degradation of nucleic acids complexed to asialoglycoprotein-polylysine carriers. Nucleic Acids Res. 22 (1994) 5439–5446.
Katayose, S. and Kataoka, K. Remarkable increase in nuclease resistance of plasmid DNA through supramolecular assembly with poly(ethylene glycol)-poly(L-lysine) block copolymer. J. Pharm. Sci. 87 (1998) 160–163.
Zhang, Y. and Anchordoquy, T.J. The role of lipid charge density in the serum stability of cationic lipid/DNA complexes. Biochim. Biophys. Acta 1663 (2004) 143–157.
Simberg, D., Weisman, S., Talmon, Y., Faerman, A., Shoshani, T. and Barenholz, Y. The role of organ vascularization and lipoplex-serum initial contact in intravenous murine lipofection. J. Biol. Chem. 27 (2003) 39858–39865.
Eliyahu, H., Servel, N., Domb, A.J. and Barenholz, Y. Lipoplex-induced hemagglutination: potential involvement in intravenous gene delivery. Gene Ther. 9 (2002) 850–858.
Karmali, P.P., Majeti, B.K., Sreedhar, B. and Chaudhuri, A. In vitro gene transfer efficacies and serum compatibility profiles of novel mono-, di-, and tri-histidinylated cationic transfection lipids: a structure-activity investigation. Bioconjug. Chem. 17 (2006) 159–171.
Tros de Ilarduya, C. and Düzgüneş, N. Efficient gene transfer by transferrin lipoplexes in the presence of serum. Biochim. Biophys. Acta. 1463 (2000) 333–342.
Fischer, D., Bieber, T., Li, Y., Elsasser, H.P. and Kissel, T. A novel nonviral vector for DNA delivery based on low molecular weight, branched polyethylenimine: effect of molecular weight on transfection efficiency and cytotoxicity. Pharm. Res. 16 (1999) 1273–1279.
Fischer, D., Li, Y., Ahlemeyer, B., Krieglstein, J. and Kissel, T. In vitro cytotoxicity testing of polycations: influence of polymer structure on cell viability and hemolysis. Biomaterials 24 (2003) 1121–1131.
Simoes, S., Slepushkin, V., Pires, P., Gaspar, R., de Lima, M.P. and Düzgüneş, N. Mechanisms of gene transfer mediated by lipoplexes associated with targeting ligands or pH-sensitive peptides. Gene Ther. 6 (1999) 1798–1807.
About this article
Cite this article
Díez, S., Miguéliz, I. & Tros de Ilarduya, C. Targeted cationic poly(D,L-lactic-co-glycolic acid) nanoparticles for gene delivery to cultured cells. Cell Mol Biol Lett 14, 347 (2009). https://doi.org/10.2478/s11658-009-0003-7
- Poly(D,L-lactic-co-glycolic acid) (PLGA)
- 1,2-dioleoyl-3-(trimethylammonium) propane (DOTAP)
- Targeted gene delivery
- Pharmaceutical nanotechnology