Abstract
Purpose. The aim of the study was to determine the critical determinants in nanoparticle-mediated gene transfection. It was hypothesized that different formulation parameters could affect the nanoparticle characteristics and hence its gene transfection.
Methods. Nanoparticles encapsulating plasmid DNA encoding for firefly luciferase were formulated using polylactide (PLA) and poly (d,l-lactide-co-glycolide) (PLGA) polymers of different compositions and molecular weights. A multiple-emulsion solvent-evaporation method with polyvinyl alcohol (PVA) as an emulsifier was used to formulate DNA-loaded nanoparticles. Gene expression of nanoparticles was determined in breast cancer (MCF-7) and prostate cancer (PC-3) cell lines.
Results. Nanoparticles formulated using PLGA polymer demonstrated greater gene transfection than those formulated using PLA polymer, and this was attributed to the higher DNA release from PLGA nanoparticles. Higher-molecular-weight PLGA resulted in the formation of nanoparticles with higher DNA loading, which demonstrated higher gene expression than those formulated with lower-molecular-weight PLGA. In addition, the nanoparticles with lower amount of surface-associated PVA demonstrated higher gene transfection in both the cell lines. Higher gene transfection with these nanoparticles was attributed to their higher intracellular uptake and cytoplasmic levels. Further study demonstrated that the molecular weight and the degree of hydrolyzation of PVA used as an emulsifier also affect the gene expression of nanoparticles.
Conclusions. Results thus demonstrate that the DNA loading in nanoparticles and its release, and the surface-associated PVA influencing the intracellular uptake and endolysosomal escape of nanoparticles, are some of the critical determinants in nanoparticle-mediated gene transfection.
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references
F. Liu and L. Huang. Development of non-viral vectors for systemic gene delivery. J. Control. Rel. 78:259-266 (2002).
M. D. Brown, A. G. Schatzlein, and I. F. Uchegbu. Gene delivery with synthetic (non viral) carriers. Int. J. Pharm. 229:1-21 (2001).
D. Lechardeur and G. L. Lukacs. Intracellular barriers to non-viral gene transfer. Curr. Gene Ther. 2:183-194 (2002).
H. Cohen, R. J. Levy, J. Gao, I. Fishbein, V. Kousaev, S. Sosnowski, S. Slomkowski, and G. Golomb. Sustained delivery and expression of DNA encapsulated in polymeric nanoparticles. Gene Ther. 7:1896-1905 (2000).
S. Prabha, W. Z. Zhou, J. Panyam, and V. Labhasetwar. Size-dependency of nanoparticle-mediated gene transfection: studies with fractionated nanoparticles. Int. J. Pharm. 244:105-115 (2002).
J. Panyam, W. Z. Zhou, S. Prabha, S. K. Sahoo, and V. Labhasetwar. Rapid endo-lysosomal escape of poly(D,L-lactide-co-glycolide) nanoparticles: implications for drug and gene delivery. FASEB J. 16:1217-1226 (2002).
K. Roy, H. Q. Mao, S. K. Huang, and K. W. Leong. Oral gene delivery with chitosan-DNA nanoparticles generates immunologic protection in a murine model of peanut allergy. Nature Med. 5:387-391 (1999).
V. Labhasetwar, J. Bonadio, S. A. Goldstein, and R. J. Levy. Gene transfection using biodegradable nanospheres: results in tissue culture and a rat osteotomy model. Colloid Surface B: Biointerfaces 16:281-290 (1999).
K. M. Shakesheff, C. Evora, I. I. Soriano, and R. Langer. the adsorption of poly(vinyl alcohol) to biodegradable microparticles: studied by x-ray photoelectron spectroscopy (XPS). J. Colloid Interface Sci. 185:538-547 (1997).
M. F. Zambaux, F. Bonneaux, R. Gref, P. Maincent, E. Dellacherie, M. J. Alonso, P. Labrude, and C. Vigneron. Influence of experimental parameters on the characteristics of poly(lactic acid) nanoparticles prepared by a double emulsion method. J Control. Rel. 50:31-40 (1998).
S. K. Sahoo J. Panyam S. Prabha, and V. Labhasetwar. Residual polyvinyl alcohol associated with poly (D,L-lactide-co-glycolide) nanoparticles affects their physical properties and cellular uptake. J. Control. Rel. 82:105-114 (2002).
J. Davda and V. Labhasetwar. Characterization of nanoparticle uptake by endothelial cells. Int. J. Pharm. 233:51-59 (2002).
H. Murakami, M. Kobayashi, H. Takeuchi, and Y. Kawashima. Preparation of poly(D,L-lactide-co-glycolide) nanoparticles by modified spontaneous emulsification solvent diffusion method. Int. J. Pharm. 187:143-152 (1999).
S. Liand and L. Huang. Nonviral gene therapy: promises and challenges. Gene Ther. 7:31-34 (2000).
M. Nishikawa and L. Huang. Nonviral vectors in the new millennium: delivery barriers in gene transfer. Hum. Gene Ther. 12:861-870 (2001).
V. P. Torchilin, T. S. Levchenko, R. Rammohan, N. Volodina, B. Papahadjopoulos-Sternberg, and G. G. D'Souza. Cell transfection in vitro and in vivo with nontoxic TAT peptide-liposome-DNA complexes. Proc. Natl. Acad. Sci. USA 100:1972-1977 (2003).
M. Thomas and A. M. Klibanov. Enhancing polyethylenimine's delivery of plasmid DNA into mammalian cells. Proc. Natl. Acad. Sci. USA 99:14640-14645 (2002).
M. L. Hedley, J. Curley, and R. Urban. Microspheres containing plasmid-encoded antigens elicit cytotoxic T-cell responses. Nat. Med. 4:365-368 (1998).
J. Panyam and V. Labhasetwar. Biodegradable nanoparticles for drug and gene delivery to cells and tissue. Adv. Drug Deliv. Rev. 55:329-347 (2003).
R. Wattiaux, N. Laurent, S. Wattiaux-De Coninck, and M. Jadot. Endosomes, lysosomes: their implication in gene transfer. Adv. Drug Deliv. Rev. 41:201-208 (2000).
R. Jeyanthi, R. C. Mehta, B. C. Thanoo, and P. P. DeLuca. Effect of processing parameters on the properties of peptide-containing PLGA microspheres. J. Microencapsul. 14:163-174 (1997).
N. B. Viswanathan, P. A. Thomas, J. K. Pandit, M. G. Kulkarni, and R. A. Mashelkar. Preparation of non-porous microspheres with high entrapment efficiency of proteins by a (water-in-oil)-in-oil emulsion technique. J. Control. Rel. 58:9-20 (1999).
Y. Y. Hsu, T. Haoand, and M. L. Hedley. Comparison of process parameters for microencapsulation of plasmid DNA in poly(D,L-lactic-co-glycolic) acid microspheres. J. Drug Target. 7:313-323 (1999).
R. Langer, W. D. Rhine, D. T. Hsiehand, and R. S. Bawa. Polymers for sustained release of macromolecules: Applications and control of release kinetics. In R. Baker (ed), Controlled Release Bioactive Materials, Academic Press, Bend, Oregon, 1980, pp. 83-98.
Q. R. Chenand and A. J. Mixson. Systemic gene therapy with p53 inhibits breast cancer: recent advances and therapeutic implications. Front. Biosci. 3:D997-D1004 (1998).
H. Murakami, Y. Kawashima, T. Niwa, T. Hino, H. Takeuchiand, and M. Kobayashi. Influence of degree of hydrolyzation and polymerization of poly(vinylalcohol) on the preparation and properties of poly(D,L-lactide-co-glycolide) nanoparticle. Int. J. Pharm. 149:43-49 (1997).
F. Shi, L. Wasungu, A. Nomden, M. C. Stuart, E. Polushkin, J. B. Engbertsand, and D. Hoekstra. Interference of poly(ethylene glycol)-lipid analogues with cationic-lipid-mediated delivery of oligonucleotides; role of lipid exchangeability and non-lamellar transitions. Biochem. J. 366:333-341 (2002).
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Prabha, S., Labhasetwar, V. Critical Determinants in PLGA/PLA Nanoparticle-Mediated Gene Expression. Pharm Res 21, 354–364 (2004). https://doi.org/10.1023/B:PHAM.0000016250.56402.99
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DOI: https://doi.org/10.1023/B:PHAM.0000016250.56402.99