Formulation of nanoparticles from short chain chitosan as gene delivery system and transfection against T47D cell line
Abstract
Recently numerous prototype DNA-based biopharmaceuticals can be used to control disease progression by induction and inhibitin the overexpression of genes. Since there are poor cellular uptake and rapid in vivo degradation of DNA-based therapeutics therefore the use of delivery systems to facilitate cellular internalization and preserve their activity is necessary. Cationic polymers commonly used as carriers to delivery gene because of easy to form complexes and higher stability compared to that lipoplexs. Chitosan, a cationic, are polymer most widely used in gene delivery systems because of the low toxicity, and biocompatible. The aim of this study was to formulate nanoparticles of short chain chitosan-pEGFP-C1 and short chain chitosan/TPP-pEGFP-C1 by coaservation complex method. Stability test of the formula was performed by incubating the nanoparticles complex with DNase I and Artificial Intestinal Fluid. Cytotoxicity and transfection studies were evaluated against T47D cell line. The diameter of Chitosan-pEGFP-C1 and chitosan/TPP-pEGFP-C1 nanoparticles were on the range of 56–282.8 nm. The zeta potential wasdetermined to be +14.03 - +16.6 mV. Stability studies showed that chitosan-pEGFP-C1 and chitosan/TPPpEGFP-C1 nanoparticles were stable, undegradable by DNase I and artificial intestinal fluid. Cytotoxic Assay of Chitosan-pEGFP-C1 and chitosan/TPP-pEGFPC1 nanoparticles (pH 4.0) showed that the viability of cell was > 90% for all formulas. EGFP-C1 plasmid gene delivered by chitosan nanoparticles can be expressed in T47D cell culture. According to these results chitosan and chitosan/TPP nanoparticles had potentially to be used as a non-viral vector system delivery for gene therapy.
Key words:Chitosan, Nanoparticles, Plasmid EGFP-C1, Cell culture T47D
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Counis, M., F. and Torriglia, A., 2000, DNases and Apoptosis, Biochem. Biophys. Res. Commun.
Biochim. Biol. Cell., 78:405-414
Hirano, S., Seino, H., Akiyama, Y., and Nonaka, I.,1990, dalam Geebelein, C., G., Dunn, R., L., Progress in Biomedical Polymers, Plenum Press, New York, 283
Indrawati, M.I.M., 2010, Formulasi Nanopartikel Menggunakan Kitosan Rantai Pendek dan Transfeksinya pada Sel Kanker SP-C1, Tesis, Fakultas Farmasi Universitas Gadjah Mada, Yogyakarta.
Kommareddy, S. and Amiji, M., 2005, Preparation and Evaluation of Thiol-modified Gelatin Nanoparticles for Intracellular DNA Delivery in Response to Glutathione, Bioconjug.Chem., 16:1423-1432
Lai, Y., Drobinskaya, I., Kolossov, E., Chen, C. and Linn, T., 2008, Genetic Modification of Cells for Transplantation, Adv. Drug Deliv Rev, 60(2):146–59.
LeHoux, J.G. and Grondin, F., 1993, Some Effects of Chitosan on Liver Function in The Rat, Endocrinology, 132: 1078-1084
Martien, R., and Loretz, B., 2006, Oral Gene Delivery: Design of Polymeric Carrier systems shielding toward Intestinal Enzymatic Attack, Biopolymers 83(4):327-36
Martien, R., and Loretz, B., 2007, Chitosan Thioglycolic Acid Conjugate: An Alternative Carrier for Oral Nonviral Gene Delivery?, J. Biomed. Mater. Res. A,82(1):1-9
Martien, R., Sandbichler L.B. and Schnurch, A., B., 2008, Thiolated Chitosan Nanoparticles: Transfection Study in the Caco-2 Differentiated Cell Culture, Nanotechnol., 19:045101
Mosmann, T., 1983, Rapid Colorimetric Assay for Cellular Growth and Survival Application to Proliferation and Cytotoxicity Assays, J. Immunol. Methods, 65:55–63.
Mutmainah, N., 2010, Formulasi Nanopartikel Menggunakan Kitosan Rantai Sedang dan Transfeksinya pada Sel Kanker SP-C1, Tesis, Fakultas Farmasi Universitas Gadjah Mada, Yogyakarta.
Nguyen, L.T., Atobe, K., Barichello, J.M. Ishida, T., and Kiwada, H., 2007, Complex Formation with Plasmid DNA Increases The Cytotoxicity of Cationic Liposomes, Biol. Pharm. Bull., 30(4):751-7
Patil, S.D., Rhodes, D.G. and Burgess, D.J., 2005, DNA-based Therapeutics and DNA Delivery Systems: A Comprehensive Review, AAPS J, 7: E61 - E77
Roy, K., Mao, H.Q., Huang, S.K. and Leong, K.W., 1999, Oral Gene Delivery with Chitosan-DNA Nanoparticles Generates Immunologic Protection in A Murine Model of Peanut Allergy, Nat. Med., 4:387-391.
Sambrook, J., Fritsch, E., F. and Maniatis, T., 1989, Molecular Cloning, A Laboratry Manual 2nd, Cold
Spring Harbor Laboratory Press, volume 1.
Supriatno, K., and Harada, 2005, Basic Investigation on The Development of Molecular Targetting Therapy Against Cyclin-dependent Kinase Inhibitor p27Kip1 in Head and Neck Cancer Cells, Int. J. Oncol., 27:627-635
Vihola, H., Laukkanen, A., Valtola, L., Tenhu, H. and Hirvonen, J., 2005, Citotoxicity of Thermosensitive Polymers Poly(N-Isopropylacrylamide), Poly(N-Vynilcaprolactam), Biomaterials, 26(16):3055-3064
DOI: http://dx.doi.org/10.14499/indonesianjpharm0iss0pp204-211
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