Efficiency of plasmid delivery and expression after lipid-mediated gene transfer to human cells in vitro

Zsuzsa Bebök, Anna M. Abai, Jian Yun Dong, Scott A. King, Kevin L. Kirk, Gabor Berta, Brian W. Hughes, Andrew S. Kraft, Stephen W. Burgess, Walter Shaw, Philip L. Felgner, Eric J. Sorscher

Research output: Contribution to journalArticle

30 Scopus citations

Abstract

Cationic liposome-mediated gene transfer has become increasingly important in the development of experimental therapies for human diseases, such as melanoma, human immunodeficiency virus infection, cystic fibrosis and alpha-1 antitrypsin deficiency. However, very little is known about the mechanisms by which lipid-mediated gene transfer occurs. We studied the kinetics of plasmid delivery and expression by using this technique. Plasmid entry in the cystic fibrosis respiratory epithelial cell line 2CFSME0-, as well as in two other cell lines (HeP 2g and HeLa) occurred in 95 to 100% of cells within 1 hr of the initiation of lipid-mediated gene transfer. In hepatic and respiratory cells, transcription of a construct containing the cystic fibrosis transmembrane conductance regulator was observed in more than 80% of the cell population; similarly high levels of plasmid utilization were obtained in studies of HLA-B7 expression in human melanoma cells. Studies directly relevant to current human trials of lipid-mediated gene transfer indicate that plasmid entry, transcription and translation are often surprisingly efficient, and may occur in nearly 100% of human cells in culture when sensitive methods for detection are used. Furthermore, conventional X-gal immunohistochemistry markedly underestimates transfection efficiency during transient gene expression. These studies point to a new mechanistic understanding of the features that limit expression by using cationic liposomes.

Original languageEnglish (US)
Pages (from-to)1462-1469
Number of pages8
JournalJournal of Pharmacology and Experimental Therapeutics
Volume279
Issue number3
StatePublished - Dec 1 1996
Externally publishedYes

ASJC Scopus subject areas

  • Molecular Medicine
  • Pharmacology

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    Bebök, Z., Abai, A. M., Dong, J. Y., King, S. A., Kirk, K. L., Berta, G., Hughes, B. W., Kraft, A. S., Burgess, S. W., Shaw, W., Felgner, P. L., & Sorscher, E. J. (1996). Efficiency of plasmid delivery and expression after lipid-mediated gene transfer to human cells in vitro. Journal of Pharmacology and Experimental Therapeutics, 279(3), 1462-1469.