Development of an effective gene delivery system: A study of complexes composed of a peptide-based amphiphilic DNA compaction agent and phospholipid

Eric A. Murphy, Alan J. Waring, Jason C. Murphy, Richard C. Willson, Kenneth J. Longmuir

Research output: Contribution to journalArticle

20 Scopus citations

Abstract

We recently described a basic technology to efficiently combine compacted DNA with phospholipids and hydrophobic peptides, to produce homogenous complexes that are completely resistant to nuclease. We have developed this technology further to form gene delivery complexes that transfect cells effectively in vitro. In addition to plasmid DNA, the complexes contained two basic components: (i) a DNA compacting peptide (-CGKKKFKLKH), either conjugated to lipid or extended to contain (WLPLPWGW-) and (ii) either phosphatidylethanolamine or phosphatidylcholine. Complexes containing a 5.5-fold charge equivalence (peptide charge/DNA charge) of WLPLPWGWCGKKKFKLKH and 5 nmol dimyristoleoylphosphatidyl-ethanolamine/μg DNA produced the highest luciferase gene expression, exceeding 1 × 109 relative light units/s/mg protein (>3 μg luciferase per mg protein). These complexes transfected OVCAR-3, COS-7 and HeLa cells at either similar or superior levels when compared to polyethylenimine or lipofectamine complexes. With green fluorescent protein reporter gene, >50% of HeLa cells were positive 30 h after addition of these complexes. Furthermore, these optimal complexes were the least sensitive to pre-treatment of cells with chloroquine, indicating efficient endosomal escape. Our results indicated that self-assembling complexes of plasmid DNA, amphiphilic peptide and phosphatidyl-ethanolamine are highly effective non-viral gene delivery systems.

Original languageEnglish (US)
Pages (from-to)3694-3704
Number of pages11
JournalNucleic Acids Research
Volume29
Issue number17
DOIs
StatePublished - Sep 1 2001

ASJC Scopus subject areas

  • Genetics

Fingerprint Dive into the research topics of 'Development of an effective gene delivery system: A study of complexes composed of a peptide-based amphiphilic DNA compaction agent and phospholipid'. Together they form a unique fingerprint.

Cite this