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Single histidine residue in head-group region is sufficient to impart remarkable gene transfection properties to cationic lipids: evidence for histidine-mediated membrane fusion at acidic pH

Gene Therapy volume 10, pages 1206–1215 (2003)Cite this article

Abstract

Presence of endosome-disrupting multiple histidine functionalities in the molecular architecture of cationic polymers, such as polylysine, has previously been demonstrated to significantly enhance their in vitro gene delivery efficiencies. Towards harnessing improved transfection property through covalent grafting of endosome-disrupting single histidine functionality in the molecular structure of cationic lipids, herein, we report on the design, the synthesis and the transfection efficiency of two novel nonglycerol-based histidylated cationic amphiphiles. We found that L-histidine-(N,N-di-n-hexadecylamine)ethylamide (lipid 1) and L-histidine-(N,N-di-n-hexadecylamine,-N-methyl)ethylamide (lipid 2) in combination with cholesterol gave efficient transfections into various cell lines. The transfection efficiency of Chol/lipid 1 lipoplexes into HepG2 cells was two order of magnitude higher than that of FuGENETM6 and DC-Chol lipoplexes, whereas it was similar into A549, 293T7 and HeLa cells. A better efficiency was obtained with Chol/lipid 2 lipoplexes when using the cytosolic luciferase expression vector (pT7Luc) under the control of the bacterial T7 promoter. Membrane fusion activity measurements using fluorescence resonance energy transfer (FRET) technique showed that the histidine head-groups of Chol/lipid 1 liposomes mediated membrane fusion in the pH range 5–7. In addition, the transgene expression results using the T7Luc expression vector convincingly support the endosome-disrupting role of the presently described mono-histidylated cationic transfection lipids and the release of DNA into the cytosol. We conclude that covalent grafting of a single histidine amino acid residue to suitable twin-chain hydrophobic compounds is able to impart remarkable transfection properties on the resulting mono-histidylated cationic amphiphile, presumably via the endosome-disrupting characteristics of the histidine functionalities.

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Acknowledgements

We express our gratitude to Dr JP Behr (Strasbourg, France) for initiating this Indo-French collaboration. We thank warmly Drs L Huang and M Brisson (University of Pittsburgh, Pittsburgh, PA, USA) for 293T7 cells and pT7Luc, and Miss Laëtitia Million for her help in FRET measurements. This work was supported by grants from Agence Nationale de Recherche sur le Sida (ANRS) and Association pour la Recherche sur le Cancer (ARC). AC and VVK gratefully acknowledge the financial support received from the Department of Biotechnology, Government of India, New Delhi. PM is Research Director at INSERM. CP is Professor Assistant at the University of Orléans.

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Authors and Affiliations

  1. Division of Lipid Science and Technology, Indian Institute of Chemical Technology, India

    V V Kumar & A Chaudhuri

  2. Centre de Biophysique Moléculaire, CNRS UPR 4301, Orleans, Cedex, France

    C Pichon, M Refregiers, B Guerin & P Midoux

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  1. V V Kumar
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  4. B Guerin
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Kumar, V., Pichon, C., Refregiers, M. et al. Single histidine residue in head-group region is sufficient to impart remarkable gene transfection properties to cationic lipids: evidence for histidine-mediated membrane fusion at acidic pH. Gene Ther 10, 1206–1215 (2003). https://doi.org/10.1038/sj.gt.3301979

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