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Histidine containing peptides and polypeptides as nucleic acid vectors
Patrick Midoux1, Eric LeCam, Dominique Coulaud
1Centre de Biophysique Moléculaire, UPR 4301 CNRS, rue Charles Sadron, F-45071 Orléans 02, France.
Somatic Cell and Molecular Genetics
|May 31, 2003
Summary
Histidine-rich peptides enhance nucleic acid delivery into mammalian cells. These peptides, including histidylated oligolysine (HoK) and histidylated polylysine (HpK), significantly improve the transfer of plasmid DNA (pDNA) and oligonucleotides (ODN) into the cytosol and nucleus.
Area of Science:
- Biotechnology and Genetic Engineering
- Cell Biology and Molecular Medicine
- Drug Delivery Systems
Background:
- Efficient delivery of nucleic acids (plasmid DNA and oligonucleotides) into the cytosol of mammalian cells is crucial for gene therapy and research.
- Endocytosis is a primary pathway for cellular uptake, but endosomal escape remains a bottleneck for effective cytosolic delivery.
- Poly-L-histidine's membrane destabilization capacity at endosomal pH offers a potential strategy for improving nucleic acid transfer.
Purpose of the Study:
- To evaluate the efficacy of histidine-rich peptides (H5WYG, HoK, HpK) in enhancing the delivery of plasmid DNA (pDNA) and oligonucleotides (ODN) into mammalian cells.
- To investigate the role of polyhistidylated molecules in facilitating endosomal escape and cytosolic/nuclear delivery of nucleic acids.
- To assess the potential of these peptide-nucleic acid complexes as nonviral gene delivery systems.
Main Methods:
- Design and synthesis of histidine-rich peptides: H5WYG, histidylated oligolysine (HoK), and histidylated polylysine (HpK).
- Formation and characterization of complexes between peptides and nucleic acids (pDNA, ODN), including particle size and stability analysis (PEGylation).
- Assessment of transfection efficiency and biological activity of antisense ODN using these complexes in mammalian cells, with pH-dependent studies.
Main Results:
- H5WYG peptide facilitated cell membrane permeabilization at pH 6.4, enhancing pDNA transfection and enabling ODN loading into the cytosol and nucleus.
- HoK formed 35 nm spherical particles with ODN, and HpK formed 100 nm rod/toroid particles with pDNA; PEGylation improved stability.
- HoK/ODN complexes increased antisense ODN biological activity by over 20-fold, while HpK/pDNA complexes showed 3-4.5 orders of magnitude higher transfection efficiency than polylysine/pDNA complexes.
Conclusions:
- Polyhistidylated molecules, through imidazole protonation in endosomes, are effective agents for cytosolic delivery of nucleic acids.
- Histidylated polylysine (HpK) complexed with plasmid DNA (pDNA) represents a promising nonviral gene delivery system with significantly enhanced transfection efficiency.
- These findings highlight the potential of histidine-based peptides for advancing nucleic acid-based therapeutics and research tools.