Jean-Jacques Yaouanc

Chemical vectors for gene delivery: a current review on polymers, peptides and lipids containing histidine or imidazole as nucleic acids carriers

DNA/cationic lipid (lipoplexes), DNA/cationic polymer (polyplexes) and DNA/cationic polymer/cationic lipid (lipopolyplexes) electrostatic complexes are proposed as non‐viral nucleic acids delivery systems. These DNA‐nanoparticles are taken up by the cells through endocytosis processes, but the low capacity of DNA to escape from endosomes is regarded as the major limitations of their transfection efficiency. Here, we present a current report on a particular class of carriers including the polymers, peptides and lipids, which is based on the exploitation of the imidazole ring as an endosome destabilization device to favour the nucleic acids delivery in the cytosol. The imidazole ring of histidine is a weak base that has the ability to acquire a cationic charge when the pH of the environment drops bellow 6. As it has been demonstrated for poly(histidine), this phenomena can induce membrane fusion and/or membrane permeation in an acidic medium. Moreover, the accumulation of histidine residues inside acidic vesicles can induce a proton sponge effect, which increases their osmolarity and their swelling. The proof of concept has been shown with polylysine partially substituted with histidine residues that has caused a dramatic increase by 3–4.5 orders of magnitude of the transfection efficiency of DNA/polylysine polyplexes. Then, several histidine‐rich polymers and peptides as well as lipids with imidazole, imidazolinium or imidazolium polar head have been reported to be efficient carriers to deliver nucleic acids including genes, mRNA or SiRNA in vitro and in vivo. More remarkable, histidylated carriers are often weakly cytotoxic, making them promising chemical vectors for nucleic acids delivery.

Progress in Cationic Lipid-Mediated Gene Transfection: A Series of Bio- Inspired Lipids as an Example

Over the last several years, various gene delivery systems have been developed for gene therapy applications. Although viral vector-based gene therapy has led to the greatest achievements in animal and human studies, synthetic non-viral vectors have also been developed as they offer several advantages over viral systems, including lower immunogenicity and greater nucleic acid packaging capacity. Nevertheless, the transfection efficiency of the current non-viral gene carriers still needs to be improved, especially as regards direct in vivo transfection. In particular, cationic lipid/nucleic acid complexes (termed lipoplexes) have been the subject of intensive investigation with a view to optimize their performance and to better understand their mechanisms of action, and consequently to design new approaches to overcome the critical barriers of cationic liposome-mediated gene delivery. A possible strategy may rely on considering the membrane constituents and properties of the vast variety of living organisms as a source of inspiration for the design of biocompatible, non-toxic and effective novel artificial liposomal systems. Thus, the present forward-looking review provides an overview of the progress already made during the last years in the field of cationic lipid-mediated gene transfection and also focuses on a series of novel bio-inspired lipids for both in vitro and in vivo gene transfection.

Synthesis and Transfection Activity of New Cationic Phosphoramidate Lipids: High Efficiency of an Imidazolium Derivative

In an effort to enhance the gene‐transfer efficiencies of cationic lipids and to decrease their toxicities, a series of new phosphoramidate lipids with chemical similarity to cell membrane phospholipids was synthesised. These lipids contained various cationic headgroups, such as arginine methyl ester, lysine methyl ester, homoarginine methyl ester, ethylenediamine, diaminopropane, guanidinium and imidazolium. Their transfection abilities, either alone or with the co‐lipid DOPE, were evaluated in HEK293–T7 cells. We found that imidazolium lipophosphoramidate 7 a/DOPE lipoplexes gave the most efficient transfection with low toxicity (15 %). The luciferase activity was 100 times higher than that obtained with DOTAP/DOPE lipoplexes. The size, ζ potential, pDNA–liposome interactions and cellular uptakes of the lipoplexes were determined. No definitive correlation between the ζ potential values and the transfection efficiencies could be established, but the uptake of lipoplexes by the cells was correlated with their final transfection efficiencies. Our results show that imidazolium phosphoramidate lipids constitute a potential new class of cationic lipids for gene transfer.

Cationic Phosphonolipids Containing Quaternary Phosphonium and Arsonium Groups for DNA Transfection with Good Efficiency and Low Cellular Toxicity

Replacing the ammonium polar head in cationic lipids 1 (A=N) by a phosphonium or an arsonium group (A=P, As) improves their properties as synthetic vectors for DNA transfection. The increased volume of the cationic head is supposed to modify the interactions of the vector with the solvent and DNA.

A novel cationic lipophosphoramide with diunsaturated lipid chains: synthesis, physicochemical properties, and transfection activities.

Cationic lipophosphoramidates constitute a class of cationic lipids we have previously reported to be efficient for gene transfection. Here, we synthesized and studied a novel lipophosphoramidate derivative characterized by an arsonium headgroup linked, via a phosphoramidate linker, to an unconventional lipidic moiety consisting of two diunsaturated linoleic chains. Physicochemical studies allowed us to comparatively evaluate the specific fluidity and fusogenicity properties of the liposomes formed. Although corresponding lipoplexes exhibited significant but relatively modest in vitro transfection efficiencies, they showed a remarkably efficient and reproducible ability to transfect mouse lung, with in vivo transfection levels higher than those observed with a monounsaturated analogue previously described. Thus, these results demonstrate that this diunsaturated cationic lipophosphoramidate constitutes an efficient and versatile nonviral vector for gene transfection. They also invite further evaluations of the transfection activity, especially in vivo, of gene delivery systems incorporating the lipid reported herein and/or other lipids bearing polyunsaturated chains.

A new route to cyclen, cyclam and homocyclen

Abstract Cyclen, cyclam and homocyclen have been synthesized from the corresponding butanedione-protected linear tetramines. The cyclization step is followed by a facile deprotection of the rigidifying moiety.

Novel neutral imidazole-lipophosphoramides for transfection assays

New helper lipids, possessing an imidazole polar head, have been synthesized and included in formulations for transfection assays; these new helper lipids can improve the transfection by a factor of up to 100 compared to the use of DOPE as co-lipid.

GENERAL ROUTE FOR THE SYNTHESIS OF MONO N-ALKYLATED DERIVATIVES OF TETRAAZAMACROCYCLES

Abstract The selective synthesis of mono N-substituted derivatives of tetraazamacrocycles can be achieved using a new boron triprotection easily removed after alkylation.

Selective gene delivery in dendritic cells with mannosylated and histidylated lipopolyplexes

We report for the first time preparation of mannosylated and histidylated lipopolyplexes (Man-LPD100) with uptake and transfection selectivity for dendritic cells (DCs). Man-LPD100 were prepared by addition of mannosylated and histidylated liposomes (Man-Lip100) on preformed PEGylated histidylated polylysine/DNA polyplexes. Man-Lip100 comprised a cationic [O,O-dioleyl-N-(3N-(N-methylimidazolium iodide)propylene) phosphoramidate)] lipid, a neutral [O,O-dioleyl-N-histamine Phosphoramidate] co-lipid and β-D-mannopyranosyl-N-dodecylhexadecanamide (Man-lipid). At the best, Man-Lip100 containing 11 mol % Man-lipid was obtained. We found that dialysis of liposomes completely abolished cytotoxicity. We showed that the uptake of Man11-LPD100 by the murine DC line (DC2.4 cells) was at least 10-fold higher than that of Lac6-LPD100. A confocal microscopy study with DC2.4 cells expressing Rab5-EGFP or Rab7-EGFP, revealed that DNA uptake occurred through clathrin-mediated endocytosis. The transfection of DC2.4 cells with Man11-LPD100 containing DNA encoding luciferase gene gave luciferase activity two to three times higher (9 × 105 RLU/mg protein) than with non-mannosylated LPD100. In contrast to the latter, it was inhibited by 90% in the presence of mannose. Overall, the results indicate that mannosylated and histidylated LPD is a promising system for a selective DNA delivery in DCs.

Condensation of glyoxal with triethylenetetraamine. Stereochemistry, cyclization and deprotection

Abstract A mixture of bis-aminal isomers was obtained from aqueous glyoxal-triethylenetetraamine condensation. The irreversible isomerization of the species was observed and a new synthesis of cyclen proposed.