Conception, synthese, et évaluation de systemes non cationiques de vectorisation de l'ADN

La recherche de vecteurs non viraux pour la thérapie génique est un domaine très développé. Les systèmes cationiques montrent une forte efficacité de transfection in vitro, mais cette activité est considérablement diminuée in vivo car les complexes ADN/vecteur, du fait de leur charge globale positive, sont rapidement éliminés de la circulation sanguine. Les lipopolythiourées sont des systèmes non cationiques qui représentent une alternative aux lipides cationiques: ils permettent de réduire de moitié l'élimination précoce après une injection intraveineuse. Dans ce mémoire est décrite la synthèse d'une famille de seize lipopolythiourées dont la structure repose sur une tête polaire branchée à deux motifs thiourée. Ces lipides présentent une grande diversité au niveau de l'ancre hydrophobe, de l'espaceur, du répartiteur et des terminaisons. L'évaluation de cette famille a été réalisée de façon systématique et la recherche du mécanisme d'action a été entreprise. Ces études ont permis la mise au point de lipopolythiourées d'une formulation facile et possédant un pouvoir transfectant du même ordre de grandeur que celui des lipides cationiques

Conception, synthèse et évaluation de systèmes de vectorisation non cationiques de l' ADN

PARIS-BIUSJ-Thèses (751052125) / SudocPARIS-BIUSJ-Physique recherche (751052113) / SudocSudocFranceF

Pharmaceutical Applications of Molecular Tweezers, Clefts and Clips

Synthetic acyclic receptors, composed of two arms connected with a spacer enabling molecular recognition, have been intensively explored in host-guest chemistry in the past decades. They fall into the categories of molecular tweezers, clefts and clips, depending on the geometry allowing the recognition of various guests. The advances in synthesis and mechanistic studies have pushed them forward to pharmaceutical applications, such as neurodegenerative disorders, infectious diseases, cancer, cardiovascular disease, diabetes, etc. In this review, we provide a summary of the synthetic molecular tweezers, clefts and clips that have been reported for pharmaceutical applications. Their structures, mechanism of action as well as in vitro and in vivo results are described. Such receptors were found to selectively bind biological guests, namely, nucleic acids, sugars, amino acids and proteins enabling their use as biosensors or therapeutics. Particularly interesting are dynamic molecular tweezers which are capable of controlled motion in response to an external stimulus. They proved their utility as imaging agents or in the design of controlled release systems. Despite some issues, such as stability, cytotoxicity or biocompatibility that still need to be addressed, it is obvious that molecular tweezers, clefts and clips are promising candidates for several incurable diseases as therapeutic agents, diagnostic or delivery tools

Lipopolythiourea Transfecting Agents: Lysine Thiourea Derivatives

International audienceSynthetic vectors represent an alternative to recombinant viruses for gene transfer. We have recently explored the transfection potential of a class of noncationic lipids bearing thiourea moieties as DNA associating headgroups. The encouraging results obtained with lipopolythioureas derived from serinol prompted us to investigate further this family of vectors. In the present study, we considered the transfection properties of a series of derivatives based on a different thiourea polar headgroup bearing a lysine scaffold. The synthesis of these compounds could be readily achieved in 3 steps with good yields. We found that these lipopolythioureas (LPT) might be considered as alternative systems for gene transfection, since their activity reached the same magnitude range as cationic vectors in the presence of serum. LPT with 14-carbon length chains appeared to be more efficient as transfecting agent than the ones with shorter chains. Toxicity studies proved that the hydration film 2 method led to particles well tolerated both by the cells in vitro and by the mice in vivo. The ability to induce gene expression in vivo was demonstrated by intratumoral injection. Finally, biodistribution studies showed that the quantity recovered in the blood circulation, 2h after systemic injection, was improved as compared to cationic lipids

Bile acid-based drug delivery systems for enhanced doxorubicin encapsulation: Comparing hydrophobic and ionic interactions in drug loading and release

Doxorubicin (Dox) is a drug of choice in the design of drug delivery systems directed towards breast cancers, but is often limited by loading and control over its release from polymer micelles. Bile acid-based block copolymers present certain advantages over traditional polymer-based systems for drug delivery purposes, since they can enable a higher drug loading via the formation of a reservoir through their aggregation process. In this study, hydrophobic and electrostatic interactions are compared for their influence on Dox loading inside cholic acid-based block copolymers. Poly(allyl glycidyl ether) (PAGE) and poly(ethylene glycol) (PEG) were grafted from the cholic acid (CA) core yielding a star-shaped block copolymer with 4 arms (CA-(PAGE-b-PEG)4) and then loaded with Dox via a nanoprecipitation technique. A high Dox loading of 14 wt% was achieved via electrostatic as opposed to hydrophobic interactions with or without oleic acid as a cosurfactant. The electrostatic interactions confer a pH responsiveness to the system. 50% of the loaded Dox was released at pH 5 in comparison to 12% at pH 7.4. The nanoparticles with Dox loaded via hydrophobic interactions, did not show such a pH responsiveness. The systems with Dox loaded via electrostatic interactions showed the lowest IC50 and highest cellular internalization indicating the pre-eminence of this interaction in Dox loading. The blank formulations are biocompatible and did not show cytotoxicity up to 0.17 mg/mL. The new functionalized star block copolymers based on cholic acid show great potential as drug delivery carriers

Switchable Lipid Provides pH-Sensitive Properties to Lipid and Hybrid Polymer/Lipid Membranes

Blending amphiphilic copolymers and lipids constitutes a novel approach to combine the advantages of polymersomes and liposomes into a new single hybrid membrane. Efforts have been made to design stimuli-responsive vesicles, in which the membrane’s dynamic is modulated by specific triggers. In this investigation, we proposed the design of pH-responsive hybrid vesicles formulated with poly(dimethylsiloxane)-block-poly(ethylene oxide) backbone (PDMS36-b-PEO23) and cationic switchable lipid (CSL). The latter undergoes a pH-triggered conformational change and induces membrane destabilization. Using confocal imaging and DLS measurements, we interrogated the structural changes in CSL-doped lipid and hybrid polymer/lipid unilamellar vesicles at the micro- and nanometric scale, respectively. Both switchable giant unilamellar lipid vesicles (GUV) and hybrid polymer/lipid unilamellar vesicles (GHUV) presented dynamic morphological changes, including protrusions and fission upon acidification. At the submicron scale, scattered intensity decreased for both switchable large unilamellar vesicles (LUV) and hybrid vesicles (LHUV) under acidic pH. Finally, monitoring the fluorescence leakage of encapsulated calcein, we attested that CSL increased the permeability of GUV and GHUV in a pH-specific fashion. Altogether, these results show that switchable lipids provide a pH-sensitive behavior to hybrid polymer/lipid vesicles that could be exploited for the triggered release of drugs, cell biomimicry studies, or as bioinspired micro/nanoreactors

Lipothioureas as Lipids for Gene Transfection: A Review

International audienceNon-viral gene therapy requires innovative strategies to achieve higher transfection efficacy. A few years ago, our group proposed bioinspired lipids whose interaction with DNA was not based on ionic interactions, but on hydrogen bonds. We thus developed lipids bearing a thiourea head which allowed an interaction with DNA phosphates through hydrogen bonds. After a proof of concept with a lipid bearing three thiourea functions, a molecular and cellular screening was performed by varying all parts of the lipids: the hydrophobic anchor, the spacer, the linker, and the thiourea head. Two lipothiourea-based structures were identified as highly efficient in vitro transfecting agents. The lipothioureas were shown to reduce non specific interactions with cell membranes and deliver their DNA content intracellularly more efficiently, as compared to cationic lipoplexes. These lipids could deliver siRNA efficiently and allowed specific cell targeting in vitro. In vivo, thiourea lipoplexes presented a longer retention time in the blood and less accumulation in the lungs after an intravenous injection in mice. They also induced luciferase gene expression in muscle and tumor after local administration in mice. Therefore, these novel lipoplexes represent an excellent alternative to cationic lipoplexes as transfecting agents. In this review we will focus on the structure activity studies that permitted the identification of the two most efficient thiourea lipids

Design, Synthesis, and Evaluation of Enhanced DNA Binding New Lipopolythioureas

International audienceNonviral gene delivery is limited to a large extent by the cationic nature of most of the chemical vector. We have shown that lipopolythioureas interact with DNA. However, lipopolythioureas were not very efficient at transfecting cells, probably due to reduced interaction between the noncationic synthetic lipid and the cell membrane. Here, we report that liposomes made from a new thiourea lipid, DPPC, and a lipid bearing an RGD ligand allowed very efficient entry of the lipopolythioureas into integrin R v 3 expressing cells. In addition, we show that a stable interaction between DNA and lipopolythiourea could be obtain with two thiourea groups. Moreover, the addition of a hydrophilic terminus improves the formulation of these new DNA binding agents