JCJC SIMI 8 - JCJC - SIMI 8 - Chimie du solide, colloïdes, physicochimie

Macromolecular membrane permeabilizers with crown ethers located each 3 carbons on the polymer backbone – MIMIC3C

Biomimetic oligomers for cancer treatment

Crown-ethers containing membranar permeabilizers, interaction with artificial and biological membranes

Membrane permeabilization induced by ion-binding cavities containing polymers

Lipid bilayers provide the basic structure of cell membranes and act as an impermeable barrier to the passage of most molecules. However, internal specific cellular transport processes exist to allow them to cross the cell membrane. This regulation of bilayer properties makes controlled incorporation of external agents very difficult. Designing novel synthetic agents that are able to affect membrane properties and induce a controlled permeabilization while keeping the integrity of membrane is hence a major challenge for academic chemistry and biophysics, applications in pharmaceutical research, and biotechnologies. <br />In this context, the overall objective is to develop a new type of membrane permeabilizers based on ion selective cavities such as crown-ethers that bind selectively K+ and Na+ (crucial ions for the metbolism). Their originality lies in a unique structure with single or geminated macrocycles located on each three carbons along the main chain. They are thus named “3C” poly(crownethers). <br />We propose to explore structure-activity relationships within the set of oligomeric membrane permeabilizers prepared. In order to evaluate the impact of these new poly(crown-ethers) as membrane permeabilizers, we propose to synthesize vinylic analogues, “2C” poly(crownethers) (substituents located on each two carbons along the main chain) and to compare their ability to permealize lipidic membrane. <br />

Polymers are obtained by polymerization of crown-ethers binding cavities containing monomers previously prepared in laboratory. “3C” poly(crown-ethers) are obtained according to a methodology we recently developed and that leads to well controlled polymer size and polydisperdity. The synthesis of vinylic analogues (« 2C » poly(crownethers)) with close chemical structure but a different spacial repartition of crown-ethers along the main chain has been explored by polymerization of crown-ethers containing methacrylate monomers.
Complexation properties of monomers and polymers are studied by titration (using microcalorimetry and alcali metal salts extraction).
Interaction of polymers with artificial bilipidic membranes is investigated by conductance measurements on planar bilipid membranes (in collaboration with the Slovac Academy of science) and by fluorescence spectroscopy on vesicles (using a method described in the literature and and set-up in the laboratory).

A new family of poly(crown-ethers) with different size and crown-ether size and density has been prepared and fully characterized. The so-called “3C” poly(crownethers) display binding properties towards sodium and potassium. Due to an active impuritity that induces sample ageing, lots of data obtained from fluorescence spectroscopy on vesicles have been discarded and it has not been possible to establish structure/properties relationships. Nevertheless, in the framework of an Hubert Curien grant with the Slovak academy of science in Bratislava, it has been shown that “3C” poly(crownethers) could form cation selective pores within bilipidic planar membranes. Moreover, biological assays performed in collaboration with CRRET laboratory in Créteil revealed the anti-proliferation ability of the molecules towards cancer cells (experiments with cells and mice have been done). That has been patented.
Polymerization of crownether substituted methacrylate monomers has been explored in order to obtain “2C”poly(crownethers). This was not successful as uncontrolled or spontaneous polymerization has been observed.
Due to impurity and synthesis problems described above, a new synthesis pathway based on the post-modification of polymeric scaffolds has been developed. In collaboration with Département de Chimie Moléculaire in Grenoble, this strategy appeared to be successful to prepare hydrosoluble biomimetic oligomers. In particular, it has been used to developed new polymers with pendant carbohydrates (glycopolymers) with protein recognition properties.

Post-modification of a polymeric scaffold to prepare new glycopolymers for protein recognition is the first step of a new project in collaboration with Département de Chimie Moléculaire in Grenoble. The overall objective is the development of sulfated oligosaccharides mimics (called glycoaminoglycans) to investigate their interaction with some proteins involved in normal and pathological biological processes.

- An article on the synthesis and ion binding properties of a new family of poly(crownethers).
- A note showing the ability of poly(crownethers) to form cation-selective pores within bilipidic membranes.
- A patent motivated by the potential impact of poly(crownethers) as anti-proliferative agents.
- A communication on glycopolymers for protein recognition and their synthesis by a post-modification strategy.
- A conference preprint dealing with polymers for molecular (ion or protein) recognition

Lipid bilayers provide the basic structure of cell membranes and act as an impermeable barrier to the passage of most molecules. However, internal specific cellular transport processes exist to allow transmembrane exchanges. This regulation makes controlled incorporation of external agents, such as drug molecules very difficult.
Designing novel synthetic agents that are able to affect membrane properties and induce a controlled permeabilization while keeping the integrity of membrane is hence a major challenge for academic chemistry and biophysics, applications in pharmaceutical research, and biotechnologies.

In this context, MIMIC3C aims to develop an original family of polymeric membranar permeabilizers. In order to generate ion-selective pores, polymers with pendant crown-ethers will be considered. A new type of poly(crown-ethers) is proposed. Their originality lies on a unique structure with single or geminated macrocycles located on each three carbons along the main chain. This particular stereochemistry induces absolutely unusual inter-substituents distances. Polymerization will be performed according to a metal free activation developed by Barbier et al. A phosphazene base is used in association with a protic molecule to initiate the polymerization. This novel initiating system leads to a rather high reactivity allowing the polymerization of bulky substituents containing monomers. Three structural parameters will be varied: the polymer size, the crown-ether size (i.e. ion selectivity) and the number of crown-ether per repetition unit.

The first objective is to synthezise this new family of membrane permeabilizers.

In contrast to small-and-medium-sized organic molecules, the prediction of exact conformations and mode of action of polymeric permeabilizers within a lipidic membrane is much more complex and largely uninvestigated.
The second objective is to explore structure-activity relationships within the set of oligomeric membrane permeabilizers prepared. In particular, we will study the influence of the three structural parameters mentionned previously on the polymer membrane permeabilization properties (pore diameter, permeation kinetics, ion selectivity, inclusion stability).
The permeabilization properties will be investigated using two methods: fluorescence kinetics in vesicles (HPTS assays) and conductance measurements on planar bilayers (BLM experiments). Both methods are complementary. Indeed, whereas fluorescence emission spectroscopy reflects the global membrane permeabilization, electrical measurements can provide information on the conductance and the dynamics of a single pore. HTPS assays will be developed in our group. BLM experiments will be performed in collaboration with phycisists (Intitute of Molecular Physiology and Genetics, Slovakia).

Finally, in order to evaluate the impact of these new poly(crown-ethers) as membrane permeabilizers, we propose to synthesize vinylic analogues and to compare their ability to permealize lipidic membrane. For this purpose, substituted methyl methacrylates appear to be the most convenient and will be prepared and their permeabilization properties will be studied acoording to the same methods.

Project coordination

Valessa BARBIER (CENTRE NATIONAL DE LA RECHERCHE SCIENTIFIQUE - DELEGATION REGIONALE ILE-DE-FRANCE SECTEUR EST) – barbier@icmpe.cnrs.fr

The author of this summary is the project coordinator, who is responsible for the content of this summary. The ANR declines any responsibility as for its contents.

Partner

ICMPE - UMR 7182 CENTRE NATIONAL DE LA RECHERCHE SCIENTIFIQUE - DELEGATION REGIONALE ILE-DE-FRANCE SECTEUR EST

Help of the ANR 161,549 euros
Beginning and duration of the scientific project: October 2011 - 36 Months

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