Copolymers Derived from Two Active Esters: Synthesis, Characterization, Thermal Properties, and Reactivity in Post-Modification.

Copolymers with two distinguished reactive repeating units are of great interest, as such copolymers might open the possibility of obtaining selective and/or consequent copolymers with different chemical structures and properties. In the present work, copolymers based on two active esters (pentafluorophenyl methacrylate and p-nitrophenyl methacrylate) with varied compositions were synthesized by Cu(0)-mediated reversible deactivation radical polymerization. This polymerization technique allows the preparation of copolymers with high to quantitative conversion of both comonomers, with moderate control over dispersity (D = 1.3-1.7). Additionally, by in-depth study on the composition of each copolymer by various techniques including elemental analysis, NMR, FT-IR, and XPS, it was possible to confirm the coherence between expected and obtained composition. Thermal analyses by DSC and TGA were implemented to investigate the relation between copolymers' composition and their thermal properties. Finally, an evaluation of the difference in reactivity of the two monomer moieties was confirmed by post-modification of copolymers with a primary amine and a primary alcohol as the model.

Interactions of Organosilanes with Fibrinogen and Their Influence on Muscle Cell Proliferation in 3D Fibrin Hydrogels.

Silanization of biomacromolecules has emerged as a fruitful approach to prepare hybrid biohydrogels. However, very little is known about interactions between organosilanes and biopolymers in solution. Here we focused on fibrin, a protein of interest in the biomedical field, whose self-assembly process and resulting gel structure are highly sensitive to experimental conditions. Three main silanes were selected to decipher the relative influence of the silanol groups and organic functions. Whereas no protein denaturation was observed, silanes bearing hydrophobic groups had a surfactant-like behavior and could improve the dispersion of fibrinogen molecules, impacting gel formation kinetics and rheological properties. 3D cultures of myoblasts evidenced that organosilanes could promote or impede cell proliferation, suggesting interactions of silanols with fibrin. These results demonstrate that the two sides of the coin of organosilane reactivity are relevant at different stages of fibrin gel formation and must be considered for future development of hybrid biomaterials.

Bridging ß-Cyclodextrin Prevents Self-Inclusion, Promotes Supramolecular Polymerization, and Promotes Cooperative Interaction with Nucleic Acids.

A bridge to assemble: Cyclodextrins bridged with an ammonium linker bearing a hydrophobic substituent can efficiently form supramolecular polymers and avoid the competing self-inclusion and head-to-head processes. Furthermore, the self-assembling cyclodextrin derivative interacts in a highly cooperative manner with DNA, as demonstrated by compaction experiments. It also interacts cooperatively with siRNA and allows its transfection.

Synthesis and characterization of a thermoresponsive polyoxometalate-polymer hybrid.

We report the synthesis of the first organo-POM with thermoresponsive properties. Our concept will provide chemists with a new tool to design POMs whose solubility is reversibly controllable through an external stimulus. POM-polymer TBA(7)[POM]-poly(N,N-diethylacrylamide) (POM-PDEAAm), was prepared by grafting PDEAAm-NH(2) (obtained by RAFT polymerization) onto the activated Dawson acyl-POM, α(2)-[P(2)W(17)O(61)SnCH(2)CH(2)C(=O)](6-). Extensive MS analysis was used to monitor the chain-functionalization steps and to confirm the formation of the hybrid. Aqueous solutions of the (NH(4))(7)[POM-PDEAAm] exhibited a LCST of 38 °C. Thus, the solubility/aggregation of the hybrid was reversibly controlled by changing the temperature. Above 38 °C, the solution became cloudy, and cleared again upon cooling. Dynamic light scattering (DLS) revealed the formation of small aggregates in the range 100 nm. We assumed that the charged POM head units prevented the formation of the larger-scattering aggregates that are usually observed for PDEAAm, and promoted the formation of micelle-like structures. The conjugate exhibited a temperature transition, which was different from that of the polymer and depended on the counterions associated with the POM. This result demonstrates the potential for merging organic (in this case, polymer) and inorganic structures to afford materials that exhibit new properties.

Conformational control of hydrogen-bonded aromatic bis-ureas.

The phenylurea moiety is a ubiquitous synthon in supramolecular chemistry because it contains strong complementary hydrogen bonding groups and is synthetically very accessible. Here we investigate the possibility to strengthen self-association by conformational preorganization of the phenylurea moiety. In fact, we show that it is possible to strongly enhance intermolecular interactions between hydrogen bonded aromatic bis-ureas by substitution at the ortho positions of the phenylurea groups. Ortho substituents enforce a noncoplanar conformation of the urea and phenyl moieties better suited for hydrogen bonding. Substitution by methyl groups is more efficient than with larger groups, probably because of reduced steric hindrance. These effects have been demonstrated in the case of two different supramolecular architectures, which points to the probable generality of the phenomenon. In addition, this study has led to the discovery of a new bis-urea able to form very stable self-assembled nanotubes in toluene up to high temperatures (boiling point) or low concentrations (10(-7) M) and in chloroform down to 3 × 10(-4) M.

Cationic and anionic lipoplexes inhibit gene transfection by electroporation in vivo.

BACKGROUND: Nonviral gene therapy still suffers from low efficiency. Methods that would lead to higher gene expression level of longer duration would be a major advance in this field. Lipidic vectors and physical methods have been investigated separately, and both induced gene expression improvement. METHODS: We sought to combine both chemical and physical methods. Cationic or anionic lipids can potentially destabilize the cell membrane and could consequently enhance gene delivery by a physical method such as electrotransfer. A plasmid model encoding luciferase was used, either free or associated with differently-charged lipoplexes before electrotransfer. RESULTS: Electrotransfer alone strongly enhanced gene expression after intramuscular and intradermal injection of naked DNA. On the other hand, cationic and anionic lipoplex formulations decreased gene expression after electrotransfer, whereas poorly-charged thiourea-based complexes, brought no benefit. Pre-injection of the lipids, followed by administration of naked DNA, did not modified gene expression induced by electroporation in the skin. CONCLUSIONS: The results obtained in the present study suggest that packing of DNA plasmid in lipoplexes strongly decreases the efficiency of gene electrotransfer, independently of the lipoplex charge. Non-aggregating complexes, such as poorly-charged thiourea-based complexes, should be preferred to increase DNA release.

Evaluation of the muscle gene transfer activity of a series of amphiphilic triblock copolymers.

BACKGROUND: Amphiphilic triblock copolymers such as the polyethylene oxide-polypropylene oxide-polyethylene oxide L64 (PEO(13)-PPO(30)-PEO(13)) significantly increase transgene expression after injection of DNA/polymer mixtures into skeletal muscles. To better understand the way such copolymers act, we studied the behaviour of different poloxamers, including L64, both in vitro and in vivo. METHODS: The in vitro and in vivo transfection activity of five copolymers that differ either by their molecular weight or by their hydrophilic/hydrophobic balance was evaluated. Furthermore, we also studied the membrane permeabilizing properties of the poloxamers. RESULTS: The results obtained indicate that, after intramuscular administration of DNA/poloxamer formulations, all five compounds were able to significantly increase the expression levels of luciferase compared to an injection of naked DNA. Using a LacZ expression cassette, up to 30% of the muscle fibers expressed the reporter gene. Furthermore, we show that the effect can be obtained using different promoters. Finally, we document that, to some extent, all five poloxamers possess membrane permeabilizing properties. CONCLUSIONS: Taken together, the results obtained in the present study show that there is a large flexibility in terms of molecular weight and EO/PO ratio for obtaining increased levels of transgene expression in vivo.

A single thiourea group is not enough to get stable thiourea lipoplexes.

The preparation of a new family of lipothiourea is reported using an automatic synthetic workstation. In these compounds the headgroups were made from single thiourea derivatives. The physicochemical properties and the transfection efficiency of several members of the family were studied. It was found that in the presence of DMPC small lipoplexes could be prepared. In opposite to the previously described di- and tri-lipothioureas most of these liposomes are unstable overtime. In addition, even the stable ones show no transfecting efficiency. All these data demonstrate that at least two thiourea groups are necessary to produce stable lipoplexes, to condense DNA and to give efficient transfection.

Controlling the nano-bio interface to build collagen-silica self-assembled networks.

Bio-hybrid networks are designed based on the self-assembly of surface-engineered collagen-silica nanoparticles. Collagen triple helices can be confined on the surface of sulfonate-modified silica particles in a controlled manner. This gives rise to hybrid building blocks with well-defined diameters and surface potentials. Taking advantage of the self-assembling properties of collagen, collagen-silica networks are further built-up in solution. The structural and specific recognition properties of the collagen fibrils are well-preserved within the hybrid assembly. A combination of calorimetry, dynamic light scattering, zetametry and microscopy studies indicates that network formation occurs via a surface-mediated mechanism where pre-organization of the protein chains on the particle surface favors the fibrillogenesis process. These results enlighten the importance of the nano-bio interface on the formation and properties of self-assembled bionanocomposites.

A comprehensive study in triblock copolymer membrane interaction.

Poloxamers are triblock copolymers made of poly(ethylene glycol)-(poly(propylene glycol))-poly(ethylene glycol). They have been shown to enhance gene transfer in the muscle, and co-administration of polymers and DNA appeared to be crucial to obtain this effect. It is questionable then if some interaction occurs between polymers and DNA. Polymer interaction with membranes represents a second crucial point due to the central hydrophobic part of the triblock copolymers. Besides, the question of the polymer spanning or adsorbing to the surface has not been solved by now. We addressed these issues by means of sensitive techniques that allowed working in diluted conditions and gaining in comprehension of gene transfection. By means of simultaneous time-correlated single-photon counting and fluorescence correlation spectroscopy, we have shown that the diffusion time of a single DNA molecule and PicoGreen lifetime was not altered in the presence of the triblock copolymer L64. Polypropylene (glycol) interactions with dodecylphosphocholine micelles were shown to occur at a deep level by (1)H NMR using doxyl probes located at the head or the lipid extremity of the micelles. The polypropylene (glycol) also interacted with lipid bilayers in a manner dependent on the cholesterol content, as shown by differential scanning calorimetry using liposomes. This interaction destabilised the membrane and allowed the release of small molecules. Finally, molecular dynamic simulation of the copolymer L64 in the presence of dodecylphosphocholine showed that the hydrophobic core of the polymer formed an extremely tight cluster, whose dimensions excluded the possibility of polymer spanning across the lipidic micelles. The simulation positively correlated with the destabilising effect observed on the liposomal membrane models.