New methods for the preparation of porous biomedical scaffolds have been explored for applications in tissue engineering and drug delivery. Scaffolds with controlled pore morphologies have been generated which incorporate cyclodextrin-drug inclusion complexes as the drug delivery component. Supercritical CO2 was explored as the main processing fluid in the complex formation and in the foaming of the polymer scaffold. The co-solvents, ethanol, ethyl acetate and acetone, were explored in each stage, as needed, to improve the solvent power of CO2. The first goal was to promote cyclodextrin-drug complex formation. Complex formation by traditional methods was compared with complex formation driven by processing in supercritical CO2. Complex formation was promoted by melting the drug in supercritical CO2 or in CO2 + co-solvent mixtures while in the presence of cyclodextrin. Some drugs, such as piroxicam, are prone to degradation near the drug's ambient melting temperature. However, this approach using CO2 was found to circumvent drug thermal degradation, since drug melting temperatures were depressed in the presence of CO2. The second goal was to produce porous polymeric matrices to serve as tissue engineering scaffolds. Poly(lactide-<i>co</i>-glycolide) and poly(ε-caprolactone) were investigated for foaming, since these biomedical polymers are already commonly used and FDA approved. Polymer foaming with CO2 is an alternative approach to conventional solvent-intensive methods for porosity generation. However, two major limitations of polymer foaming using CO2 as the only processing fluid have been reported, including the formation of a non-porous outer skin upon depressurization and limited pore interconnectivity. Approaches to circumvent these limitations include the use of a co-solvent and controlling depressurization rates. The effect of processing parameters, including foaming temperatures and depressurization rate, as well as co-solvent addition, were examined in polymer foaming using CO2. Drug release dynamics were compared for foams incorporated with either pure drug, cyclodextrin-drug physical mixture or cyclodextrin-drug complex. Pore morphology, polymer choice and drug release compound choice were found to alter drug release profiles. / Ph. D.

L’Imagerie par Résonance Magnétique est une technique d’imagerie possédant une des plus hautes résolutions d’images, utilisée pour le diagnostic clinique et en recherche. Cependant cette technique souffre d’une faible sensibilité intrinsèque due au faible contraste naturel entre les différents tissus. Pour pallier à cette limitation, des agents de contraste, principalement des complexes de Gd(III), sont actuellement utilisés et continus d’être développés. Ce projet de thèse s’inscrit dans la volonté de concevoir de nouveaux agents de contrastes plus efficaces et plus stables en utilisant la β- et l’α-cyclodextrines (CD) comme plateforme capables d’intégrer trois ou quatre fonctions pyridine-carboxylates ou pyridine-phosphonates, connues pour leur pouvoir chélatant du Gd(III). La première partie des travaux concerne les aménagements fonctionnels des CDs afin de pouvoir les fonctionnaliser régiosélectivement avec des bras chélatants du Gd(III) de types pyridino-carboxylate et pyridino-phosphonate. La deuxième partie traite de la caractérisation physico-chimique des nouveaux systèmes complexes Gd-ligands synthétisés (Gd-Lα et Gd-Lβ). Les principales propriétés caractérisant les agents de contraste ont été mesurées. Ainsi, de très bonne relaxivités ont été obtenue (25 mM-1s-1 < r1 < 40 mM-1s-1). Cependant, les stabilités des complexes se sont avérées faibles (4,24 < log KGdL < 5,58). En perspectives, il est nécessaire d’obtenir un juste équilibre entre une forte relaxivité et une stabilité élevée pour assurer la non-libération du Gd(III) toxique. / Magnetic Resonance Imaging is one of the most important and non-invasive tools for clinical diagnostics and biomedical researches. Nevertheless, this modality suffers from intrinsic low sensitivity. To overcome this limitation, contrast agents, mostly based on polyaminocarboxylate complexes of gadolinium are used. The aim of this project was to design new Gd(III)-based contrast agents using β- and α-cyclodextrins (CD), known to generate high relaxivity, functionalized with pyridine-carboxylate and pyridine-phosphonate ligands, known for their good affinity with the lanthanide cations. Here we first investigated the regio-functionalization of the β-CD on the primary face to access to scaffold called Lβ with four pyridine-carboxylate ligands. To develop an efficient and reproducible synthesis, HPLC analysis was implemented. Then, using these conditions, carboxylate- and phosphonate-ester, precursors to access α-CD ligands Lα and Lα’, were synthesized. The carboxylate-ester deprotection step successfully afforded the ligand noted Lα contrary to the deprotection of phosphonate ester which still have to be explored. Characterizations of these complicated Gd-ligands systems (Gd-Lα et Gd-Lβ) were reported. Their stability and relaxivity were measured and very good relaxivities were obtained (25 mM-1s-1 < r1 < 40 mM-1s-1). These new structures open the way to an improvement in term of stability.

L’utilisation de siRNA est une nouvelle approche thérapeutique très prometteuse. Néanmoins leur transfection à visée thérapeutique est un réel défi. Les obstacles à franchir pour élaborer des agents de transfection sûrs et fiables sont nombreux. Afin de les contourner nous nous sommes attachés à la construction d’un système dynamique qui, à l’image des virus, est constitué de briques moléculaires, s’emboitant et interagissant avec des acides nucléiques selon des interactions supramoléculaires. Ainsi, nous avons élaboré des polymères supramoléculaires polycationiques à base de monomères de cyclodextrines pontées, fonctionnalisées par un groupement adamantyle. Ce type de conjugué pallie un problème manifeste dans la littérature concernant les assemblages de β-CD souvent insolubles ou bien auto-inclus. L’ajout éventuel d’une autre fonction cationique pour améliorer l’interaction avec les siRNA a aussi été réalisé. Ainsi, la capacité à s’auto-assembler de quatre composés a été étudiée par RMN-1H, RMN-ROESY, ITC, RMN-DOSY, et SANS. Par ailleurs, ces composés ont montré une certaine capacité à complexer et à protéger les siRNA. L’un de ces composés a de plus montré une bonne aptitude à transfecter des siRNA in vitro, sans induire de toxicité. Les assemblages CD-siRNA ont finalement été observés par cryo-TEM et ont montré la formation de fibres, organisées de manière hiérarchique et hautement coopérative. Nous avons ainsi créé des assemblages supramoléculaires uniques à base d’acides nucléiques, rappelant la structure, la taille et la fonction d’un virus. / SiRNA based therapeutics are very promising. A key challenge for their development is the design of sophisticated, safe and effective delivery methods. To address all the biological obstacles for the conception of such therapeutics, we focused on the construction of a virus-like dynamic system, built with molecular bricks, able to self assemble and to interact with nucleic acid through supramolecular interactions. Bridged cyclodextrin based supramolecular polymers were developed to form host-guest interactions. To do so, cyclodextrins were conjugated with cationic and hydrophobic moiety in a spatially controlled way. These conjugates solved problems well known in the literature about the self-inclusion and the solubility in water of such molecules. The ability to self-assemble of 4 compounds were studied by RMN-1H, RMN-ROESY, ITC, RMN-DOSY and SANS. All these compounds showed a good capability to complex and protect siRNA. Moreover, one of these compounds is able to transfect siRNA in vitro without any toxicity, and therefore, to induce gene silencing. Assembly of CD and siRNA were finally observed by cryo-microscopy, which showed long fibres organised in a hierarchical and cooperative manner. This unique system is therefore strongly reminiscent of the structure, size and function of a virus.

This diploma work is devoted to the preparation and analytical testing of modified gold electrodes in QCM. The sensitivity and selectivity response towards selected aromatic analytes in aqueous phase was monitored. Studied analytes were 1-methylnaphthalene, 2-methylnaphthalene, naphthalene, o-xylene and toluene. These compounds are frequently used in chemistry industry and other industrial branches and, are known to be harmful to human organism and environment. Using QCM method it is possible to detect them rapidly and relatively easily.

The main aim of this work was to prepare and characterize a new chiral stationary phase (CSP). The CSP was prepared by dynamic coating of sulfobutylether- β-cyclodextrin (SBE-β-CD) on a strong anion-exchange stationary phase (SP). The selectivity and stability of the newly prepared CSP were tested on the sets of chiral and achiral analytes. The next separation system used was composed of C18 SP with the addition of SBE-β-CD as a chiral selector into the mobile phase. The set of chiral analytes contained amino acids phenylalanin, tyrosine, tryptophan, their derivatives and dipeptides glycine-DL-tryptophan and glycine-DL- phenylalanin. Four groups of mixtures of blocked and unblocked dipeptide isomers were tested for achiral separations. Dipeptides used for the mixtures had the same molecular formula but the sequence of amino acids was reversed. Measurements were carried out in reversed phase separation mode and hydrophilic interaction liquid chromatography. Mobile phases composed of methanol as an organic modifier and four different aqueous parts: (i) deionized water, (ii) aqueous solution of formic acid (pH 2.10), (iii) 20mM ammonium acetate buffer (pH 4.70) and (iv) 10mM ammonium acetate buffer (pH 8.80) in various volume ratios. Newly prepared SBE-β-CD CSP was more suitable for separations of...

The aim of this diploma thesis was to prepare two new chiral stationary phases by dynamic coating of sulphobutylether-β-cyclodextrin (SBE-β-CD) with varying degrees of substitution onto strong anion-exchange stationary phases. The enantioselective potential and stability of the newly prepared chiral stationary phases were tested using a set of chiral analytes. The set contained structurally diverse analytes, i.e. benzodiazepines (oxazepam, lorazepam), phenothiazines (thioridazine, promethazine), β-blockers (labetalol, pindolol, propranolol, alprenolol), profens (carprofen, fenoprofen, flurbiprofen, indoprofen), flavanones (6-hydroxyflavanone, 7-hydroxyflavanone), DL-tryptophan and its derivatives (5-OH-DL-tryptophan, 5-F-DL-tryptophan, DL-tryptophan butylester and blocked aminoacid (t-Boc-DL-tryptophan)), dipeptides (glycyl-DL-phenylalanine, glycyl-DL-tryptophan) and Troger's base. Measurements were carried out in reversed-phase high-performance liquid chromatography. Mobile phases consisted of methanol/formic acid (pH 2.10) and methanol/10mmol l-1 ammonium acetate buffer (pH 4.00) in various volume ratios. The chiral stationary phase containing hexasubstituted SBE-β-CD was suitable for enantioseparation of eleven analytes. Four of them were baseline enantioresolved and seven partially. The chiral...

På senare tid har forskning syftat till att främja miljövänligare teknik inom alla fält. Det visar sig inom materialframställningen som en önskan att använda material som inte är beroende av olja, inte tillverkade med tungmetaller samt nedbrytbara med en minimal miljöpåverkan. Detta har bland annat gett organiska katalysatorer en större plats inom forskningen. En av dessa är cyclodextrin, en cyklisk oligosackarid som har påvisats bilda makromolekylara komplex med andra molekyler. En aspekt av detta är att den kan hydrolysera polymerer då den bildar komplex med hydrofila molekyler och kan aktivera dessa genom vätebindning. Vilket Harada et. al. visade kunde utnyttjas for att polymerisera cycliska estrar. Detta examensarbete utforskar en del av det arbetet genom att dels verifiera polymerisationstekniken som ar en lösningsmedelsfri polymerisation dels försök till att framstalla en ny dimer av cyclodextrin som ska effektivt kunna polymerisera cycliska estrar mer effektivt an tidigare. Den nya dimern bygger på en thiourea-länk. Aven om polymerisation med vanlig cyclodextrin har visats fungera sa kan den föreslagna  dimern inte polymerisera lika effektivt. Syntesen ar problematisk och annars åtråvarda egenskaper hos cyclodextrin, som makromolekylär komplexbildning, kan inhibera polymerisationen och tidigare uppreningssteg.

Raloxifene is a highly insoluble, highly metabolized serum estrogen receptor modulator approved for use in the treatment of osteoporosis. Hydroxybutenyl-beta-cyclodextrin (HBenBCD) is a novel solubility enhancer previously demonstrated to increase the oral bioavailability of tamoxifen, letrozole, and itraconazole. The current study evaluated the pharmacokinetics of raloxifene in oral and intravenous formulations with HBenBCD in male Wistar-Hannover rats. Analytical methodology to measure raloxifene and its metabolites was developed by measuring raloxifene metabolism in vitro. Formulation with HBenBCD significantly increased raloxifene oral bioavailability. Mean ± S.D. oral bioavailabilities were 2.6 ± 0.4% for raloxifene formulated with microcrystalline cellulose, 7.7 ± 2.1% for a solid capsule formulation of raloxifene:HBenBCD complex, and 5.7 ± 1.3% for a liquid-filled capsule formulation containing raloxifene:HBenBCD/PEG400/H2O. Relative to raloxifene/microcrystalline filled capsules, the presence of HBenBCD in the solid capsule formulation afforded: (i) a decrease in raloxifene Tmax (2.5 ± 0.5 h versus 4.0 ± 0.5 h); (ii) a two-fold increase in raloxifene Cmax and a three-fold increase in raloxifene AUC; and (iii) a 12-fold increase in raloxifene glucuronide Cmax and a 6.5-fold increase in raloxifene glucuronide AUC. Hence, these studies demonstrate that raloxifene formulations containing HBenBCD significantly increased the oral bioavailability in rats relative to formulations that did not contain HBenBCD.

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Cardiac stem cells (CSCs) have emerged as promising cell candidates to regenerate damaged hearts, because of the potential in differentiating to cardiomyocytes. However, the differentiation is difficult to trigger without inducers. Here we reported that β-cyclodextrin (β-CD) increased the expression of cardiac transcription factors (Nkx2.5 and GATA4), structural proteins (cardiac Troponin T, cTnt), transcriptional enhancer (Mef2c) and induced GATA4 nucleus translocation in adult resident CSCs, thus β-CD could be used to enhance myogenic transition. As the differentiation process was accompanied by autophagy, we constructed the Atg5 knockdown cell line by using the Atg5 siRNA lentivirus, and the myogenic conversion was blocked in Atg5 knockdown cells, which suggested that β-CD induces the cardiomyocytes transition of resident CSCs through autophagy. Furthermore, we found that JNK/STAT3 and GSK3β/β-catenin was the downstream pathways of β-CD-induced autophagy and differentiation using the inhibitors. Moreover, β-CD performed its functions through improving intracellular cholesterol levels and affecting cholesterol efflux. Collectively, our results reveal that β-CD as a novel tool to induce myogenic transition of CSCs, which could mobilize the resident CSCs or used together with CSCs to enhance the therapy effects of CSCs on damaged hearts. In addition, the clarified molecular mechanisms supported the new targets for inducing cardiomyocyte differentiation.