No description available.

The increasing number of nucleic acid-based therapeutics demonstrates the potential to treat diseases at the genetic level. Although oligonucleotides show clinical potential, challenges remain including nuclease degradation, rapid clearance when administered systemically, low cell permeability, and limited distribution to tissues of interest. This is largely imparted by the polyanionic phosphate backbone, which produces unfavourable electrostatic interactions at cell membranes. As a result, their clinical translation is dependent on delivery technologies that improve stability, facilitate cell entry, and increase target affinity. Spherical nucleic acids (SNAs) consist of radially orienting linear nucleic acids onto a nanoparticle core, conferring them a three-dimensional, spherical architecture. These structures enter cells readily and display distinct properties that are independent of their nanoparticle core. Accordingly, we decided to replace the intrinsically anionic phosphodiester linkage of DNA with a phosphoramidate linkage (P-N), allowing us to incorporate new functionality at the phosphate backbone. With this handle, we inserted cationic and hydrophobically modified functional groups that were compatible with nanoscale architectures, giving rise to new properties relevant in biological contexts. Specifically, amine and guanidinium derivatized functional groups provided SNAs with a ~10-fold increase in cell uptake at early incubation times compared with unmodified SNAs. This demonstrates that we can tune the behaviour of SNAs with phosphate backbone modifications in a highly controlled manner. We hypothesize that the stringent control over location and placement of functional groups within the SNA framework will afford them favourable interactions at cell membranes, not only increasing their cell uptake, but also access to alternative uptake mechanisms and potency as therapeutics. / Thesis / Master of Science (MSc) / Oligonucleotides are short synthetic sequences of DNA or RNA that have the capacity to treat diseases at the genetic level. However, they face challenges such as degradation, low cell uptake, and poor tissue distribution. To overcome this issue, we plan to incorporate chemical modifications at the phosphate backbone of oligonucleotides to make them more stable and facilitate more favourable interactions at cell membranes. Conferring oligonucleotides into a 3D arrangement further enhances their stability and cell uptake relative to linear oligonucleotides. By densely functionalizing them onto a nanoparticle core, we can create spherical nucleic acids (SNAs). We hypothesize that the modifications imparted onto the phosphate backbone of linear oligonucleotides will translate their properties into SNAs. The new properties afforded to the SNAs will provide increased cell uptake, alternative uptake mechanisms, and access to cytosolic and nuclear targets, highlighting their potency and therapeutic potential.

Cette thèse s’inscrit dans le cadre de synthèses de matériaux innovants contenant des entités cationiques que sont le guanidinium et l’ammonium. Ces entités cationiques confèrent des propriétés intéressantes et fonctionnelles pour chacun des systèmes ioniques suivants : tensio-actifs, liquides ioniques et ionosilices. A cet effet, nous avons procédé à l’étude de deux familles de composés : les sels de guanidiniums et les ionosilices. Pour les sels de guanidiniums, nous avons étudié la formation et les propriétés d’auto-assemblage de tensio-actifs en utilisant différentes techniques de mesures (conductivité, tension de surface et calorimétrie). Ce remarquable synthon moléculaire qu’est le guanidinium a été aussi mis en avant comme liquide ionique pour l’extraction du méthyl orange, du diclofenac et du chromate. Quant aux ionosilices, bien qu’ils présentent aussi des propriétés intéressantes pour l’extraction ionique et l’adsorption de principes actifs, leur mise en forme reste cependant un paramètre clef pour cibler leur application. En effet, la mise en forme des ionosilices en nanoparticules permet l’extension des applications dans le domaine de la nanomedecine. Ainsi, durant cette thèse, des nanoparticules avec des sous-structures ioniques ammoniums sont synthétisées pour la première fois et utilisées comme nano-vecteur pour le relarguage d’un anti-inflammatoire (diclofenac). Par ailleurs, dans le but d’une extraction ionique en flux continu, des matériaux contenant des fonctions ioniques sous forme de monolithe ont été synthétisés à partir de précurseur ammonium par voie sol gel. Ainsi cette thèse nous a permis de trouver les éléments théoriques, illustratifs et expérimentaux des différentes facettes de la formation de matières à base d’unités cationiques aux propriétés remarquables que sont les sels de guanidiniums et les sels d’ammoniums. / This dissertation deals with innovative synthetic materials bearing cationic entities that are guanidinium and ammonium. These cationic entities give interesting and functional properties for each ionic system studied: surfactant, ionic liquid and ionosilica. For this purpose, we investigated two families groups composed of: guanidiniums salts and ionosilica. Regarding guanidiniums salts, we studied the formation and self-assembly behavior of guanidinium surfactants using different measurement techniques (conductivity, surface tension and calorimetry). This remarkable molecular synthon that represents guanidinium was also highlighted as an ionic liquid for the extraction of methyl orange, diclofenac and chromate. As for ionosilicas, although they also have advantageous properties for ion extraction and adsorption of the active ingredients, however their shaping remains a key parameter for targeting their application. In fact, the design of ionosilica material as nanoparticle allows applications extension in the field of nanomedicine. So during this thesis, nanoparticles containing ammonium substructures were synthesized for the first time and used as a nano-vector to deliver an anti-inflammatory drug (diclofenac). Furthermore, with the aim of ionic extraction in continuous flow, materials containing ionic functions as monolith were synthesized from ammonium precursor via sol gel route. This thesis allowed us to find the theoretical, experimental and illustrative elements of the different aspects of materials formation based on cationic entities with remarkable properties that are guanidiniums and ammonium salts.

L'objet de ce travail repose sur la recherche, le développement de nouveaux composés antibactériens, devant, selon le concept initial, interagir avec la paroi bactérienne. Ces derniers sont de nature polycationique, intégrant sur une structure calixarènique tridimensionnelle ou aromatique plan, des fonctions guanidiniums en nombre variable. Certains ont été développés en tant que prodrogues hydrosolubles, capables de libérer in vivo un principe actif (acide nalidixique). Le chapitre I porte sur introduction, l'étude bibliographique des calixarènes et leurs applications en biologie. Dans chapitre II, nous présentons les résultats (synthèse et caractérisation) obtenus pour les parents partiellement substitués du tétra-para-(guanidinoéthyl)calix[4]arène. Le but de ces synthèses est d'appréhender le rôle, l'importance du nombre et le positionnement des fonctions guanidines sur le calixarène. Nous abordons au chapitre III, les différentes étapes de synthèse, de caractérisation et l'évaluation par CLHP de nouvelles prodrogues hydrosolubles. Le comportement de prodrogue potentiel est ainsi démontré pour l'un d'entre eux. Dans le quatrième chapitre sont décrites les synthèses et caractérisations de nouvelles structures poly-ioniques, intégrant un motif central de type benzène, porteur d'un nombre variable de motifs latéraux para-(guanidinoéthyl)phényléther, et conçues comme une alternative aux calixarènes précédemment développés. Le chapitre V est dédié aux résultats des évaluations biologiques de l'ensemble de ces composés. Nous montrons que plusieurs des dérivés préparés, prodrogues ou non, présentent une activité anti-bactérienne très intéressante / The main objective of this work is based on the search and the development of new antibacterial compounds able to interact with a bacterial wall. The proposed compounds are of polycationic nature, elaborated on calixarene structure or an aromatic platform and containing guanidinium functions in various number and position. Some of them were developed as water-soluble prodrugs of the quinolone: nalidixic acid. The chapter I talk about introduction and a bibliographical study on calixarenes and their applications in biology. The chapter II presents the results obtained for the synthesis, the characterization of the partially substituted analogues of the tétra-para (guanidinoéthyl) calix[4]arene. The objective of this part is to understand the role and the importance of the number and the positioning of the guanidines functions on the calixarene core. In chapter III, are presented the various steps leading to the synthesis, the characterization and the evaluation by HPLC ways of new water-soluble prodrugs. The work is centered on a set of ionic derivatives of calix[4]arenes carriers of nalidixic acid. The prodrug behavior was demonstrated for one of them. The chapter IV presents the syntheses and characterizations of star-like poly-ionic structures are described, based on a benzene core, including a variable number of para (guanidinoéthyl) phényléther subunits. The chapter V is dedicated to the results of the biological evaluations of all the structure synthetized. For some compounds, viability and cellular toxicity were evaluated on eukaryotic cells. We show that many of the new prepared derivatives, prodrugs or not, present very interesting anti-bacterial activities

Une série de précurseurs contenant des sous-structures organo-ioniques à base d'entités imidazolium, guanidinium, ammonium et zwitterionique ainsi que des précurseurs neutres contenant des sous-structures amido-thiol, amine-thiol et amine a été synthétisée. Ces précurseurs ont été utilisés pour la synthèse de matériaux silices hybrides nanostructurés par voie template. La formation de matériaux nanostructurés a été réalisée par des réactions d'hydrolyse-polycondensation par l'utilisation de différents agents de structuration. Ce travail avait pour but la détermination des principaux facteurs influençant la structuration des matériaux. Ainsi, nous avons préparé une série d'ionosilicates nanostructurés par une nouvelle méthode de structuration qui met en jeu des interactions spécifiques entre des paires d'ions ‘précurseur cationique - surfactant anionique' et ‘précurseur anionique - surfactant cationique'. Cette approche a permis d'accéder aux matériaux originaux de type PMO ionique. Finalement, nous avons utilisé pour la première fois des surfactants de guanidinium comme ‘template' dans la synthèse des silices ioniques nanostructurées de morphologie sphérique. Les matériaux nanostructurés contenant des sous-structures amine, amine-thiol, ammonium et zwitterionique développés au cours de ce travail présentent des surfaces spécifiques élevées et une bonne accessibilité des sites organiques. Ces propriétés font de ces matériaux des systèmes de choix pour des applications en catalyse ou en séparation. / A series of precursors containing organo-ionic substructures such as imidazolium, guanidinium, ammonium and zwitterionic entities and several neutral precursors containing thiol-amide, thiol-amine and amino groups were successfully synthesized. These precursors were used for the synthesis of nanostructured silica hybrid materials containing ionic substructures via soft templating approaches. The formation of structured materials was achieved using template directed hydrolysis polycondensation procedures in the presence of various structure directing agents. The goal of this study was the determination of the parameters influencing the structuring of the materials. Thus, we prepared a series of nanostructured ionosilicates using a new method of structuring that is based on specific interactions between ‘cationic precursor - anionic surfactant' and ‘anionic precursor - cationic surfactant' ion pairs. This new strategy allowed the synthesis of ionic 'periodic mesoporous organosilicas'. At the end of this thesis, we used a new ‘guanidinium' type template in the preparation of nanostructured i-silica hybrid materials with a spherical morphology. Nanostructured ionosilicates bearing amine, amino-thiol, ammonium and zwitterionic substructures prepared in this work present high specific surface areas and a high accessibility of the organic functional sites. Due to these features, these materials have large potential in the fields of catalysis and separation.

In Streptococcus pyogenes, the protein SiaA (HtsA) is part of a heme uptake pathway system and involved in heme transfer from Shp to the ABC transporter. SiaA mutants, in which alanine replaces the axial histidine (H229) and methionine (M79) ligands, as well as a lysine (K61) and cysteine (C58) located near the heme propionates, are reported. Studies on a mutant of a cysteine expected to be at a distance from the propionates (C47A) are also reported. The coordination state and spin state of the selected mutants were determined via Resonance Raman studies. The pKa values of mutants ranged from 9.0 to 9.4, which were close to the pKa of the WT SiaA (9.7). The midpoint reduction potential of lysine (K61A) mutant was determined by spectroelectrochemical titration to be 61 ± 3 mV vs. SHE, similar to the WT protein (68 ± 3 mV). The addition of guanidinium hydrochloride resulted in protein denaturation that could show more than one process and occurred over days. The ease of protein unfolding was directly related to the extent of interaction of the residues with the heme: changes in the axial ligands resulted in far greater changes in heme protein stability than changes in the residues near the heme propionates. The causative agent of diphtheriae, Corynebacterium diphtheriae, imports heme via an ABC uptake transporter. In this research, two of the five proteins in the heme uptake pathway of C. diphtheriae were studied. These proteins were HmuT, lipoprotein component of the ABC transporter, and HtaA, the heme receptor. UV-visible spectroscopy and fluorescence spectroscopy showed that HmuT protein as isolated bound a porphyrin, rather than heme. Electrospray ionization mass spectrometry (ESI-MS) studies suggested that two tetrapyrroles were bound. To assess stability of this protein towards heme release, thermal denaturation studies were performed. For HtaA, UV-visible and fluorescence spectroscopy also showed the protein as isolated was also bound a porphyrin, rather than heme. Homology studies showed that HtaA protein is quiet distant from homologous heme uptake proteins and could be a member of novel heme binding domain family.

Aromatic stacked salt bridges are increasingly observed to play an important role in biology, suggesting that the two separate weak interactions cooperate with each other to mediate molecular recognition in a biological solution. In this thesis an in depth study was carried out in attempt to find the contribution of the guanidinium-carboxylate-aromatic triad in biological systems. Two different small molecule systems are used to carry out the study. From the results of the two chapters I proposed here that stacking aromatic ring enhances the salt bridge through desolvation effect. This hypothesis was also tested in a protein-protein interaction (Grb2 SH3 domain/SOS interaction). The most ideal peptide inhibitor cannot be obtained due to the synthetic difficulties. Limited result showed that increasing the hydrophobic area of the hot spot in this protein-protein interaction enhances the interaction. In researching the guanidinium-carboxylate-aromatic triad, we were inspired to study the pre-organization effect of 1,3,5-triethyl-2,4,6-trisubstituted benzene template. A computational and literature study done in this thesis showed that the installation of ethyl or methyl groups at 1,3,5 positions leads to consistent increases in binding affinity relative to unsubstituted hosts, but the amount of increase is non-trivial and varies with different substitutes. The installation of ethyl or methyl groups at 1,3,5 positions leads to consistent but relatively small increases in binding affinity relative to unsubstituted hosts. / Graduate

Die fortschreitende Entwicklung auf dem Gebiet der kontrollierten Polymerisationstechniken ermöglicht heutzutage die Synthese definierter Makromoleküle. Durch das Design der Primärstruktur dieser Makromoleküle kann ein starker Einfluss auf die sich bildende Sekundärstruktur ausgeübt werden. Um den Grad der Kontrolle über den Polymerisationsprozess zu erhöhen sollten in dieser Arbeit photoschaltbare Azobenzolfunktionalitäten in Guanidin- und Thioharnstoffmotive integriert werden, so dass durch die reversibel photoschaltbare Ausbildung von Wasserstoffbrücken die vorliegenden Polymerisationsprozesse beeinflusst werden können. In einem Ansatz wurden dazu neuartige azobenzolverknüpfte Guanidin- und Thioharnstoffkatalysatoren für die Ringöffnungspolymerisation (ROP) von Lactid (LA) synthetisiert. Im Fall der photoschaltbaren Guanidinkatalysatoren wurde eine Synthesemethode entwickelt, welche die zu Beginn langwierige Aufreinigung der Katalysatoren bedeutend vereinfacht und somit die Darstellung mehrerer Katalysatorgenerationen ermöglichte. Die erste Guanidinkatalysatorgeneration zeigte keine katalytische Aktivität. Durch die Synthese von verschiedenen Referenzguanidinen und deren Einsatz in der ROP von LA konnten die aromatischen Substituenten der Guanidinfunktionalität als Ursache der katalytischen Inaktivität identifiziert werden. Daraufhin wurde eine mit zwei Alkylsubstituenten versehene zweite Generation synthetisiert und erfolgreich in der ROP von Lactid eingesetzt. In einem anderen Ansatz wurde versucht ein azobenzolverknüpftes Guanidinium-Carboxylat-Zwitterion so zu gestalten, dass es als photoschaltbares Monomer zur Bildung supramolekularer Polymere verwendet werden kann. Hierzu wurden zwei Generationen photoschaltbarer Zwitterionen synthetisiert. Die Eigenschaften der zweiten Generation wurden mit verschiedenen spektroskopischen Methoden untersucht. Dabei wurden Hinweise auf die Bildung eines supramolekualren Polymers gefunden. / Following the progressive development in the field of controlled polymerizations, it is now possible to synthesize well defined macromolecular structures. Controlling the primary structure in these macromolecules significantly influences the secondary structure, allowing the preparation of smart materials. In order to improve the achievable degree of control, this work aims to incorporate azobenzene functionalities into guanidine and thiourea moieties and, through the photo triggered reversible formation of hydrogen bridges, influence polymerization processes. Novel azobenzene substituted guanidine and thiourea catalysts for the ring opening polymerization (ROP) of lactide were synthesized. In the case of the photoswitchable guanidine catalysts, a new synthetic protocol was developed to overcome the difficult purification of the catalysts, allowing the facile preparation of multiple catalyst generations. The first generation of photoswitchable guanidines showed catalytic activity. Synthesis of reference guanidine catalysts demonstrated a negative effect between aromatic guanidine substituents and the catalytic performance. Following this observation, a second generation of alkyl substituted guanidine catalysts was synthesized and applied successfully in the ROP of lactide. In a concurrent approach, a guanidinium carboxylate zwitterion was rendered photoswitchable by the incorporation of an azobenzene functionality and used as a monomer in supramolecular polymerization processes. The first generation of photoswitchable zwitterions showed promising photochemical properties, but its poor solubility in apolar, aprotic solvents prevented a final proof of the concept. To achieve this, a second generation of photoswitchable zwitterions was synthesized, incorporating solubilizing functionalities into the molecular design. The properties of the second generation zwitterion were examined by various spectroscopical methods, indicating the formation of supramolecular polymers.