No description available.

The delivery of immunoreactive macromolecules to tumour cells in solid, heterogeneously perfused tumours is a major problem in the effectiveness of immunotherapy. To help optimize the new experimental treatment method, a published mathematical model of macromolecule transport (Baxter & Jain 1989,1990,199la) was appraised and verified experimentally. Computational and analytical tools were developed to predict the interstitial plasma fluid pressure and velocity distributions in well perfused spherical tumours. Their published analytical solutions of the formulation were found to have some errors and were corrected in this work. To check the validity of the formulation, a series of animal experiments was performed to quantify the total vascular volume, and plasma fluid extravasation rate in SKOV3ipl human ovarian tumour xenografts in nude mice. The results compared well with the theoretically predicted total plasma fluid extravasation rate. Computer codes were also developed to predict the spatial and temporal distributions of intact IgG and its F(ab')₂ and Fab/Fab' fragments in well perfused spherical tumours using the formulation proposed by Baxter & Jain (1989,1990,1991a). The cases of non-binding and binding macromolecules were treated separately. The codes for both the interstitial pressure and macromolecule distributions were written to include a radially variable vessel surface area for transcapillary exchange per unit volume of tumour tissue (SN). The sensitivity of the overall tumour perfusion to variation of (a) the macromolecule m.w., binding affinity, and metabolism, (b) SN, tumour radius, and (c) microvascular permeability were investigated. Comparison of the theoretical predictions with available experimental data leads to the realization of a number of shortcomings in the previously proposed formulations. Finally, a computational method for deriving the effective spherically symmetric spatial distributions for the vascular volume density, and SN from tumour serial sections was developed. This bridges the gap between the actual topology of vascular distributions in tumours and the format of current formulations. / Thesis / Master of Science (MS)

La translocation, le passage d'une macromolécule à travers un pore inséré dans une membrane, est impliquée dans de nombreux processus biologiques. On peut citer comme exemple le transport d'ARN ou de protéines entre les composants de la cellule, et l'infection d'une cellule par le passage d'un ADN viral à travers la membrane cellulaire. Aujourd'hui la translocation est aussi la base d'applications technologiques, comme le fait d'utiliser les pores en tant que détecteurs pour le séquençage rapide de molécules ou en tant que filtre moléculaire. La compréhension du processus de translocation est importante à la fois d'un point de vue fondamental et pour la fabrication de nouveaux dispositifs de translocation à usage spécifique. Dans cette thèse, nous réalisons des expériences et des simulations informatiques pour étudier certains des effets les plus importants mis en jeu lors de la translocation.Nous utilisons des simulations informatiques avec un modèle à ``gros grain'' pour étudier qualitativement l'influence d'une interaction attractive entre les parois du pore et un polymère en train de transloquer. Nous montrons que la position de l'interaction influence la fréquence d'entrée et le temps de résidence du polymère dans le pore. La fréquence d'entrée est plus grande lorsque l'entrée du pore est attractive. Le comportement du temps de résidence avec la longueur du polymère est qualitativement et quantitativement affecté par la position de l'interaction dans le pore. Cependant, quelle que soit la position de l'interaction, nous observons que le temps de translocation augmente linéairement avec la longueur du polymère lorsque le polymère est plus long que le pore. Cette observation est qualitativement en accord avec des données expérimentales publiées.Lorsque la translocation est lente, la corrélation entre les mouvements des monomères confinés dans le pore peut jouer un rôle important. Cet effet n'a pas été pris en compte jusqu'à présent. Nous développons un nouveau modèle pour la translocation de polymères, inspiré par le processus d'exclusion asymétrique (ASEP process), qui permet d'étudier spécifiquement cet effet. Nous montrons que les mouvements corrélés des monomères confinés dans le pore génèrent un comportement du temps de résidence avec la longueur du polymère qui est qualitativement similaire à ce qui est habituellement interprété comme la présence d'une barrière d'énergie libre dans le processus de translocation, même lorsqu'une telle barrière n'existe pas. Notre modèle réduit fortement le temps de simulation comparé aux simulations de dynamique moléculaire traditionnelles (quelques secondes contre quelques mois pour un système similaire). Cette accélération provient de l'idéalisation des portions du polymère à l'extérieur du pore. Une telle idéalisation est également présente dans les modèles largement utlisés de type Fokker-Planck, mais dans notre cas le comportement de la partie de la chaîne confinée dans le pore est mieux modélisé.Enfin nous réalisons des expériences pour tester l'existence d'un flot électro-osmotique (EOF) à travers le nanopore d'alpha-hémolysine de staphylococcus aureus. Malgré de nombreux travaux ces dernières années, la question de l'EOF à travers l'un des nanopores biologiques les plus utlisés fait toujours débat. Nous montrons qu'un EOF existe à travers l'alpha-hémolysine et qu'il contrôle la fréquence d'entrée et le temps de résidence de molécules neutres (beta-cyclodextrines) dans le nanopore. La force de l'EOF dépend du type de cation en solution. En particulier nous montrons que l'EOF est plus fort en présence de LiCl que de KCl. / Translocation, the passage of a macromolecule through a pore inserted in a membrane, is involved in many biological processes. Examples include the transport of RNA or proteins between cell components, and the infection of a cell by the passage of a viral DNA through the cell membrane. Today translocation is also the basis of technological applications, such as using pores as sensors for fast molecule sequencing or molecular sieves. The comprehension of the translocation process is important both from a fundamental point of view and for the design of new translocation setups for specific uses.In this thesis both experiments and computer simulations are used to investigate some of the most important effects at work during translocation.Coarse-grained computer simulations are used to study qualitatively the influence of an attractive interaction between the pore walls and a translocating polymer. The location of the interaction is shown to influence both the entry frequency and residence time of the polymer in the pore. The entry frequency is greater when the pore entry is attractive. The behaviour of the residence time with the polymer length is qualitatively and quantitatively affected by the location of the interaction within the pore. Nevertheless, regardless of the location of the interaction, a linear increase of the residence time with polymer length occurs when the polymer becomes longer than the pore. This observation is in qualitative agreement with published experimental data.In the case of slow translocation the correlation between the movements of the monomers confined in the pore may be important. This effect has not been considered previously. A new model of polymer translocation, inspired by the asymmetric exclusion process (ASEP), is developped which enables to specifically investigate this effect. The correlated movements of the monomers confined in the pore are shown to give rise to a behaviour of the residence time with polymer length which is qualitatively similar to what is usually interpreted as the presence of a free-energy barrier in the translocation process, even when such barrier is absent. Our model greatly reduces the simulation time compared to traditional molecular dynamics simulations (several seconds versus several months for similar systems). This speed up comes from the idealization of the portions of the polymer outside the pore. Such idealization is also present in the widely used Fokker-Planck models, but in our case the behaviour of the portion of the chain confined in the pore is better modelled.Finally, experiments are performed to probe the existence of an electro-osmotic flow (EOF) through the nanopore of alpha-hemolysin, from staphylococcus aureus. Despite numerous works during past years, the question of EOF through one of the most commonly used biological nanopores is still under debate. An EOF is shown to exist through alpha-hemolysin and to control the entry frequency and residence time of neutral molecules (beta-cyclodextrins) in the nanopore. The strength of the EOF depends on the type of cations in solution. In particular EOF is shown to be stronger in LiCl solution than in KCl solution.

No description available.

Les interactions biologiques sont régies par des mécanismes complexes, qui mêlent différentes échelles, de temps comme de taille. C’est le cas du ribosome, un complexe nucléoprotéique responsable de la traduction de l’ARNm en protéines, et ce faisant, une cible thérapeutique primordiale. Or la taille du ribosome procaryote 70S (2.4 MDa) rend difficile l’applications des techniques classiques de criblage de ligands. Au cours de ma thèse, j’ai exploré la possibilité d’utiliser la RMN du fluor pour caractériser les interactions entre des ligands et le ribosome procaryote. Cette approche a été motivée par l’apport de nouvelles méthodes de détection pouvant coupler la versatilité de la RMN (Résonance Magnétique Nucléaire) avec les propriétés de l’atome de fluor. L’atome 19F se prête parfaitement à la RMN, avec son rapport gyromagnétique proche du proton et son abondance isotopique naturelle de 100%. De plus, le fluor est bio-orthogonal au Vivant. Enfin, les caractéristiques physico-chimiques du fluor sont bien exploitées dans la pharmacopée (un quart des antibiotiques en possèdent un groupement). / Biological interactions are under the control of complex mechanisms, across different scales, in time of in size. It is particularly true for the ribosome, a nucleoprotein responsible for the mRNA translation into proteins, and thus, a primary therapeutic target. The size of the prokaryotic 70S ribosome (2.4 MDa) is a problem for the application of classical ligand screening method. During my thesis, I explored using fluorine NMR to characterize the interaction between ligands and the prokaryotic ribosome. This strategy was motivated by new detection approaches that can combine NMR (Nuclear Magnetic Resonance) versatility with the fluorine atom properties. The 19F atom is perfectly suited for NMR, with its gyromagnetic ratio close to the proton one and its isotopic abundance of 100%. Moreover, the fluorine is absent from natural compounds. Finally, the physicochemical characteristics of fluorine are well exploited in the pharmacopeia (a fourth of all antibiotics has a fluorine moiety).

This work details the attachment of dendrimers to proteins, peptides and single stranded DNA (ssDNA). Dendrimers based on melamine satisfy many of the synthetic demands in the field of bioconjugate chemistry including: monodispersity, synthetic flexibility and scalability. The solution-phase syntheses of both ssDNA-dendrimer and peptide-dendrimer bioconjugates is described, and thorough characterization by matrix-assisted laser desorption ionization/ time-of-flight (MALDI-TOF) mass spectrometry, UV-vis spectroscopy, fluorescence spectroscopy, and polyacrylamide gel electrophoresis is discussed. Non-covalent DNA-dendrimer complexes have been shown to facilitate antisense gene delivery, but are vulnerable to dissociation and subsequent enzymatic degradation within the cell. In an effort to prepare biocompatible antisense agents capable of effectively shielding ssDNA from intracellular nuclease digestion, disulfide-linked ssDNA-dendrimers were prepared and rigorously characterized to rule out the possibility of an electrostatic-based interaction. Hybridization assays were performed to determine if the covalently-attached dendrimer affected the ability of the attached ssDNA strand to anneal with a complementary sequence to form double-stranded DNA (dsDNA)-dendrimers. Results indicate that ssDNA-dendrimer conjugates readily anneal to complementary ssDNA strands either in solution or attached to gold surfaces. Nuclease digestions of conjugates in solution suggested that enzymatic manipulation of dsDNA-dendrimers is possible, offering promise for DNA-based computation and other fields of DNA-nanotechnology. Much larger bioconjugates consisting of dendrimers, proteins and peptides were prepared with the goal of obtaining molecular weights sufficient for enhanced permeability and retention (EPR) in tumors. While the dendrimer provides the advantages of a purely synthetic route for drug delivery, the protein portion of the bioconjugate provides a monodisperse, macromolecular scaffold for the non-covalent self-assembly of the dendrimers. The strategy presented herein is based on the strong interaction between biotin and the 60 kD tetrameric protein streptavidin. Each monomer of streptavidin is capable of binding 1 biotin molecule, thus when biotin functionalized peptide-dendrimers are added to streptavidin they bind to form a cluster of dendrimers, or a megamer. The biotinylated peptides that link the dendrimers to the streptavidin core provide a way to actively target specific cell types for drug delivery. Megamer formation through the addition of tetrameric streptavidin was successful as indicated by MALDI-TOF, UV-vis titration and gel electrophoresis assays.

Metal/organic/indium tin oxide (ITO) structures, including OLEDs, are demonstrated to contain multiple nonvolatile conductance states that can be programmed by the application of an external bias above a certain threshold voltage (Vth). These conductance states are stable and in turn can be probed by the use of a bias lower in value than Vth. The unbiased retention time of states is greater than several weeks, and more than 48,000 write-read-rewrite-read cycles have been performed with minimal degradation. It is found that the programming of a continuum of conductance states is possible, and techniques to do so are outlined. The electrical conductivity of the highest and lowest states can differ by six orders of magnitude. Switching speeds below 50 ns are shown, resulting in an energy requirement of about 100 pJ to switch from one conductance state to another. The memory phenomenon is shown to be influenced by the active layer thickness and anode/surface roughness while temperature dependence is limited. The electrical characteristics of these devices are consistent with metal diffusion or filament phenomena found in metal-insulator-metal structures, suggesting a possible mechanism by which the states are stored.Electroluminescent devices employing several new organic-inorganic lumophore-functionalized macromolecules are presented. In this study, macromolecules incorporating several lumophores covalently bonded to the vertices of a cubical core structure based on Polyhedral Oligomeric Silsesquioxane (POSS) in multiple configurations are implemented as light-emitting centers. The hole-transporting polymer poly(N-vinylcarbazole) (PVK) and electron-transporting additive 2-(4-biphenylyl)-5-(4-tert-butylphenyl)1,3,4-oxadiazole (PBD) are used as a two-part host to enhance the carrier transport in these simple solution-processed single-layer devices. A study of energy transfer in several systems is carried out to understand the requirements needed to create white-light emission from a single macromolecule. A single macromolecule incorporating twenty-one blue and one yellow lumophore is shown to exhibit field-independent stable white-light electroluminescence with Commission Internationale de l'Eclairage (CIE) coordinates of (0.31, 0.37). An external quantum efficiency of 0.55 percent and a maximum brightness of 1600 cd/m2 are attained with simple solution-processed single-layer devices. High solubility and ease of purification give these macromolecule white-light emitters advantages over their small molecule and polymeric type counterparts.

We performed coarse-grained molecular dynamics simulations of fully dissociated polyelectrolytes in the presence of monovalent and multivalent counterions and charged oligomers. The dynamics of these systems has been studied to follow internal dynamics of polyelectrolyte and counterions. The dynamics is presented via the mean square displacement which we obtain from the simulation trajectory and, moreover, via curves of Fluorescence Correlation Spectroscopy (FCS) obtained also from the simulation trajectory. FCS is an experimental technique which can be used to study condensation of counterions on polyelectrolytes. Therefore, we analyze the FCS curves and investigate applicability of various analytical models to fit these FCS curves. The aim of this work is a discussion about the fitting of the FCS curves and presentation of our observations for dynamics of counterions during condensation on the polyelectrolyte.

<p> This project proposal is focused on the development of a novel class of Viscosity Index improvers. A bench-top method for living anionic polymerization has been set up and tested in many aspects. A new difulvene derivative, 1,4-di(6'-6'-methyl-fulvyl)benzene, was synthesized. The living anionic polymer chain was terminated on this difulvene. Cyclopentadiene groups were formed in the middle of the macromolecule chain during the termination. A doubled molecular weight of polystyrene was obtained after the termination. The cyclopentadienes were then changed to other functional groups by Diels-Alder reaction with dimethyl acetylenedicarboxylate, ozonolysis of double bonds, and bromination and reaction with amine. The infrared spectra and nuclear magnetic resonance spectra of the resulting polymers showed new peaks for the new attached groups.</p> / Thesis / Master of Science (MSc)

Yes / Measurement of molecular weight is an integral part of macromolecular and polymer characterization which usually has limitations. Herein, we present the use of a bench-top 80 MHz NMR spectrometer for diffusion-ordered spectroscopy as a practical and rapid approach for the determination of molecular weight/size using a novel solvent and polymer-independent universal calibration. / Royal Society. Grant Number: URF∖R1∖180274. Engineering and Physical Sciences Research Council. Grant Numbers: EP/V037943/1, EP/V007688/1, EP/V036211/1