Advances in molecular medicine have produced a large amount of information about genes that translate to therapeutic molecules when expressed in living cells. There is an increasing interest in nonviral methods for gene delivery, to address all concerns on non-toxic, easy, and possibly efficient delivery systems. In this paper we introduced a new attractive approach for non-viral transferring of genetic materials on demand. By using lipofectin reagent (1:1 molar ratio of DOPE:DOTMA. DOPE: L-α-doleoyl posphatidylethanolamine; DOTMA: N-[1-(2,3-dideyloxy) propyl]-n,n,n-trimethylammonium chloride), the lipid/DNA complexes (lipoplexes) can be electrostatically adsorbed on the gold microelectrode surface. The resulting lipoplexes molecules can be subsequently removed from the surface by applying -1.0 V (vs. Ag/AgCl) in physiological phosphate buffer medium (pH 7.4). This electrochemically controlled-release process has been extensively examined by gel electrophoresis (GE), electrochemical quartz crystal microbalance (EQCM), infrared spectroscopy (IR), and square wave voltammetry (SWV) techniques. The lipoplex composition has been addressed for efficient gene delivery protocol, based on their different charge ratios. The results from different techniques coincided, as also verified by the repetitive control experiments. This in-vitro electrically - triggered release protocol for genetic material offers the current gene delivery arsenal a new, simple, and non-viral alternative.

Thesis (M.S.)--Massachusetts Institute of Technology, Dept. of Nutrition and Food Science; and, (M.S.)--Massachusetts Institute of Technology, Dept. of Chemical Engineering, 1981. / MICROFICHE COPY AVAILABLE IN ARCHIVES AND SCIENCE / Includes bibliographical references. / by Rajan Sohansingh Bawa. / M.S.

<div>The advent of Patient-Centric therapy demands technologies capable of producing multiple versions of a given product, each tailored for specific segments of the population/individual, but in a time- and cost-effective manner. Prevailing manufacturing methods for oral dosage forms do not easily lend themselves for the transition to the Patient-Centric area. The purpose of this research was to develop a formulation/manufacturing platform technology meeting the flexibility requirements for Patient-Centric formulation and product development for oral dosage forms. The approach is based on the molecular designing and manufacturing of the dosage form. The dosage form consists of a 3D assembly of prefabricated functional modules, each with a specific pharmaceutical performance function. </div><div>The characterization of individual modules showed that solvent casting produced API-loaded HPMC films with homogeneous content distribution. The release profile of 3D assemblies was significantly influenced by the physicochemical properties of single modules. API-loading, thickness, and diameter had a significant effect on the release kinetics. In contrast, the hydrophobicity of the casting substrate did not affect the release kinetics. The initial geometry of the final 3D assembly given by the number of modules and their diameter was proved to have a significant impact on the release kinetics as well. </div><div>The 3D assemblies were used to produce dosage forms with customizable release profiles. Two API-loaded thin HPMC-based films with fast (FRA) and slow (SRB) release rates were produced by the solvent casting method. Accurate dose control (API loading) was accomplished by varying the number of individual modules in the 3D assemblies, whereas control of release kinetics was achieved by combining different ratios FRA and SRB film modules in the assembled dosage form. </div><div>The modular design was also tested for its ability to generate a dosage form of a weak-base API. This part was accomplished using a module containing citric acid (CA) interspaced between weak-base loaded FRA modules. Characterization of the 3D assemblies that were devoid of CA modules showed that the API release rate from modular assemblies containing weekly basic API exhibited strong pH-dependence. The 3D assemblies featuring CA modules in their design exhibited nearly pH-independent release kinetics. </div><div>Electrospinning was used as an enabling technology to produce HPMC-based fibrous films. HPMC films were able to encapsulate a wide variety of APIs with different aqueous solubility. All fibers produced were in the range of a few hundred nanometers to a few microns. X-ray diffraction and differential scanning calorimetry exhibited the amorphous or crystalline state of the API dispersed. Disintegration and release tests showed the fast dissolution of the fibrous system. </div><div><br></div>

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<p> Polyurea microcapsules are micron-scale, hollow polymer spheres commonly used in agriculture to encapsulate pesticides for controlled diffusive release onto target crops. Diffusion of these active materials through a protective polymer wall offers a safer and more effective method of delivery compared to the direct spraying of crops with toxicants. The approach we are taking to control the release rate is to coat pre-formed porous polyurea capsules with a separate release-controlling outer layer. This allows us to separately optimize the load-bearing capsule wall and the release control layer, an approach commonly used in other membrane diffusion systems.</p> <p> Montmorillonite clay incorporation into polymer matrices can reduce membrane permeability by forcing diffusants to take a tortuous path around the stacked silicate sheets. Effective formation of clay-polyurea composites requires the delamination of clay particles into thin sheets with high aspect ratios, and their incorporation into polyurea microcapsules either during interfacial polymerization, or post-polymerization. The net negative surface charge of the silicate sheets should facilitate their initial binding to the cationic polyurea surfaces, as well as subsequent binding of polycations to the clay-coated polyurea capsules to create layer-by-layer (LbL) capsule assemblies with decreasing release rates of internal materials.</p> <p> The main focus of this project is to gain a fundamental understanding of montmorillonite clay and polyurea microcapsules, and the development of a model polyurea composite capsule for release rate analysis. Emphasis will be placed on the reduced permeability of microcapsules coated with clay by LbL assembly post-polymerization, followed by an exploration of further layering with polycations.</p> / Thesis / Master of Science (MSc)

Molecular recognition is an important tool for developing tunable controlled release systems and fabricating biosensors with increased selectivity and sensitivity. The development of polymer-based materials that exploit molecular recognition events such as host-guest complexation, enzyme-substrate and enzyme-inhibitor interactions and nucleic acid hybridization was pursued in this thesis. Using polymers as an anchor for molecular recognition can enhance the affinity, selectivity, and the capacity for immobilization of recognition units, enabling the practical use of affinity-based systems in real applications. To introduce the potential for immobilization while preserving or enhancing the affinity of small molecule recognition units, the affinity of derivatized cyclodextrins for the hydrophobic drug, dexamethasone, was investigated. Cyclodextrins (CDs) are molecules that possess a hydrophilic exterior and a hydrophobic cavity capable of accommodating a wide range of small molecule guests. Analysis of the solubilization capacities, thermodynamic parameters and aggregative potentials of carboxymethyl and hydrazide derivatives of CDs established the dextran-conjugated βCD derivative as an ideal carrier of hydrophobic drugs and the hydrazide βCD derivative as an optimal solubilizer of lipophilic pharmaceuticals, both alone and when incorporated in a polymer-based drug delivery vehicle. To enable non-covalent immobilization and stabilization of biomacromolecular recognition units, a printed layer hydrogel was investigated as a selective diffusion barrier for analyte sensing and enzyme inhibitor recognition. A printable hydrogel platform was developed from an established injectable system composed of aldehyde- and hydrazide-functionalized poly(oligoethylene glycol methacrylate) polymers. The printed layer hydrogel effectively immobilized a wide range of enzymes and protected enzyme activity against time-dependent and protease-induced denaturation, while facilitating the diffusion of small molecules. Furthermore, to demonstrate the potential of the printed film hydrogel immobilization layer to enhance the selectivity of the target, the printable hydrogel platform was used to develop a microarray-based assay for the screening of inhibitors of the model enzyme, β-lactamase. The assay was able to accurately quantify dose-response relationships of a series of established inhibitors, while reducing the required reagent volumes in traditional drug screening campaigns by 95%. Most significantly, the assay demonstrated an ability to discriminate true inhibitors of β-lactamase from a class of non-specific inhibitors called promiscuous aggregating inhibitors. Finally, to enable non-covalent immobilization of DNA recognition units, the printable hydrogel-based microarray was tested for its ability to immobilize DNA recognition sites and promote the detection of DNA hybridization events. A long, concatameric DNA molecule was generated through rolling circle amplification and was used as a sensing material for the detection of a small, fluorophore labeled oligonucleotide. The printable hydrogel was able to effectively entrap the rolling circle amplification product. Properties of the printable hydrogel were investigated for their ability to support the detection of DNA hybridization events. / Thesis / Doctor of Philosophy (PhD) / This thesis describes the development of polymer-based materials that exploit molecular recognition events for drug delivery and biosensing applications. First, cyclodextrins (CDs) are molecules that are capable of binding a wide range of small molecules. A comprehensive analysis of the complexation properties of CD derivatives revealed critical insight regarding their application in polymer-based drug delivery vehicles. Second, a printable hydrogel platform was developed to support the immobilization and activity of biomolecules and establish a biosensing interface that facilitates the diffusion of small molecules but not molecular aggregates. A microarray-based assay was developed by employing the printed hydrogel interface for the screening of inhibitors of the model enzyme, β-lactamase, and the detection of DNA hybridization events.

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