Evaporative Edge Lithography: A New Method for Assaying the Effect of Lipophilic Drugs on Migration and Outgrowth of Cells over Patterned Surface

Cells sense and respond to topographical cues in their microenvironment that influence growth, development, and migration. Cell migration and outgrowth assays have been used to study cellular movement or changes in cellular morphology and topography. Such assays are promising tools in drug discovery, especially when implemented with high-throughput and high-content imaging systems. These techniques have also been useful for screening and analyzing the effect of different compounds on neurite outgrowth and topography which in turn may lead to the discovery of beneficial targets for regeneration of nervous tissue.;Typically, high-throughput screening of large chemical libraries is employed during the early stages of discovering new drug entities. However, these screening assays do not utilize different topographical surfaces. Many common techniques such as the scratch wound assay are limited in their compatibility with patterned surfaces. Therefore, there is a need to develop novel technologies capable of identifying potentially therapeutic compounds in early stage of drug discovery processes that can regulate cell behaviors and are not limited in their throughput and compatibility with patterned surfaces.;A potentially scalable approach is the "fence" assay in which cells are cultured on topographical surfaces which are partially covered by a removable barrier. Upon removal of the barrier, cells are free to spread and migrate on the freshly uncovered topographies. In this thesis, a novel technique called evaporative edge lithography (EEL) is demonstrated as an approach to miniaturize the fence assay and can be used for high-throughput screening (HTS) in early stages of drug discovery. Furthermore, EEL is a new method to fabricate lipid-based drug delivery microarrays.;Lipid multilayer micro-patterns offer a promising approach to applications such as drug screening and biosensing that require well defined patterns and fluidity. It is shown in this thesis that the factors that govern stability and instability of lipid multilayer nanostructures upon immersion using fluorescence microscopy and observed the following four mechanisms of lipid multilayer instability and strategies are derived to control immersion stability based on these findings: (1) Dissolution by the air/water interface; (2) Disruption by shearing from flowing solution; (3) Spreading at the solid-liquid interface; (4) Diffusion into solution. Based on these studies, a lipid multilayer microarray was developed that is suitable for cell-based assays without detectible cross-contamination by culturing cells on lipid patterns.;It is shown in this thesis that this assay is compatible with poorly soluble lipophilic drug compounds that pose a challenge for HTS microarray assays. EEL was demonstrated for topographically patterned surfaces for screening compounds on adherent cells. Lipophilic compounds including docetaxel and BFA were screened using EEL with cultured HeLa cells to test if migration is affected and can be quantified with this approach. These results indicate that docetaxel and BFA were delivered locally into cells locally from surface supported lipid films and significantly inhibited cellular migration. Subsequently, EEL was used to screen docetaxel on cultured primary olfactory bulb neuronal cells to test the effect on neurite outgrowth.;EEL is a novel approach that allows delivery and subsequent study of the effects of poorly water-soluble drugs on cell migration as well as in vitro screening of different drugs for their effects on cell structures and functions. In addition, this migration assay is a scalable and promising approach for high throughput drug screening microarrays since multiple drug compounds at different dosages can be screened simultaneously on the same surface. This work will advance future studies in developing a portable assay capable of screening lipophilic cancer and neurotropic compounds for topographically-driven cell outgrowth and migration.