Year of Publication

2016

Degree Name

Doctor of Philosophy (PhD)

Document Type

Doctoral Dissertation

College

Engineering

Department

Chemical and Materials Engineering

First Advisor

Dr. Brad Berron

Abstract

Cancer metastasis directly accounts for an estimated 90% of all cancer related deaths and is correlated with the presence of malignant cells in systemic circulation. This observed relationship has prompted efforts to develop a fluid biopsy, with the goal of detecting these rare cells in patient peripheral blood as surrogate markers for metastatic disease as a partial replacement or supplement to tissue biopsies. Numerous platforms have been designed, yet these have generally failed to support a reliable fluid biopsy due to poor performance parameters such as low throughput, low purity of enriched antigen positive cells, and insufficiently low detection thresholds to detect poor expressed surface markers of target cell populations. This work describes the development of a rapid cell sorting technology called Antigen Specific Lysis (ASL) based on photo-crosslinked polymer encapsulation to isolate tumor cells in suspension.

In the first study, we characterize the chemical and structural properties of the surface-initiated polymer films formed directly on mammalian cell surfaces. Coated populations are shown to remain highly viable after coating formation. Biomolecular transport is examined though film coatings on cellular substrates using fluorescent, time-resolved confocal microscopy and diffusivity estimates are generated for these materials. In the next study, a lysis-based cell isolation platform is described in which marker positive cells can be specifically coated in a heterogeneous cell suspension. Anionic surfactants lyse virtually 100% of uncoated cells while fully encapsulated cells remain protected, and are then easily collected by centrifugation. We report that purified cells are released from polymeric coatings to yield viable and functional populations. We monitor cell response throughout the isolation process by multiple techniques, and report viability >80% after the sorting process. Lastly, we examine the response of process yield on the level of photoinitiator loading on target populations. Streptavidin-fluorochrome loading was quantitatively assessed on a panel of markers, both epithelial and mesenchymal, on representative model breast and lung cancer cells. We report that ASL is fundamentally capable of achieving 50-60% yield which is promising for fluid biopsy applications. Finally, both EpCAM and metastatic targeting strategies are then compared to covalently biotinylated samples to inform future robust targeting strategies.

Digital Object Identifier (DOI)

http://dx.doi.org/10.13023/ETD.2016.270

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