Identifier

etd-11152014-131055

Degree

Master of Science in Engineering Science (MSES)

Department

Biological and Agricultural Engineering

Document Type

Thesis

Abstract

This project explored a gene-regulated chemotherapy using a silver nanoparticle (SNP) conjugated with deoxyribozyme (DNAzyme) oligonucleotides which target a mutated gene in select cancer cells, sensitize them to doxorubicin treatment. Light exposure to the SNP-DNAzyme conjugates disengages the oligonucleotides and permits specific cleavage of the Kirsten Rat Sarcomal Oncogene Homolog (K-RAS) mRNA. These conjugates could provide spatiotemporal specificity in killing only those photoexposed cells with the mutant gene. Synthesis, functionalization and characterization of citrate and hydroxypropyl cellulose SNP conjugates confirmed attachment and photolytic release of the thiol-modified 10-23 DNAzyme. Gel electrophoresis was used to demonstrate DNAzyme photoactivation, showing greater K-RAS RNA degradation when disengaged compared to the SNP-tethered form. DNAzyme in the tethered form was also protected from DNAse degradation compared to photolyzed DNAzyme. Characterization of the toxicity and localization of the nanoparticle drug delivery system constructed for the release of a photolabile DNA oligonucleotide was checked within several sets of cells to check for temporal and spatial control efficiency. MTS, alamar blue, and flow cytometry assays were performed to assess cell viability in several sets of cell cultures, including HEK293 and MCF-7 (wildtype K-RAS), SW480 and MDA-MB-231 (mutant K-RAS), and 3T3 (negative control) lines. Following the 5-day experimental protocol involving staggered treatment with SNP-DNAzyme, UV light, and doxorubicin, no cell group showed the intended pattern of necrosis in mutant K-RAS cells without morbidity in controls or partial treatments. Thus further evaluation of K-RAS+/- cells which respond consistently in viability assays is necessary before this strategy can be deemed of potential as a targeted therapeutic.

Date

2014

Document Availability at the Time of Submission

Release the entire work immediately for access worldwide.

Committee Chair

Monroe, Todd

Included in

Engineering Commons

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