Date of Award
Honors College Thesis
Chemistry and Biochemistry
Chemistry and Biochemistry
Cancer is one of the leading causes of death for people in the United States. Despite the numerous cancer treatments available, many of these chemotherapy treatments cause negative side effects such as hair and weight loss. With this in mind, in the past 15 years, researchers have discovered a mechanism known as RNA interference, which can be used to control certain genes and their activity. This mechanism is based on the function of siRNA molecules, which can be inserted into cells.(12) These siRNA molecules can be manipulated to target and eliminate specific cancer cells. However, these siRNA molecules are normally associated with inconvenient problems such as instability and a short half-life. These siRNA molecules also face an uphill task of entering these targets cells. In most cases, cellular entryways are highly selective based on the size and type of cargo.(1)(6) With this in mind, researchers have discovered that gold nanoparticles (AuNPs) can be a suitable transport vehicle for siRNA molecules into targeted cells due to their easy synthesis, easy modification for specific pathways, and high biocompatibility with living tissue.(1)(2) Studies have shown that AuNPs can increase the half-life of siRNA molecules by six fold. Before assembling these gold nanoparticles with siRNA, they must be coated with a reagent of branched polyethyleneimine (C12-PEI) chains in order to increase the stability of the gold nanoparticles, thus producing newly-formed AuPEI conjugates. Subsequently, the PEI will bind with siRNA based on their electrostatic interaction, forming a well-dispersed nanoparticle with uniform size and narrow size distribution. (8)
In Fall 2011, the Huang research group encountered several problems with the synthesis of these AuPEI conjugates. These experiments revealed data suggesting that aggregation occurred at certain concentrations of PEI. Experimental data also suggested that aggregation had occurred when loading these AuPEI conjugates with tRNA. With these problems in mind, the Spring 2012 semester of research has been geared towards determining the optimal PEI concentration for the synthesis of AuPEI conjugates. More specifically, the research effort was geared towards exploring how many PEI molecules are bound to each gold nanoparticle. The Huang research group designed a C12PEI solution incorporated with fluorescein isothiocyanate (FITC) in order to gain a closer look into the synthesis of these AuPEI-FITC conjugates. By monitoring the fluorescence of these AuPEI-FITC conjugates, the results could provide information about optimal synthesis conditions. The Huang research group has conducted numerous experiments to study the effects of incubation time, temperature, and DTT (dithiothreitol) concentration on the amount of PEI-FITC on each gold nanoparticle. Based on the experimental results and its statistical analysis, the Huang research group has successfully determined that an estimated 1,000 PEI molecules are bound to each gold nanoparticle. Upon determining PEI density, the goal of the Summer 2012 semester was to optimize the AuPEI-tRNA conjugate assembly system. Based on extensive experimental data and analysis, the optimized conditions for assembly have been established for future research efforts.
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Nosser, Albert J., "The Characterization of AuNP-PEI Conjugates for siRNA Delivery to Cancer Cells" (2013). Honors Theses. 179.