OPTIMIZING THYROID HORMONE RECEPTOR PROBES FOR USE IN FLUORESCENT LABELLING
| Author(s) | Fendt, Stephen | |
| Date Accessioned | 2018-09-27T18:17:46Z | |
| Date Available | 2018-09-27T18:17:46Z | |
| Publication Date | 2018-05 | |
| Abstract | There has been renewed interest in the fluorescent thyroid hormone receptor probe, JZ01, previously developed in the Koh lab. This probe selectively binds to the thyroid hormone receptor, TR, and is useful in labeling and isolating cells involved in various cancers. With cells labelled by a fluorescent probe, researchers can use fluorescence-activated cell separation (FACS) to isolate their cells of interest. The first goal of this project is to resynthesize JZ01 for use in cell isolation. A second goal of this project is to create a more selective labelling technique. It has become apparent that the selectivity of JZ01 between TRα and TRβ subtypes could use improvement. Collaborators began facing issues when using the newly synthesized JZ01 in their studies as they intended to differentiate cells overexpressing TRβ alone. Ideally, this fluorescent label would selectively bind the thyroid receptor β over its α subtype. To block the fluorescent labelling of the thyroid receptor α, CO31 was synthesized. This triiodothyronine (T3) analog was previously created by Cory Ocasio, another Koh Group graduate. His research found that CO31 formed selective interactions with TRα that were ten to twenty-times greater than interactions with the TRβ subtype [9]. Using these molecules in conjunction, CO31 would theoretically form selective interactions with TRα that would allow JZ01 to more selectively label TRβ in cells. With a more selective probe, collaborators would be able to more efficiently use FACS sorting in their studies. The third goal of this project is to create improvements in the thyroid hormone probe itself. JZ01 is a fluorescein analog which presents several issues when working with cells. First, fluorescein is susceptible to photobleaching meaning it is moderately unstable and prone to breakdown [2,4]. Additionally, this analog can prove to be toxic to cells and its hydrophobic qualities hinders its labelling of cells [4]. Therefore, to create an optimal tool for FACS sorting, it is necessary to synthesize analogs that are better taken up by cells, are less toxic at concentrations use to label, and are more structurally stable. One such analog would include a BODIPY label. Using this analog would theoretically improve labeling by reducing the cellular toxicity associated with the fluorescein. Additionally, BODIPY is a more stable fluorescent analog meaning that the final label will show an increase in stability [2,6]. Finally, BODIPY is known for having a high fluorescent quantum yield even in water meaning that this probe will be able to near 100% efficiency, even in water [6]. SF01 was created by making small alterations in the synthesis of JZ01 to more efficiently couple BODIPY in a final step. The same scaffold structure is used meaning that a relatively similar utility should be expected when using this BODIPY coupled probe. | en_US |
| Advisor | John Koh | |
| Program | Biochemistry | |
| URL | http://udspace.udel.edu/handle/19716/23846 | |
| Publisher | University of Delaware | en_US |
| Keywords | Biochemistry, thyroid hormone receptors, fluorescent labelling | en_US |
| Title | OPTIMIZING THYROID HORMONE RECEPTOR PROBES FOR USE IN FLUORESCENT LABELLING | en_US |
| Type | Thesis | en_US |